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  • Industrial Chemistry  (9,655)
  • gene expression
  • 1
    Electronic Resource
    Electronic Resource
    The interferon-α regulation of interferon regulatory factor 1 (IRF-1) and IRF-2 has therapeutic implications in carcinoid tumors (2000)
    Springer
    Annals of oncology 11 (2000), S. 707-714 
    Details
    ISSN: 1569-8041
    Keywords: gene expression ; IRF-1 and IRF-2 ; p68 ; PKR ; signal transduction
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract Background:Interferon regulatory factor 1 (IRF-1) has beendemonstrated to possess antiproliferative and tumor suppressor functions, onthe contrary, IRF-2 has been suggested to induce oncogenetic effect in somecell lines, but not evaluated in tumor patients. Patients and methods:In 35 carcinoid tumor patients, expressionsof IRF-1 and IRF-2 were investigated by immunohistochemistry and their valueswere analyzed with clinical treatment response. In carcinoid tumor cell line,Bon1, effects of IFN-α on the expression of both IRF-1 and IRF-2 mRNAs andproteins were determined by Northern blot, RNase protection assays and Westernblot analysis. Results:IFN-α up-regulated the expression of IRF-1 and IRF-2both in vivoand in vitro. In carcinoid tumors, IFN-αtreatment led to a significant increase in the expression of IRF-1 (P〈 0.001) and IRF-2 (P 〈 0.001). Moreover, the IRFs inductionwas correlated with the clinical response of IFN-α treatment, althoughtheir baseline values were not predictive. In addition, expressions of IRF-1and IRF-2 were significantly correlated with the p68kinaseexpression (P = 0.032 and P = 0.0176, respectively) and theexpression of IRF-1 protein was positively correlated with that of IRF-2(r = 0.671,P = 0.0001) tested in the same specimens. Conclusions:IRF-1 as well as IRF-2 have therapeutic implicationsin carcinoid tumors during treatment with interferon-α and IRFs inductionmight be used as indicators of response to treatment with interferon-α.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1008314804492
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  • 2
    Electronic Resource
    Electronic Resource
    Preservation of Gene Expression Ratios Among Multiple Complex cDNAs After PCR Amplification: Application to Differential Gene Expression Studies (2000)
    Springer
    Journal of structural and functional genomics 1 (2000), S. 1-7 
    Details
    ISSN: 1570-0267
    Keywords: cDNA ; PCR cDNA ; TaqMan Analysis ; gene expression ; Pearson's correlation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Comparative gene expression studies are often limited by low availability of tissue and poor quality of extractable mRNA. Collective PCR amplification of minute quantities of mRNA has great potential for overcoming these limitations. However, there remains significant concern about the effects of amplification on the absolute and relative abundance of individual mRNAs that could complicate subsequent gene expression studies. To address this problem, we systematically compared the relative abundance of many specific mRNAs from complex cDNA preparations (from tissue and cultured cells) both before and after amplification by PCR. Our results demonstrated that, as expected, the absolute abundance of different mRNAs in a cDNA library is altered in an unpredictable manner by PCR amplification. However, we found that the concentration ratios of specific mRNAs among different cDNA preparations were routinely well conserved after PCR amplification. Thus, for the purpose of comparative expression studies for specific mRNAs in two (or more) complex cDNAs, PCR-amplified cDNA is equally useful as unamplified cDNA. These results provide a rigorous experimental validation and offer a theoretical treatment to support the utility of PCR amplified cDNA for differential gene expression studies. We conclude that the inherent difficulties in performing differential screening studies such as gene chip and array analyses on limited amounts of biological materials can be overcome by a PCR amplification step without compromising data quality.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1011337409258
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  • 3
    Electronic Resource
    Electronic Resource
    Dexamethasone induced cardiac hypertrophy in newborn rats is accompanied by changes in myosin heavy chain phenotype and gene transcription (2000)
    Springer
    Molecular and cellular biochemistry 209 (2000), S. 165-174 
    Details
    ISSN: 1573-4919
    Keywords: myosin heavy chain ; gene expression ; hypertrophy ; dexamethasone ; promoter function
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract Cardiac hypertrophy has been observed in newborn infants treated with dexamethasone (DEX). This study was undertaken to examine whether DEX-induced hypertrophy in newborn rats is associated with redistribution of cardiac myosin heavy chain (MHC) isoforms and if so, the effects involve transcriptional regulation. Newborn rats were injected with either DEX (1 mg/kg/day; s.c.) or equivalent volume normal saline for 1, 3, 5, 7 or 9 days. Hypertrophy was quantified by heart dry/wet wt ratios, heart/body wt ratios, and total protein content of the myocardium. Changes in the expression of cardiac MHC mRNA were characterized by northern blot and slot blot analyses, using isoform specific probes for a- and β-MHC genes. DEX effect on α-MHC gene transcription was analyzed by transiently transfecting various α-MHC promoter/CAT reporter constructs into primary cultures of cardiac myocytes derived from one day old rat pups. DEX administration into newborn rats produced significant cardiac hypertrophy ranging from 23% at day 1 to 59% at 9 days. The hypertrophy was accompanied by immediate increase (83%) in steady state level of the α-MHC mRNA within one day and a maximum increase (148%) at 7 days of treatment. The steady state level of β-MHC mRNA declined by 25% at day 1 and a maximum decrease of 54% at day 7 of DEX treatment. The changes in MHC mRNA were also reflected in their protein levels as determined by V1 and V3 isozyme analysis. DEX treatment of primary cultures of cardiomyocytes following transfection with a-MHC promoter/CAT reporter constructs resulted in increased CAT expression in a dose dependent manner. The minimum α-MHC gene sequences responding to DEX treatment were located between the -200 to -74-bp region of the gene, resulting in 2-fold and 6-fold activation of CAT reporter after 0.05 and 0.1 mM doses of DEX, respectively. Our data indicate that DEX induced cardiac hypertrophy in newborn rats is accompanied by increased expression of α-MHC and decreased expression of β-MHC. The α-MHC effects are mediated in part through transcriptional mechanisms.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1007128300430
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  • 4
    Electronic Resource
    Electronic Resource
    Mechanisms of transcriptional activation of cAMP-responsive element-binding protein CREB (2000)
    Springer
    Molecular and cellular biochemistry 212 (2000), S. 5-9 
    Details
    ISSN: 1573-4919
    Keywords: transcriptional regulation ; gene expression ; coactivator ; repressor
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract The CREB-CREM transcription factors are the main gene regulatory effectors of the cAMP signaling pathway. The investigations of this family of transcription factors had a profound impact on the understanding of signaling-induced gene transcription. Here we discuss some key aspects of the underlying biology, review transcriptional activation by CREB proteins through transcription cofactors and present novel insights into the context- and position-specific function of CREB on complex genes.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1007111818628
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  • 5
    Electronic Resource
    Electronic Resource
    Upstream regulatory elements in chick heme oxygenase-1 promoter: A study in primary cultures of chick embryo liver cells (2000)
    Springer
    Molecular and cellular biochemistry 209 (2000), S. 17-27 
    Details
    ISSN: 1573-4919
    Keywords: AP-1 ; cobalt chloride ; gene expression ; heme oxygenase ; oxidative stress ; sodium arsenite
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract Previously, chick heme oxygenase-1 (cHO-1) gene was cloned by us and two regions important for induction by sodium arsenite were identified. These two regions were found to contain consensus sequences of an AP-1 (-1580 to -1573) and a MRE/cMyc complex (-52 to -41). In the current study, the roles of these two elements in mediating the sodium arsenite or cobalt chloride dependent induction of cHO-1 were investigated further. DNA binding studies and site-directed mutagenesis studies indicated that both the AP-1 and MRE/cMyc elements are important for the sodium arsenite induction, while cobalt chloride induction involves only the AP-1 element. Electrophoretic mobility shift assays showed that nuclear proteins binding to the AP-1 element was increased by both sodium arsenite or cobalt chloride treatment, whereas the binding of proteins to the MRE/cMyc element showed a high basal expression in untreated cells and the binding activity was only slightly increased by sodium arsenite treatment. Site-directed mutagenesis studies showed that, to completely abolish sodium arsenite induction, both the AP-1 and MRE/cMyc elements must be mutated; mutation of either element alone resulted in only a partial effect. In contrast, a single mutation at AP-1 element was sufficient to reduce the cobalt chloride induction almost completely. The MRE/cMyc complex plays a major role in the basal level expression, and shares some similarities to the upstream stimulatory factor element (USF) identified in the promoter regions of mammalian HO-1 genes and other stress regulated genes. Because sodium arsenite is known to cause oxidative stress and because activation of AP-1 proteins has been shown to be a key step in the oxidative stress response pathway, we also explored the possibility that the induction of the cHO-1 gene by sodium arsenite is mediated through oxidative stress pathway(s) by activation of AP-1 proteins. We found that pretreatment with antioxidants (N-acetyl cysteine or quercetin) reduced the induction of the endogenous cHO-1 message or cHO-1 reporter construct activities induced by sodium arsenite or cobalt chloride. These antioxidants also reduced the protein binding activities to the AP-1 element in the electrophoretic mobility shift assays. In summary, induction of the cHO-1 gene by sodium arsenite or cobalt chloride is mediated by activation of the AP-1 element located at -1,573 to -1,580 of the 5′ UTR.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1007025505842
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  • 6
    Electronic Resource
    Electronic Resource
    Kidney ischemia-reperfusion: Modulation of antioxidant defenses (2000)
    Springer
    Molecular and cellular biochemistry 205 (2000), S. 1-11 
    Details
    ISSN: 1573-4919
    Keywords: kidney ; ischemia-reperfusion injury ; free radicals ; reactive oxygen species ; gene expression ; antioxidant enzymes
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract Reactive oxygen species (ROS; O2-, H2O2, and OH·), normal by-products of cellular metabolic processes, are kept in control by antioxidant enzymes, such as catalase, glutathione peroxidase (GPX) and superoxide dismutases (SODs). To understand the role of antioxidant enzymatic defenses against ROS injury following ischemia-reperfusion, we examined the effect on kidney exposed to varying periods (30, 60 or 90 min) of ischemia followed by different periods of reperfusion. The enzymatic activities and protein levels of catalase, GPX, CuZnSOD and MnSOD were relatively unaffected at 30 min of ischemia followed by 0, 2 or 24 h reperfusion. However, 60 or 90 min of ischemia followed by 0, 2 or 24 h of reperfusion resulted in a decrease in activities and protein levels which paralleled the duration of ischemic injury. MnSOD activity tended to recover towards normal during reperfusion. Examination of the mRNA levels of these antioxidant enzymes demonstrated a severe decrease in mRNA levels of catalase and GPX at a time point of minimal ischemic injury (30 min of ischemia followed by reperfusion) suggesting that loss of mRNA of catalase and GPX may be the first markers of alterations in cellular redox in ischemia-reperfusion injury. Greater loss of mRNA for catalase, GPX and CuZnSOD were observed following longer periods (60 or 90 min) of ischemia. The mRNA for MnSOD was upregulated at all time points of ischemia-reperfusion injury. Actually, the greater decrease in mRNAs for catalase, GPX and CuZnSOD in the acute phase (within 24 h) subsequently showed a further decrease in these enzyme activities in the subacute phase (72 or 120 h after ischemia). These enzyme activities in the 30 min ischemia group, but not in the 90 min group, already showed tendencies for normalization at 120 h after ischemia. To understand the molecular basis of the loss of mRNA of these antioxidant enzymes during ischemia-reperfusion injury, we examined the rate of transcription by nuclear run-on assays. The similar rates of transcription in control and kidney exposed to ischemia-reperfusion indicates that the loss of mRNA for catalase, GPX and CuZnSOD are possibly due to the increased rate of turnover of their mRNAs. These studies suggest that expression of antioxidant genes during ischemia-reperfusion are not coordinately expressed and the differential loss of antioxidant enzymes may be the contributing factor(s) towards the heterogeneous renal tissue damage as a result of ischemia-reperfusion induced oxidative stress.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1007047505107
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  • 7
    Electronic Resource
    Electronic Resource
    Transcriptional regulation of cyclooxygenase-2 in the Human Microvascular Endothelial Cell Line, HMEC-1: Control by the combinatorial actions of AP2, NF-IL-6 and CRE elements (2000)
    Springer
    Molecular and cellular biochemistry 203 (2000), S. 41-51 
    Details
    ISSN: 1573-4919
    Keywords: prostaglandin ; cyclooxygenase ; transcriptional regulation ; gene expression ; promotor activation ; transcription ; endothelial cells
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract Interleukin-1β (IL-1) is a potent inducer of cyclooxygenase-2 (COX-2) and prostaglandin biosynthesis in many types of cells, yet little is known about the molecular mechanisms regulating IL-1 mediated prostanoid biosynthesis in the endothelium of the microvasculature. Therefore, we examined the cis- and trans-acting factors regulating IL-1-induced COX-2 expression in the human microvascular endothelial cell line, HMEC-1. IL-1 enhanced steady state levels of COX-2 protein and mRNA synthesis by ≈ 2-fold which preceded a 2-fold increase in PGFα biosynthesis. Expression of a series of COX-2 promoter-luciferase constructs in IL-1 treated HMEC-1 cells revealed that the 'full length' (-1432/+59 bp) promoter was 10 times more active than the SV-40 promoter/enhancer and that it could be further activated by IL-1. Surprisingly however, all except for the shortest COX-2 promoter construct retained the ability to respond to IL-1 and luciferase activity driven by -191/+59 bp COX-2 promoter was as responsive to IL-1 as the full-length promoter. Moreover, site-directed promoter mutagenesis and electophoretic mobility shift assays (EMSA) indicate that the combinatorial actions of AP2, NF-IL6, and CRE elements are critical for both constitutive and IL-1-inducible COX-2 promoter activity. Understanding the mechanism(s) regulating COX-2 gene expression and prostaglandin biosynthesis in the microvasculature has important implications with regard to inflammation and angiogenesis in vivo.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1007045600664
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  • 8
    Electronic Resource
    Electronic Resource
    Characterization of a 5′-flanking region supporting the transcription of mouse thymosin β-4 in mouse NIH3T3 cells (2000)
    Springer
    Molecular and cellular biochemistry 203 (2000), S. 163-167 
    Details
    ISSN: 1573-4919
    Keywords: thymosin β-4 ; gene expression ; chloramphenicol acetyltransferase ; NIH3T3 cells ; interferon response
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract Expression of the gene coding for thymosin β-4 (Tβ-4), the major G-actin sequestering peptide in the cell, is regulated mainly at the level of transcription. In this study, we examined the nucleotide sequence of the 5′-flanking region (from - 2202 to - 881) of the mouse Tβ-4 gene, and demonstrated that the DNA fragment from -278 to +410 of this gene was capable of directing the expression of a chloramphenicol acetyltransferase reporter gene in NIH3T3 cells. However, expression of the reporter gene in cells cannot be induced by interferon-a treatment even though a rapid activation of endogenous Tβ-4 gene by this cytokine was observed. These results suggest that the projected interferon-stimulated response element (ISRE) might reside in other parts of the mouse Tβ-4 gene.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1007020619788
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  • 9
    Electronic Resource
    Electronic Resource
    The effect of thioacetamide on the activity and expression of cytosolic rat liver glutathione-S-transferase (2000)
    Springer
    Molecular and cellular biochemistry 211 (2000), S. 103-110 
    Details
    ISSN: 1573-4919
    Keywords: thioacetamide ; glutathione-S-transferase ; rat liver ; transcription ; gene expression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract The effect of thioacetamide (TA), an hepatotoxic and hepatocarcinogenic compound, on the expression and activity of the cytosolic enzyme glutathione-S-transferase (GST) was studied in rat liver. Four h following the administration of 14C-labeled thioacetamide (10 mg/Kg), several subunits of GST were found to be radioactively labeled. A single sublethal dose of TA (250 mg/Kg) decreased by three-fold the expression of classα GST at 24-48 h of treatment, but did not significantly affect the transcription of class μ GST. The activity of the enzyme toward 1-chloro-2,4-dinitrobenzene was mildly inhibited (66% of the control) by a 24 h TA treatment and gradually increased thereafter. It is proposed that the covalent binding of TA or its derivative to the GST subunits does not affect the activity of the enzyme. Nevertheless, GST activity inhibition is due to the deleterious effect of TA on GST transcription.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1007114801362
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  • 10
    Electronic Resource
    Electronic Resource
    Apolipoprotein E gene expression is reduced in apolipoprotein A-I transgenic mice (2000)
    Springer
    Molecular and cellular biochemistry 209 (2000), S. 125-129 
    Details
    ISSN: 1573-4919
    Keywords: apolipoprotein E ; apolipoprotein A-I ; gene expression ; transgenic mouse
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract The levels of plasma apolipoprotein (apo) E, an anti-atherogenic protein involved in mammalian cholesterol transport, were found to be 2-3 fold lower in mice over-expressing human apoA-I gene. ApoE is mainly associated with VLDL and HDL-size particles, but in mice the majority of the apoE is associated with the HDL particles. Over-expression of the human apoA-I in mice increases the levels of human apoA-I-rich HDL particles by displacing mouse apoA-I from HDL. This results in lowering of plasma levels of mouse apoA-I. Since plasma levels of apoE also decreased in the apoA-I transgenic mice, the mechanism of apoE lowering was investigated. Although plasma levels of apoE decreased by 2-3 fold, apoB levels remained unchanged. As expected, the plasma levels of human apoA-I were almost 5-fold higher in the apoAI-Tg mice compared to mouse apoA-I in WT mice. If the over-expression of human apoA-I caused displacement of apoE from the HDL, the levels of hepatic apoE mRNA should remain the same in WT and the apoAI-Tg mice. However, the measurements of apoE mRNA in the liver showed 3-fold decreases of apoE mRNA in apoAI-Tg mice as compared to WT mice, suggesting that the decreased apoE mRNA expression, but not the displacement of the apoE from HDL, resulted in the lowering of plasma apoE in apoAI-Tg mice. As expected, the levels of hepatic apoA-I mRNA (transgene) were 5-fold higher in the apoAI-Tg mice. ApoE synthesis measured in hepatocytes also showed lower synthesis of apoE in the apoAI-Tg mice. These studies suggest that the integration of human apoA-I transgene in mouse genome occurred at a site that affected apoE gene expression. Identification of this locus may provide further understanding of the apoE gene expression.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1007107712725
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  • 11
    Electronic Resource
    Electronic Resource
    CRE-decoy oligonucleotide-inhibition of gene expression and tumor growth (2000)
    Springer
    Molecular and cellular biochemistry 212 (2000), S. 29-34 
    Details
    ISSN: 1573-4919
    Keywords: cAMP ; transcription factor-decoy oligonucleotides ; CRE ; Ap-1 ; p53 ; tumor growth ; gene expression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract Nucleic acid molecules with high affinities for a target transcription factor can be introduced into cells as decoy cis-elements to bind these factors and alter gene expression. This review discusses a synthetic single-stranded palindromic oligonucleotide, which self-hybridizes to form a duplex/hairpin and competes with cAMP response element (CRE) enhancers for binding transcription factors. This oligonucleotide inhibits CRE- and Ap-1-directed gene transcription and promotes growth inhibition in vitro and in vivo in a broad spectrum of cancer cells, without adversely affecting normal cell growth. Evidence presented here suggests that the CRE-decoy oligonucleotide can provide a powerful new means of combating cancers, viral diseases, and other pathological conditions by regulating the expression of cAMP-responsive genes.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1007144618589
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  • 12
    Electronic Resource
    Electronic Resource
    Catecholamines and angiotensinogen gene expression in kidney proximal tubular cells (2000)
    Springer
    Molecular and cellular biochemistry 212 (2000), S. 73-79 
    Details
    ISSN: 1573-4919
    Keywords: adrenergic receptors ; renin-angiotensin system (RAS) ; gene expression ; kidney
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract To investigate the molecular mechanism(s) of action of catecholamines on the expression of the angiotensinogen (ANG) gene in kidney proximal tubular cells, we used opossum kidney (OK) cells with a fusion gene containing the 5′-flanking regulatory sequence of the rat ANG gene fused with a human growth hormone (hGH) gene as a reporter, pOGH (rANG N-1498/+18), permanently integrated into their genomes. The level of expression of the ANG-GH fusion gene was quantified by the amount of immunoreactive-hGH (IR-hGH) secreted into the medium. The addition of norepinephrine (NE), isoproterenol (a β1/β2-adrenergic receptor (AR) agonist) and iodoclonidine (an α2-AR agonist) stimulated the expression of the ANG-GH fusion gene in a dose-dependent manner, whereas the addition of epinephrine and phenylephrine (α1-AR agonist) had no effect. The stimulatory effect of NE was blocked by the presence of propranolol (β-AR blocker), atenolol (β1-AR blocker), yohimbine (α2-AR blocker), Rp-cAMP (an inhibitor of cAMP-dependent protein kinase AI & AII) and staurosporine (an inhibitor of protein kinase C), but was not blocked by ICI 118, 551 (β2-AR blocker) and prazosin (α1-AR blocker). The addition of a combination of isoproterenol and iodoclonidine or a combination of 8-Bromo-cAMP (8-Br-cAMP) and phorbol 12-myristate (PMA) synergistically stimulated the expression of the ANG-GH fusion gene as compared to the addition of isoproterenol, iodoclonidine, 8-Br-cAMP or PMA alone. Furthermore, the addition of NE, 8-Br-cAMP or PMA stimulated the expression of pOGH (rANG N-806/-779/-53/+18), a fusion gene containing the putative cAMP responsive element (CRE, ANG N-806/-779) upstream of the ANG promoter (ANG N-53/+18) in OK cells, but had no effect on the expression of fusion genes containing the mutant of the CRE. Gel mobility shift assays revealed that the ANG-CRE binds with the DNA-binding domain (bZIP 254-327) of the cAMP-responsive binding protein (CREB). The binding of the labeled ANG-CRE to CREB (bZIP254-327) was displaced by unlabeled ANG-CRE and the CRE of the somatostatin gene but not by the mutants of the ANG-CRE. Finally, NE stimulated the phosphorylation of CREB in OK cells. These studies demonstrate that the molecular mechanism(s) of NE action on the expression of the ANG gene in OK cells may be mediated via both the PKA and PKC signalling pathways and via the phosphorylation of CREB. The phosphorylated CREB then interacts with the CRE in the 5′-flanking region of the ANG gene and subsequently stimulates the gene expression.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1007152821315
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  • 13
    Electronic Resource
    Electronic Resource
    Control of cardiomyocyte gene expression as drug target (2000)
    Springer
    Molecular and cellular biochemistry 212 (2000), S. 135-142 
    Details
    ISSN: 1573-4919
    Keywords: gene expression ; catecholamines ; angiotensin II ; heart failure ; myosin ; hypertension ; eprosartan
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract Pressure overload of the heart is associated with a perturbed gene expression of the cardiomyocyte leading to an impaired pump function. The ensuing neuro-endocrine activation results in disordered influences of angiotensin II and catecholamines on gene expression. To assess whether angiotensin II type 1 receptor inhibition can also counteract a raised sympathetic nervous system activity, spontaneously hypertensive rats fed a hypercaloric diet were treated with eprosartan (daily 90 mg/kg body wt) and cardiovascular parameters were monitored with implanted radiotelemetry pressure transducers. Both, blood pressure and heart rate were increased (p 〈 0.05) by the hypercaloric diet. Although eprosartan reduced (p 〈 0.05) the raised systolic and diastolic blood pressure, the diet-induced rise in heart rate was blunted only partially. In addition to drugs interfering with the enhanced catecholamine influence, compounds should be considered that selectively affect cardiomyocyte gene expression via 'metabolic' signals.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1007181626766
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  • 14
    Electronic Resource
    Electronic Resource
    Expression of renin-angiotensin system and extracellular matrix genes in cardiovascular cells and its regulation through AT1 receptor (2000)
    Springer
    Molecular and cellular biochemistry 212 (2000), S. 203-209 
    Details
    ISSN: 1573-4919
    Keywords: angiotensinogen ; fibronectin ; gene expression ; transcriptional regulation ; cardiomyocytes ; vascular smooth muscle cells
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract Angiotensinogen (AGT) is a unique substrate of the renin-angiotensin system and fibronectin (FN) is an important component of the extracellular matrix. These play critical roles in the pathophysiological changes including cardiovascular remodeling and hypertrophy in response to hypertension. This study was performed to examine the regulation of AGT and FN gene in cardiac myocytes (CMs) and vascular smooth muscle cells (VSMCs) in response to mechanical stretch. Mechanical stretch significantly increased the AGT mRNA expression in CMs, while these stimuli did not affect FN mRNA levels. On the other hand, Mechanical stretch upregulated FN mRNA levels in VSMCs, whereas no increase in AGT mRNA levels was observed in response to stretch stimuli. An angiotensin II type 1 (AT1) receptor antagonist (CV11974) significantly decreased these stretch-mediated increases in mRNA level and promoter activity of the AGT and FN gene, whereas angiotensin II type 2 (AT2) receptor antagonist (PD123319) did not affect the induction. These results indicate that mechanical stretch activates transcription of the AGT and FN gene mainly via AT1 receptor-pathway in CMs and VSMCs. Furthermore, mechanisms regulating AGT and FN gene seem to be different between CMs and VSMCs.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1007141912654
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  • 15
    Electronic Resource
    Electronic Resource
    Regulation of angiotensin II receptors in the medullary thick ascending limb (2000)
    Springer
    Molecular and cellular biochemistry 212 (2000), S. 211-217 
    Details
    ISSN: 1573-4919
    Keywords: angiotensin receptor ; medullary thick ascending limb ; sodium intake ; primary cell culture ; gene expression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract Angiotensin II (Ang II) is an important regulator of the function of medullary thick ascending limb of loop of Henle (MTAL). Recent studies showed that changes in Ang II receptor expression occur and underlie changes in the function of proximal tubules during altered sodium intake. The present experiment was designed to determine (1) whether expression of the type 1 Ang II (AT1) receptor in the MTAL is regulated by altered sodium intake, and (2) the specific pathway(s) mediating sodium-induced AT1 expression in the MTAL. Wistar rats were fed a normal sodium (0.5%, NS), low sodium (0.07%, LS), or high sodium (4%, HS) diet for 2 weeks. Northern blot analysis and radioligand binding showed that in rats fed a normal sodium diet the rank of order for both AT1 mRNA expression and receptor density was outer medulla 〉 cortex 〉 inner medulla. Sodium restriction significantly increased both AT1 mRNA expression and receptor density in the outer medulla. In contrast, neither AT1 mRNA expression nor receptor density in the outer medulla was altered by sodium loading. Losartan treatment (3 mg/kg/per day by oral gavage for 2 weeks) prevented low sodium-induced upregulation of the AT1 receptor in the outer medulla, but it had no effect on AT1 expression in the outer medulla of rats fed a normal sodium diet. Highly purified suspensions of MTAL were isolated from rats fed a normal or low sodium diet. Low sodium intake significantly increased AT1 mRNA level by 184% and AT1 receptor density by 58% in MTALs. Primary cultures of MTAL cells were treated with PBS, Ang II (10-8 M), and Ang II + 17 octadecynoic (17 ODYA, 10 μM). Ang II caused about 2-fold increase in AT1 mRNA levels, and this increase was diminished by about 30% by the addition of 17 ODYA. We conclude that (1) sodium restriction but not sodium loading increases AT1 receptor expression in the MTAL, (2) low sodium-induced upregulation of the AT1 receptor in the MTAL is Ang II-dependent, and (3) Ang II-induced upregulation of the AT1 receptor in the MTAL is mediated, at least in part, by cytochrome P450 pathways.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1007193931309
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  • 16
    Electronic Resource
    Electronic Resource
    Role of cardiac renin-angiotensin system in sarcoplasmic reticulum function and gene expression in the ischemic-reperfused heart (2000)
    Springer
    Molecular and cellular biochemistry 212 (2000), S. 227-235 
    Details
    ISSN: 1573-4919
    Keywords: renin angiotensin system ; sarcoplasmic reticulum ; Ca2+-handling ; gene expression ; ischemia-reperfusion ; cardioprotection
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract The aim of this study was to explore the possible participation of cardiac renin-angiotensin system (RAS) in the ischemia-reperfusion induced changes in heart function as well as Ca2+-handling activities and gene expression of cardiac sarcoplasmic reticulum (SR) proteins. The isolated rat hearts, treated for 10 min without and with 30 μM captopril or 100 μM losartan, were subjected to 30 min ischemia followed by reperfusion for 60 min and processed for the measurement of SR function and gene expression. Attenuated recovery of the left ventricular developed pressure (LVDP) upon reperfusion of the ischemic heart was accompanied by a marked reduction in SR Ca2+-pump ATPase, Ca2+-uptake and Ca2+-release activities. Northern blot analysis revealed that mRNA levels for SR Ca2+-handling proteins such as Ca2+-pump ATPase (SERCA2a), ryanodine receptor, calsequestrin and phospholamban were decreased in the ischemia-reperfused heart as compared with the non-ischemic control. Treatment with captopril improved the recovery of LVDP as well as SR Ca2+-pump ATPase and Ca2+-uptake activities in the postischemic hearts but had no effect on changes in Ca2+-release activity due to ischemic-reperfusion. Losartan neither affected the changes in contractile function nor modified alterations in SR Ca2+-handling activities. The ischemia-reperfusion induced decrease in mRNA levels for SR Ca2+-handling proteins were not affected by treatment with captopril or losartan. The results suggest that the improvement of cardiac function in the ischemic-reperfused heart by captopril is associated with the preservation of SR Ca2+-pump activities; however, it is unlikely that this action of captopril is mediated through the modification of cardiac RAS. Furthermore, cardiac RAS does not appear to contribute towards the ischemia-reperfusion induced changes in gene expression for SR Ca2+-handling proteins.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1007174803993
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  • 17
    Electronic Resource
    Electronic Resource
    Attenuation of changes in sarcoplasmic reticular gene expression in cardiac hypertrophy by propranolol and verapamil (2000)
    Springer
    Molecular and cellular biochemistry 213 (2000), S. 111-118 
    Details
    ISSN: 1573-4919
    Keywords: pressure overload ; gene expression ; subcellular remodeling ; sarcoplasmic reticulum Ca2+-handling ; anti-hypertensive therapy
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract The effects of propranolol and verapamil on contractile dysfunction, subcellular remodeling and changes in gene expression in cardiac hypertrophy due to pressure overload were examined. Rats were subjected to banding of the abdominal aorta and then treated with either propranolol (10 mg/kg daily), verapamil (5 mg/kg daily) or vehicle for 8 weeks after the surgery. Depression of the left ventricular function in the hypertrophied heart was associated with decreases in myofibrillar and myosin CA2+ ATPase activities as well as Ca2+-pump and Ca2+-release activities of the sarcoplasmic reticulum (SR). The level of a-myosin heavy chain (α-MHC) mRNA was decreased while that of β-MHC mRNA was increased in the pressure-overloaded heart. The level of SR Ca2+-pump ATPase (SERCA2) mRNA and protein content for SERCA2 were decreased in the pressure overloaded heart. Treatment of the hypertrophied animals with propranolol or verapamil resulted in preservation of the left ventricular function and prevention of the subcellular alterations. Shift in the α- and β-MHC mRNA levels and changes in the expression in SERCA2 mRNA level and protein content were also attenuated by these treatments. The results suggest that blockade of β-adrenoceptors or voltage-dependent calcium channels normalizes the cardiac gene expression, prevents subcellular remodeling and thus attenuates heart dysfunction in rats with cardiac hypertrophy. Furthermore, both cardiac β-adrenoceptors and L-type Ca2+-channels may be involved in the genesis of cardiac hypertrophy due to pressure overload.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1007120332587
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  • 18
    Electronic Resource
    Electronic Resource
    Cloning and characterization of the rat TIS11 gene (2000)
    Springer
    Molecular and cellular biochemistry 213 (2000), S. 119-126 
    Details
    ISSN: 1573-4919
    Keywords: TIS11 ; an immediate early gene ; gene cloning ; gene expression ; gene organization ; promoter
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract The TIS11 gene is an immediate early gene that is induced rapidly and transiently by phorbol 12-myristate 13-acetate and various growth factors. To study transcriptional regulation of the gene, a genomic clone of rat TIS11 was isolated, and the organization of exon-intron structure and transcriptional initiation site were determined. The rat TIS11 gene consisted of 2 exons spanning approximately 2.5 kb. Several canonical sequences for binding of transcriptional factors were found in the 5′-flanking region. The 5.3 kb of the 5′-flanking region fused to a luciferase reporter gene showed promoter activity when introduced into rat pheochromocytoma PC12 cells. Analyses with serial 5′-deletion mutants suggested that the major positive regulatory region is located at the region of -241 to -76, and that the minimum promoter region is within the 76-bp upstream of the transcriptional initiation site. Gel mobility shift assays revealed that PC12 cell nuclear proteins specifically bind to the major positive regulatory region of the TIS11 gene. The identified nuclear protein components may act as the positive trans-acting factors in the basal expression of the TIS11 gene in PC12 cells.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1007172316657
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  • 19
    Electronic Resource
    Electronic Resource
    Expression and activity of antioxidant enzymes during potato tuber dormancy (2000)
    Springer
    Potato research 43 (2000), S. 383-393 
    Details
    ISSN: 1871-4528
    Keywords: Solanum tuberosum L. ; plant development ; antioxidant genes ; gene expression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Summary The expression of antioxidant genes has been analyzed in a potato plant and during tuber dormancy. Manganese superoxide dismutase (MnSOD), cytosolic copper and zinc superoide dismutase (Cu/ZnSOD), catalase class II, cytosolic ascorbate peroxidase (APX) and glutathione peroxidase (GPX) are expressed at the RNA level in all the contexts analyzed. By contrast, the expression of the iron superoxide dismutase (FeSOD) and plastidic Cu/ZnSOD seems to be limited to green tissues, as shown by northern blots and native gels. A complex DAB-peroxidase isozyme pattern (using diaminobenzidine as substrate) has been observed in different developmental contexts analyzed, but hardly observed in tubers. During tuber dormancy, MnSOD and cytosolic Cu/ZnSOD activity was relatively constant in both Désirée and Bintje varieties while catalase activity decreases. Moreover, tuber dormancy breakage did not involve significant changes in the activity of these enzymes. On the basis of these results, the possible link between active oxygen species (AOS) metabolism and dormancy is discussed.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF02360542
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  • 20
    Electronic Resource
    Electronic Resource
    Functional genomic analysis of potato tuber life-cycle (2000)
    Springer
    Potato research 43 (2000), S. 297-312 
    Details
    ISSN: 1871-4528
    Keywords: gene expression ; cDNA-AFLP ; RNA-fingerprinting ; organogenesis ; tuberisation ; dormancy ; sprouting ; cluster analysis ; metabolic pathways
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Summary Potato tuber life-cycle is composed of many individual developmental stages including tuber formation, tuber development, dormancy and sprouting. We have used cDNA-AFLP fingerprinting to analyse gene expression in 24 individual stages of development, over the period from stolon formation through sprouting. In addition to these developmental stages, different tissues were analysed to assess tissue specificity and various controls were incorporated to determine process specificity. In total around 18000 transcript derived cDNA fragments (TDFs) were visualised from which circa 2600 were included in a statistical analysis allowing general conclusions about gene expression during development. More than 200 process specific TDFs were isolated and sequenced throughout the potato tuber life-cycle. The sequence similarities of these TDFs to known genes give an insight into the kinds of processes occurring during tuberisation, dormancy and sprouting.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF02360536
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  • 21
    Electronic Resource
    Electronic Resource
    Cu, Zn superoxide dismutase deficiencyaccelerates the time course of an age-related markerin Drosophila melanogaster (2000)
    Springer
    Biogerontology 1 (2000), S. 163-169 
    Details
    ISSN: 1573-6768
    Keywords: aging ; Cu, Zn superoxide dismutase ; Drosophila melanogaster ; gene expression ; wingless
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract In the oxidative stress hypothesis of aging therandom accumulation of oxidative damage over time ispostulated to cause aging. The pace at whichoxidative damage accrues determines the rate of aging,but it is less clear how the accumulation of randomdamage could cause the stereotypic pattern of aging. It has been proposed that oxidative damage induceschanges in gene expression, translating a random inputof damage into a patterned output. In support of thiswe show that in adult Drosophila melanogaster,with a deficiency in the anti-oxidant enzyme Cu, Znsuperoxide dismutase (Sod), an increase in oxidativestress, and a shortened life span, there isacceleration in the normal age-related temporalpattern of wingless gene expression. Theacceleration in the temporal pattern of winglessgene expression is proportional to the shortened lifespan suggesting that the shortened life span of Soddeficient animals is due, not to an abnormalpathological process, but to an increase in the rateof aging.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1010039813107
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  • 22
    Electronic Resource
    Electronic Resource
    Regulation of NADPH-cytochrome P450 reductase expressed during Douglas-fir germination and seedling development (2000)
    Springer
    Plant molecular biology 44 (2000), S. 141-153 
    Details
    ISSN: 1573-5028
    Keywords: gene expression ; germination ; NADPH-cytochrome P450 reductase ; Pseudotsuga menziesii
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract NADH-cytochrome P450 is a key enzyme that transfers electrons from NADPH to the cytochrome P450 family of enzymes. To begin to determine the regulation of CPR gene expression and enzyme activity in Douglas-fir a full-length cDNA was isolated from a seedling λZAP cDNA library and the ORF was used to develop a synthetic CPR-peptide-based antiserum. Northern blot analysis indicated CPR expression was regulated both developmentally prior to seed maturation and during germination, and differentially in the cotyledons, radicle and megagametophyte of seed and seedling tissues. The CPR-peptide antiserum detected a single CPR in seed and seedling microsomes analyzed by western blot of two-dimensional SDS-polyacrylamide gels. In microsomal extracts from whole seeds and seedlings, the amount of CPR protein remained constant while NADPH:cytochrome c reductase activity increased during stratification, germination and early seedling development. In contrast to cotyledons and megagametophyte, the level of CPR protein detected in radicles was higher than expected when compared to the amount of CPR transcript.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006425025702
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  • 23
    Electronic Resource
    Electronic Resource
    The nitrate reductase promoter of birch directs differential reporter gene expression in tissues of transgenic tobacco (2000)
    Springer
    Plant and soil 221 (2000), S. 33-38 
    Details
    ISSN: 1573-5036
    Keywords: ammonium ; birch ; gene expression ; nia promoter ; nitrate ; nitrate reductase
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract A 1535 bp promoter of the nitrate reductase gene (nia) from birch (Betula pendula) and a series of 5′ deletions were fused to the β-glucuronidase (GUS) gene and introduced into Nicotiana plumbaginifolia. In transgenic plants the NR promoter sequences directed strong GUS expression in the root epidermal hair cells, and in phloem cells of leaf and stem vascular tissue. The NR promoter confers also a significant stimulation of the GUS gene expression by nitrate. These findings might indicate that nitrate flow is one of the signals involved into tissue and cell specific expression of the NR promoter GUS fusions.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1004739621352
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  • 24
    Electronic Resource
    Electronic Resource
    Molecular approaches to study plasma membrane H+-ATPases in arbuscular mycorrhizas (2000)
    Springer
    Plant and soil 226 (2000), S. 219-225 
    Details
    ISSN: 1573-5036
    Keywords: arbuscular mycorrhizas ; gene expression ; Glomus mosseae ; nutrient transport processes ; plasma membrane H+-ATPases
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract The activity of H+-ATPases of plant and fungi generates an electrochemical gradient of H+ across the cell plasma membrane that drives a number of secondary transport systems, including those responsible for the translocation of cations, anions, amino acids and sugars. During the last years, several studies have been aimed at elucidating the role of plasma membrane H+-ATPases in the nutrient exchange processes taking place between the plant and the fungus in arbuscular mycorrhizal (AM) symbiosis. This paper reviews present knowledge about plasma membrane H+-ATPases and experimental evidence supporting the involvement of H+-ATPases of both organisms in the bidirectional transport of nutrients between partners. Molecular strategies that will provide further information on the function and regulation of plasma membrane H+-ATPases in AM symbiosis are presented and discussed.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1026469109773
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  • 25
    Electronic Resource
    Electronic Resource
    Plant responses to ultraviolet-B (UV-B: 280–320 nm) stress: What are the key regulators? (2000)
    Springer
    Plant growth regulation 32 (2000), S. 27-39 
    Details
    ISSN: 1573-5087
    Keywords: ethylene ; gene expression ; jasmonic acid ; reactive oxygen species ; salicylic acid ; ultraviolet-B radiation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract Depletion of the stratospheric ozone layer is leadingto an increase in ultraviolet-B (UV-B: 280–320 nm)radiation reaching the earth's surface. This hasraised interest in the possible consequence ofincreased UV-B levels on plant growth and developmentand the mechanisms underlying these responses. Although the effects of UV-B are now wellcharacterised at the physiological level, little isknown about the cellular and molecular mechanismsinvolved. Recent studies have shown that UV-B affectsa number of important physiological processes, such asphotosynthesis, through effects on gene expression. In addition, induction of a number of defensemechanisms, such as production of UV-B screeningpigments, increase in antioxidant enzymes andinduction of pathogenesis-related proteins, are alsomediated at the level of gene expression. The signaltransduction pathways by which UV-B regulates geneexpression are at present poorly understood. Thestudies carried out to date have, however, indicateda pivotal role for reactive oxygen species as keysecond messengers acting up-stream of a number ofpathways involving the plant hormones salicylic acid,jasmonic acid and ethylene. The transduction pathwaysidentified to date and the role of intermediates inregulating tolerance to UV-B damage are discussed inthis review.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006314001430
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  • 26
    Electronic Resource
    Electronic Resource
    Biochemical and molecular aspects of fruitlet abscission (2000)
    Springer
    Plant growth regulation 31 (2000), S. 35-42 
    Details
    ISSN: 1573-5087
    Keywords: β-1,4-endoglucanase ; ethylene ; fruit ; gene expression ; polygalacturonase
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract Fruitlet abscission during fruit development is due to the activation ofpre-differentiated abscission zones (AZs) located between twig andpedicel, and/or pedicel and pericarp. Major advances on biochemicaland molecular aspects are related to β-1,4-endoglucanase (EG) andpolygalacturonase (PG), two cell hydrolases involved in the cell walldisassemblement responsible for fruit shedding. AZ activation isaccompanied by an increase in activity and transcript accumulation ofone or both enzymes. Expression of PG genes specifically related toabscission has been found in tomato flower AZ. In peach, an EG genehighly expressed in leaf and fruitlet AZs has been isolated. AZactivation is preceded by an induction of ethylene biosynthesis,paralleled by a stimulation of ACO activity and transcript accumulation.Ethylene, besides a dramatic stimulation of PG and EG, up or downregulates several other abscission related genes. The specificexpression of genes encoding for ethylene receptors in the AZ wouldsupport the hypothesis that fruitlet AZ specificity may depend on theability of this region to sense ethylene.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006338210977
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  • 27
    Electronic Resource
    Electronic Resource
    Molecular cloning and expression analysis of the mevalonate kinase gene from Arabidopsis thaliana (2000)
    Springer
    Plant molecular biology 42 (2000), S. 365-376 
    Details
    ISSN: 1573-5028
    Keywords: Arabidopsis thaliana ; β-glucuronidase (GUS) reporter gene ; gene expression ; isoprenoids ; mevalonate kinase ; transgenic plants
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Mevalonate kinase (MVK), the enzyme that catalyzes the phosphorylation of mevalonate to produce mevalonate 5-phosphate, is considered as a potential regulatory enzyme of the isoprenoid biosynthetic pathway. The Arabidopsis thaliana MVK gene corresponding to the MVK cDNA previously isolated has been cloned and characterized. RNAse protection analysis indicated that the expression of the MVK gene generates three mRNA populations with 5′ ends mapping 203, 254 and 355 nt upstream of the MVK ATG start codon. Northern blot analysis showed that the MVK mRNA accumulates preferentially in roots and inflorescences. Histochemical analysis, with transgenic A. thaliana plants containing a translational fusion of a 1.8 kb fragment of the 5′ region of the MVK gene to the β-glucuronidase (GUS) reporter gene, indicated that the MVK 5′-flanking region directs widespread expression of the GUS gene throughout development, although the highest levels of GUS activity are detected in roots (meristematic region) and flowers (sepals, petals, anthers, style and stigmatic papillae). The expression pattern of the MVK gene suggests that the role of the encoded MVK is the production of a general pool of mevalonate-5-phosphate for the synthesis of different classes of isoprenoids involved in both basic and specialized plant cell functions. Functional promoter deletion analysis in transfected A. thaliana protoplasts indicated that regulatory elements between positions −295 and −194 of the MVK 5′-flanking region are crucial for high-level MVK gene expression.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006325630792
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  • 28
    Electronic Resource
    Electronic Resource
    Isolation and characterization of two pathogen- and salicylic acid-induced genes encoding WRKY DNA-binding proteins from tobacco (2000)
    Springer
    Plant molecular biology 42 (2000), S. 387-396 
    Details
    ISSN: 1573-5028
    Keywords: gene expression ; hypersensitive responses ; plant defense responses ; salicylic acid ; tobacco mosaic virus ; WRKY DNA-binding proteins
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract A pathogen- and salicylic acid (SA)-induced DNA-binding activity has been recently identified in tobacco that is related to a previously identified class of WRKY DNA-binding proteins. To identify members of the WRKY gene family associated with this DNA-binding activity, we have attempted to isolate those WRKY genes that are induced by pathogen infection. Using a domain-specific differential display procedure, we have isolated two tobacco WRKY genes, tWRKY3 and tWRKY4, that are rapidly induced in resistant tobacco plants after infection by tobacco mosaic virus (TMV). Both tWRK3 and tWRKY4 encode proteins with a single WRKY domain that contain the conserved WRKYGQK sequence. Unlike other isolated WRKY proteins that contain the Cys2His2 zinc motif, tWRKY3 and tWRKY4 appear to contain the Cys2HisCys zinc motif. Nonetheless, both tWRKY3 and tWRKY4 are capable of binding DNA molecules with the W-box (TTGAC) element recognized by other WRKY proteins. Expression of the tWRKY3 and tWRKY4 genes could be rapidly induced not only by TMV infection but also by SA or its biologically active analogues that are capable of inducing pathogenesis-related genes and enhanced resistance. Interestingly, induction of both genes by TMV infection was still observed in resistant tobacco plants expressing the bacterial salicylate hydroxylase gene (nahG), although the levels of induction appeared to be reduced. Identification of pathogen- and SA-induced genes encoding WRKY DNA-binding proteins should facilitate future studies on the regulation and functions of this novel group of DNA-binding proteins.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006399311615
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  • 29
    Electronic Resource
    Electronic Resource
    Cellular expression and regulation of the Medicago truncatula cytosolic glutamine synthetase genes in root nodules (2000)
    Springer
    Plant molecular biology 42 (2000), S. 741-756 
    Details
    ISSN: 1573-5028
    Keywords: gene expression ; glutamine synthetase ; legume-Rhizobium symbiosis ; nitrogen assimilation ; root nodules
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract In this paper we have studied the localisation of expression of the two functional cytosolic glutamine synthetase (GS) genes, MtGSa and MtGSb, in root nodules of the model legume Medicago truncatula. We have used a combination of different techniques, including immunocytochemistry, in situ hybridisation and promoter β-glucuronidase (GUS) fusions in transgenic plants, to provide the means of correlating gene expression with protein localisation. These studies revealed that transcriptional regulation (mRNA synthesis) plays an important part in controlling GS protein levels in nodules of M. truncatula. The major locations of cytosolic GS mRNA and protein are the central tissue, the parenchyma and the pericycle of the vascular bundles. These findings indicate that in nodules, GS might be involved in other physiological processes in addition to the primary assimilation of ammonia released by the bacterial nitrogenase. The two genes show different but overlapping patterns of expression with MtGSa being the major gene expressed in the infected cells of the nodule. Promoter fragments of 2.6 kb and 3.1 kb of MtGSa and MtGSb, respectively, have been sequenced and primer extension revealed that the MtGSb promoter is expressed in nodules from an additional start site that is not used in roots. Generally these fragments in the homologous transgenic system were sufficient to drive GUS expression in almost all the tissues and cell types where GS proteins and transcripts are located except that the MtGSa promoter fragment did not express GUS highly in the nodule infected cells. These results indicate that the cis-acting regulatory elements responsible for infected-cell expression are missing from the MtGSa promoter fragment.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006304003770
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  • 30
    Electronic Resource
    Electronic Resource
    Isolation of genes predominantly expressed in guard cells and epidermal cells of Nicotiana glauca (2000)
    Springer
    Plant molecular biology 42 (2000), S. 857-869 
    Details
    ISSN: 1573-5028
    Keywords: epidermis ; gene expression ; glycine-rich protein ; lipid transfer protein ; proline-rich protein ; stomata
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Guard cells are specialized and metabolically active cells which arise during the differentiation of the epidermis. Using Nicotiana glauca epidermal peels as a source of purified guard cells, we have constructed a cDNA library from guard cell RNA. In order to isolate genes that are predominantly expressed in guard cells, we performed a differential screen of this library, comparing the hybridization of a radiolabeled cDNA probe synthesized from guard cell RNA to that from a mesophyll cell cDNA probe. Sixteen clones were isolated based on their greater level of hybridization with the guard cell probe. Of these, eight had high homology to lipid transfer protein (LTP), two were similar to glycine-rich protein (GRP), and one displayed high homology to proline-rich proteins from Arabidopsis thaliana (AtPRP2, AtPRP4) and from potato guard cells (GPP). Northern analysis confirmed that one or more NgLTP genes, NgGRP1, and NgGPP1 are all differentially expressed, with highest levels in guard cells, and low or undetectable levels in mesophyll cells and in roots. In addition, all are induced to some degree in drought-stressed guard cells. NgLTP and NgGRP1 expression was localized by in situ hybridization to the guard cells and pavement cells in the epidermis. NgGRP1 expression was also detected in cells of the vasculature. Genomic Southern analysis indicated that LTP is encoded by a family of highly similar genes in N. glauca. This work has identified members of a subset of epidermis- and guard cell-predominant genes, whose protein products are likely to contribute to the unique properties acquired by guard cells and pavement cells during differentiation.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006480107407
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  • 31
    Electronic Resource
    Electronic Resource
    Flower-predominant expression of a gene encoding a novel class I chitinase in rice (Oryza sativa L.) (2000)
    Springer
    Plant molecular biology 42 (2000), S. 883-897 
    Details
    ISSN: 1573-5028
    Keywords: chitinase function ; flower-predominant ; gene expression ; molecular cloning ; monocotyledon ; promoter
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract A flower-predominant cDNA for a gene, termed OsChia1;175, was isolated from a cDNA library of rice pistils. Northern blot and RT-PCR analyses revealed that the OsChia1;175 gene is highly expressed in floral organs (pistils, stamens and lodicules at the heading stage) but not or at an extremely low level in vegetative organs. OsChia1;175 encodes a protein that consists of 340 amino acid residues, and the putative mature protein shows 52% to 63% amino acid identity to class I chitinases of rice or other plants. The phylogenetic tree shows that the OsChia1;175 protein is a new type of plant class I chitinase in rice. The expression of OsChia1;175 in vegetative organs is not induced by several chemicals, UV, and wounding. The soluble putative mature OsChia1;175 protein expressed in Escherichia coli exhibited chitinase activity in the assay with colloidal chitin as a substrate. Genomic Southern analysis revealed that the OsChia1;175 gene was organized as a low-copy gene family. The rice genomic library was screened and a genome clone corresponding to OsChia1;175 was isolated. The transcription start sites of the OsChia1;175 gene were mapped by primer extension analysis. The 1.2 kb putative promoter region of the OsChia1;175 gene was fused to the GUS (β-glucuronidase) gene, and this chimeric gene was introduced to rice by Agrobacterium-mediated transformation. The flower-predominant gene expression was identified also in the transgenic rice plants. The high promoter activity was detected in the stigmas, styles, stamens and lodicules in transgenic plants. The possible functions of OsChia1;175 are discussed.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006401816145
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  • 32
    Electronic Resource
    Electronic Resource
    Anaerobiosis-specific interaction of tobacco nuclear factors with cis-regulatory sequences in the maize GapC4 promoter (2000)
    Springer
    Plant molecular biology 43 (2000), S. 11-21 
    Details
    ISSN: 1573-5028
    Keywords: anaerobiosis ; electrophoretic mobility shift assays ; gene expression ; glyceraldehyde-3-phosphate dehydrogenase ; Nicotiana tabacum ; transgenic plants
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The promoter of the maize glyceraldehyde-3-phosphate dehydrogenase 4 gene (GapC4) confers strong, specific and ubiquitous anaerobic reporter gene expression in tobacco. To identify factors required for heterologous anaerobic gene expression, 19 progressive 5′ and 3′ promoter deletions were linked to a chimeric GapC4 TATA box-β-glucuronidase (GUS) reporter gene construct and transformed into tobacco. In all transgenic lines aerobic expression values were in the range obtained for negative controls while histochemical GUS assays reveal some weak expression in roots only. Anaerobic induction of about 100-fold to more than 1000-fold above unspecific background is mediated by a region of about 190 bp of the GapC4 promoter. Anaerobic reporter gene induction strongly decreases upon deletion of a 20 bp fragment from −286 to −266 relative to the transcription start point. This fragment harbours putative cis-acting sequences. Electrophoretic mobility shift assays with a 50 bp fragment harbouring these cis sequences reveal a high-mobility complex that is formed with nuclear extracts from aerobic and anaerobic leaf tissue while an additional low-mobility complex is anaerobiosis-specific. The formation of the high-mobility complex requires the sequence GTGGGCCCG. The 50 bp fragment alone confers weak and orientation-dependent anaerobic induction to a GapC4 TATA box-β-glucuronidase (GUS) reporter gene.
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    URL: http://dx.doi.org/10.1023/A:1006419232075
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    Electronic Resource
    Plant A-type cyclins† (2000)
    Springer
    Plant molecular biology 43 (2000), S. 659-675 
    Details
    ISSN: 1573-5028
    Keywords: A-type cyclins ; cell cycle ; gene expression ; regulation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Although the basic mechanisms which control the progression through the cell cycle appear to be conserved in all higher eukaryotes, the unique features of the plant developmental programme must be somehow reflected in a plant-specific regulation of the factors which control cell division. In the past few years, considerable progress has been achieved in identifying the major components of the cell cycle machinery in plants, especially the cyclin-dependent kinases (CDKs) and their regulatory subunits, the cyclins. The question of how these components direct expression of specific genes at specific stages of the cell cycle, and how they are themselves regulated, constitutes a challenge for the present and for the years to come. This review summarizes our current knowledge of a particular class of plant cyclins, the A-type cyclins, which can be further subdivided into three structural groups. The putative functions of these A-type cyclins are discussed in relation to the presence of remarkable motifs in their amino acid sequences, and to their specific transcriptional regulation, protein amount and subcellular localization.
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    URL: http://dx.doi.org/10.1023/A:1006303100592
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  • 34
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    The diageotropica mutation alters auxin induction of a subset of the Aux/IAA gene family in tomato (2000)
    Springer
    Plant molecular biology 44 (2000), S. 73-84 
    Details
    ISSN: 1573-5028
    Keywords: auxin ; Aux/IAA ; dgt ; gene expression ; Lycopersicon esculentum ; signal transduction
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The diageotropica (dgt) mutation has been proposed to affect either auxin perception or responsiveness in tomato plants. It has previously been demonstrated that the expression of one member of the Aux/IAA family of auxin-regulated genes is reduced in dgt plants. Here, we report the cloning of ten new members of the tomato Aux/IAA family by PCR amplification based on conserved protein domains. All of the gene family members except one (LeIAA7) are expressed in etiolated tomato seedlings, although they demonstrate tissue specificity (e.g. increased expression in hypocotyls vs. roots) within the seedling. The wild-type auxin-response characteristics of the expression of these tomato LeIAA genes are similar to those previously described for Aux/IAA family members in Arabidopsis. In dgt seedlings, auxin stimulation of gene expression was reduced in only a subset of LeIAA genes (LeIAA5, 8, 10, and 11), with the greatest reduction associated with those genes with the strongest wild-type response to auxin. The remaining LeIAA genes tested exhibited essentially the same induction levels in response to the hormone in both dgt and wild-type hypocotyls. These results confirm that dgt plants can perceive auxin and suggest that a specific step in early auxin signal transduction is disrupted by the dgt mutation.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006437205596
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  • 35
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    Caleosins: Ca2+-binding proteins associated with lipid bodies (2000)
    Springer
    Plant molecular biology 44 (2000), S. 463-476 
    Details
    ISSN: 1573-5028
    Keywords: Arabidopsis thaliana ; calcium-binding protein ; caleosin ; EF-hand ; gene expression ; lipid bodies ; vesicles
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract We have previously identified a rice gene encoding a 27 kDa protein with a single Ca2+-binding EF-hand and a putative membrane anchor. We report here similar genes termed caleosins, CLO, in other plants and fungi; they comprise a multigene family of at least five members in Arabidopsis (AtClo1–5). Northern hybridization demonstrated that AtClo2–4 mRNAs levels were low in various tissues, while AtClo1 mRNA levels were high in developing embryos and mature seeds. Analysis of transgenic Arabidopsis plants expressing the GUS reporter under control of the AtClo1 promoter showed strong levels of expression in developing embryos and also in root tip cells. Antibodies raised against AtCLO1 were used to detect caleosin in cellular fractions of Arabidopsis and rapeseed. This indicated that caleosins are a novel class of lipid body proteins, which may also be associated with an ER subdomain.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1026564411918
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  • 36
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    Proliferation of Mitochondria in Chronically Stimulated Rabbit Skeletal Muscle—Transcription of Mitochondrial Genes and Copy Number of Mitochondrial DNA (2000)
    Springer
    Journal of bioenergetics and biomembranes 32 (2000), S. 627-634 
    Details
    ISSN: 1573-6881
    Keywords: Mitochondrial biogenesis ; copy number ; gene expression ; mitochondrial transcription factor ; nuclear—mitochondrial communication ; stimulation ; endurance training
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Physics
    Notes: Abstract Mitochondrial proliferation was studied in chronically stimulated rabbit skeletal muscle over a period of 50 days. After this time, subunits of COX had increased about fourfold. Corresponding mRNAs, encoded on mitochondrial DNA as well as on nuclear genes, were unchanged when related to total tissue RNA, however, they were elevated two- to fivefold when the massive increase of ribosomes per unit mass of muscle was taken into account. The same was true for the mRNA encoding mitochondrial transcription factor A. Surprisingly, tissue levels of mtTFA protein were reduced about twofold, together with mitochondrial DNA. In conclusion, mito chondria are able to maintain high rates of mitochondrial transcription even in the presence of reduced mtTFA protein and mtDNA levels. Therefore, stimulated mtTFA gene expression accompanies stimulated mitochondrial transcription, as in other models, but it is not sufficient for an increase of mtDNA copy number and other, yet unknown, factors have to be postulated.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1005630813227
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  • 37
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    Expression of the antibacterial peptide, cecropin, in cultured mammalian cells (2000)
    Springer
    Biotechnology letters 22 (2000), S. 151-155 
    Details
    ISSN: 1573-6776
    Keywords: antibacterial peptide ; cecropin ; gene expression ; mammalian cell
    Source: Springer Online Journal Archives 1860-2000
    Topics: Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract We have stably transfected a Chinese hamster lung cell line V79 with a recombinant gene construct where the Drosophila cecropin A2 cDNA is under the control of Rous sarcoma virus, long terminal repeat (RSV LTR). We have not only been able to demonstrate expression at the RNA level by Northern analysis but also have detected an unprocessed peptide using an antiserum raised against Hyalophora cecropin.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1005609728707
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  • 38
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    Cyclic Core Dendrimer as a New Kind of Vector for Gene Transfer into Mammalian Cells (2000)
    Springer
    Genetica 108 (2000), S. 53-56 
    Details
    ISSN: 1573-6857
    Keywords: dendritic polymer ; reporter gene ; gene expression ; transfection
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Cyclic core dendritic polymer is a new type of synthetic polymers. The ability of generation 4 of the dendrimer with a core of 1,4,7,10-tetraazacyclododecane to function as an effective gene delivery vector was investigated. Results from fluorescence in situhybridization (FISH) show that the pCH 110 plasmid DNA was transferred into human small intestine cancer metastatic ascites (HICMA) cells induced by this kind of dendrimer as a vector. The transferred LacZ, GFP and luciferase genes were highly expressed in the transfected HICMA, COS-7 and 293 cells. These studies demonstrate that the dendrimer can transfect mammalian cells in vitrowhich offers an alternatively efficient method for mammalian gene transfer.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1004047902652
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  • 39
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    Electronic Resource
    Regulation of Keratinocyte Growth Factor (KGF) and KGF Receptor mRNAs by Nutrient Intake and KGF Administration in Rat Intestine (2000)
    Springer
    Digestive diseases and sciences 45 (2000), S. 736-743 
    Details
    ISSN: 1573-2568
    Keywords: keratinocyte growth factor ; KGF receptor ; gene expression ; intestine ; nutrition
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract The aim of this study was to investigate the regulation of keratinocyte growth factor (KGF) and KGF receptor mRNAs by diet and KGF treatment in rat intestine. Fasting for three days up-regulated KGF and KGF receptor mRNA levels in ileum and increased KGF receptor mRNA expression in colon. KGF and KGF receptor mRNA levels returned toward control values with ad libitum refeeding but remained elevated when refeeding was limited to 25% of ad libitum intake. KGF treatment during nutrient repletion did not alter intestinal KGF mRNA levels but increased KGF receptor mRNA abundance in ileum and colon. We conclude that the increase in KGF and KGF receptor mRNAs induced by malnutrition may be an adaptive response to attenuate gut mucosal atrophy in this setting. The gut-trophic effects of KGF treatment may be mediated, in part, by up-regulation of the KGF receptor mRNA in small bowel and colon.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1005447827579
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  • 40
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    Electronic Resource
    Effects of increased and deregulated expression of cell division genes on the morphology and on antibiotic production of streptomycetes (2000)
    Springer
    Antonie van Leeuwenhoek 78 (2000), S. 269-276 
    Details
    ISSN: 1572-9699
    Keywords: differentiation ; FtsZ ; gene expression ; septum ; SsgA ; transmission electron microscopy
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract This paper describes the effects of increased expression of the cell division genes ftsZ, ftsQ, and ssgA on the development of both solid- and liquid-grown mycelium of Streptomyces coelicolor and Streptomyces lividans. Over-expression of ftsZ in S. coelicolor M145 inhibited aerial mycelium formation and blocked sporulation. Such deficient sporulation was also observed for the ftsZ mutant. Over-expression of ftsZ also inhibited morphological differentiation in S. lividans 1326, although aerial mycelium formation was less reduced. Furthermore, antibiotic production was increased in both strains, and in particular the otherwise dormant actinorhodin biosynthesis cluster of S. lividans was activated in liquid- and solid-grown cultures. No significant alterations were observed when the gene dosage of ftsQ was increased. Analysis by transmission electron microscopy of an S. coelicolor strain over-expressing ssgA showed that septum formation had strongly increased in comparison to wild-type S. coelicolor, showing that SsgA clearly influences Streptomyces cell division. The morphology of the hyphae was affected such that irregular septa were produced with a significantly wider diameter, thereby forming spore-like compartments. This suggests that ssgA can induce a process similar to submerged sporulation in Streptomyces strains that otherwise fail to do so. A working model is proposed for the regulation of septum formation and of submerged sporulation.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1010267708249
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  • 41
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    Expression of Four Genes of Bacteriophage MB78 from Contiguous Open Reading Frames: The Genomic Organization as Deduced by Sequence Analysis (2000)
    Springer
    Virus genes 20 (2000), S. 87-97 
    Details
    ISSN: 1572-994X
    Keywords: bacteriophage MB78 ; contiguous ORF ; unusual start codons ; genome analysis ; gene expression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract Four proteins of bacteriophage MB78 having apparent molecular weights as 35, 14, 21 and 16 kDa are expressed from 3.9 kb SalI-HindIII fragment located almost in the middle of the phage genome. Analysis of the sequence supported by some experimental evidences suggest that these four proteins are expressed from polycistronic message without any intercistronic gap. Stop and start codons of consecutive ORFs overlap and rare initiation codons are used. Computer analysis of the sequence suggests the presence of two more open reading frames within the ORFs of 35 and 16 kDa proteins but in the opposite orientation, i.e. in the complementary strand.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1008168425571
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  • 42
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    Electronic Resource
    Ethylene regulation of fruit ripening: Molecular aspects (2000)
    Springer
    Plant growth regulation 30 (2000), S. 193-200 
    Details
    ISSN: 1573-5087
    Keywords: ACC synthase ; ACC oxidase ; ethylene ; fruit ; gene expression ; regulation ; ripening
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract Progress in ethylene regulating fruit ripening concerning itsperception and signal transduction and expression of ACC synthaseand ACC oxidase genes is reviewed. ACC synthase and ACC oxidasehave been characterized and their genes cloned from various fruittissues. Both ACC synthase and ACC oxidase are encoded bymultigene families, and their activities are associated withfruit ripening. In climacteric fruit, the transition toautocatalytic ethylene production appears to be due to a seriesof events in which ACC sythase and ACC oxidase genes have beenexpressed developmentally. Differential expression of ACCsynthase and ACC oxidase gene family members is probably involvedin such a transition that ultimately controls the onset of fruitripening.In comparison to ACC synthase and ACC oxidase, less is knownabout ethylene perception and signal transduction because of thedifficulties in isolating and purifying ethylene receptors orethylene-binding proteins using biochemical methods. However, theidentification of the Nr tomato ripening mutant as anethylene receptor, the applications of new potent anti-ethylenecompounds and the generation of transgenic fruits with reducedethylene production have provided evidence that ethylenereceptors regulate a defined set of genes which are expressedduring fruit ripening. The properties and functions of ethylenereceptors, such as ETR1, are being elucidated.Application of molecular genetics, in combination withbiochemical approaches, will enable us to better understand theindividual steps leading from ethylene perception and signaltransduction and expression of ACC synthase and ACC oxidase genefamily member to the physiological responses.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006348627110
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  • 43
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    Resistance to wheat leaf rust in Aegilops tauschii Coss. and inheritance of resistance in hexaploid wheat (2000)
    Springer
    Genetic resources and crop evolution 47 (2000), S. 135-140 
    Details
    ISSN: 1573-5109
    Keywords: Aegilops tauschii ; gene expression ; genetic inheritance ; Puccinia recondita f. sp. tritici ; rust resistance ; synthetic hexaploid wheats
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract A collection of 164 Aegilops tauschii accessions, obtained from Gatersleben, Germany, was screened for reaction to leaf rust under controlled greenhouse conditions. We have also evaluated a selection of synthetic hexaploid wheats, produced by hybridizing Ae. tauschii with tetraploid durum wheats, as well as the first and second generation of hybrids between some of these resistant synthetic hexaploid wheats and susceptible Triticum aestivum cultivars. Eighteen (11%) accessions of Ae. tauschii were resistant to leaf rust among which 1 was immune, 13 were highly resistant and 4 were moderately resistant. Six of the synthetic hexaploid wheats expressed a high level of leaf rust resistance while four exhibited either a reduced or complete susceptibility compared to their corresponding diploid parent. This suppression of resistance at the hexaploid level suggests the presence of suppressor genes in the A and/or B genomes of the T. turgidum parent. Inheritance of leaf rust resistance from the intercrosses with susceptible bread wheats revealed that resistance was dominant over susceptibility. Leaf rust resistance from the three synthetics (syn 101, syn 701 and syn 901) was effectively transmitted as a single dominant gene and one synthetic (syn 301) possessed two different dominant genes for resistance.
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    URL: http://dx.doi.org/10.1023/A:1008770226330
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  • 44
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    Electronic Resource
    Mechanisms regulating differential freezing tolerance of suspension cell cultures derived from contrasting alfalfa genotypes (2000)
    Springer
    Plant cell, tissue and organ culture 61 (2000), S. 143-151 
    Details
    ISSN: 1573-5044
    Keywords: fall dormancy ; gene expression ; Medicago sativa L. ; protein ; starch ; sugar ; winter hardiness
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract A major factor limiting persistence of alfalfa (Medicago sativa L.) in the northern US is poor winter hardiness. Our hypothesis is that suspension cell cultures derived from dormant, winter-hardy alfalfa cultivars would cold acclimate and survive sub-zero temperatures better than cell cultures derived from non-dormant, non-hardy cultivars. Our objectives were (1) to determine if genetic differences in winter hardiness between dormant and non-dormant alfalfa were retained by suspension cells derived from these contrasting cultivars; and (2) to determine the physiological and biochemical bases for differences in freezing tolerance of suspension cells. Cell suspensions derived from `5262' (fall dormant) and `5929' (fall non-dormant) were cold hardened at 2 °C for 14 days. Cells were frozen in a cooling bath and cell survival determined by measuring 2, 3, 5-triphenyltetrazolium chloride (TTC) reduction. Cold acclimation improved cell survival of both cultivars to −5 °C when compared to unacclimated cells. Only acclimated cells of 5262 survived temperatures of −10 °C to −25 °C. The freezing tolerance of cold-acclimated 5262 cells was associated with high sugar and starch concentrations, lower α-amylase activities and slightly lower cell protein levels when compared to 5929. No differences in polypeptide composition were evident when comparing acclimated and unacclimated cells of 5929, but polypeptide composition did change with acclimation of 5262 cells. As expected, expression of RootCAR1 in 5262 cells increased with cold acclimation, but high levels of RootCAR1 transcript were unexpectantly found in both cold acclimated and unacclimated 5929 cells. With the exception of the RootCAR1 expression, many of the physiological responses of these alfalfa cell lines to cold acclimation were similar to those that have been reported for field-grown plants.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006408829105
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  • 45
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    Electronic Resource
    Binding of cell type-specific nuclear proteins to the 5′-flanking region of maize C4 phosphoenolpyruvate carboxylase gene confers its differential transcription in mesophyll cells (2000)
    Springer
    Plant molecular biology 44 (2000), S. 543-557 
    Details
    ISSN: 1573-5028
    Keywords: C4 photosynthesis ; maize ; phosphoenolpyruvate carboxylase ; transgenic plant ; transcription ; gene expression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract C4-type phosphenolpyruvate carboxylase (C4PEPC) acts as a primary carbon assimilatory enzyme in the C4 photosynthetic pathway. The maize C4PEPC gene (C4Ppc1) is specifically expressed in mesophyll cells (MC) of light-grown leaves, but the molecular mechanism responsible for its cell type-specific expression has not been characterized. In this study, we introduced a chimeric maize C4Ppc1 5′-flanking region/β-glucuronidase (GUS) gene into maize plants by Agrobacterium-mediated transformation. Activity assay and histochemical staining showed that GUS is almost exclusively localized in leaf MC of transgenic maize plants. This observation suggests that the introduced 5′ region of maize C4Ppc1 contains the necessary cis element(s) for its specific expression in MC. Next, we investigated whether the 5′ region of the maize gene interacts with nuclear proteins in a cell type-specific manner. By gel shift assays with nuclear extracts prepared from MC or bundle sheath cells (BSC), cell type-specific DNA-protein interactions were detected: nuclear factors PEPIb and PEPIc are specific to MC whereas PEPIa and PEPIIa are specific to BSC. Light alters the binding activity of these factors. These interactions were not detected in the assay with nuclear extract prepared from root, or competed out by oligonucleotides corresponding to the binding sites for the maize nuclear protein, PEP-I, which is known to bind specifically to the promoter region of C4Ppc1. The results suggest that novel cell type-specific positive and negative nuclear factors bind to the maize C4Ppc1 5′-flanking region and regulate its differential transcription in MC in a light-dependent manner.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1026565027772
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  • 46
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    Molecular characterization of quinolinate phosphoribosyltransferase (QPRTase) in Nicotiana (2000)
    Springer
    Plant molecular biology 44 (2000), S. 603-617 
    Details
    ISSN: 1573-5028
    Keywords: gene expression ; nicotinic acid ; pyridine alkaloids ; secondary metabolism ; polyploidy ; wound-induced
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Quinolate acid phosphoribosyltransferase (QPRTase), a key enzyme in nicotinamide adenine dinucleotide (NAD) biosynthesis, also plays an important role in ensuring nicotinic acid is available for the synthesis of defensive pyridine alkaloids in Nicotiana species. In this study, cDNAs for QPRTase were characterized from N. rustica and N. tabacum. Deduced proteins from both cDNAs are almost identical and contain a 24 amino acid N-terminal extension, not reported in other QPRTases, that has characteristics of a mitochondrial targeting sequence. In N. tabacum and N. sylvestris, both of which contain nicotine as the major pyridine alkaloid, QPRTase transcript was detected in roots, the site of nicotine synthesis, but not in leaves. QPRTase transcript levels increased markedly in roots of both species 12–24 h after damage to aerial tissues, with a concomitant rise in transcript levels of putrescine N-methyltransferase (PMT), another key enzyme in nicotine biosynthesis. In N. glauca, however, in which anabasine represents the major pyridine alkaloid, QPRTase transcript was detected in both leaf and root tissues. Moreover, wound induction of QPRTase but not PMT was observed in leaf tissues, and not in roots, 12–24 h after wounding. Southern analysis of genomic DNA from the Nicotiana species noted above, and also several others from within the genus, suggested that QPRTase is encoded by a small gene family in all the species investigated.
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    URL: http://dx.doi.org/10.1023/A:1026590521318
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  • 47
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    Hexose transporters of tomato: molecular cloning, expression analysis and functional characterization (2000)
    Springer
    Plant molecular biology 44 (2000), S. 687-697 
    Details
    ISSN: 1573-5028
    Keywords: gene expression ; heterologous expression ; H+/hexose symporter ; Lycopersicon esculentum ; quantitative PCR ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract A full-length (LeHT2) and two partial (LeHT1 and LeHT3) cDNA clones, encoding hexose transporters, were isolated from tomato (Lycopersicon esculentum) fruit and flower cDNA libraries. Southern blot analysis confirmed the presence of a gene family of hexose transporters in tomato consisting of at least three members. The full-length cDNA (LeHT2) encodes a protein of 523 amino acids, with a calculated molecular mass of 57.6 kDa. The predicted protein has 12 putative membrane-spanning domains and belongs to the Major Facilitator Superfamily of membrane carriers. The three clones encode polypeptides that are homologous to other plant monosaccharide transporters and contain conserved amino acid motifs characteristic of this superfamily. Expression of the three genes in different organs of tomato was investigated by quantitative PCR. LeHT1 and LeHT3 are expressed predominantly in sink tissues, with both genes showing highest expression in young fruit and root tips. LeHT2 is expressed at relatively high levels in source leaves and certain sink tissues such as flowers. LeHT2 was functionally expressed in a hexose transport-deficient mutant (RE700A) of Saccharomyces cerevisiae. LeHT2-dependent transport of glucose in RE700A exhibited properties consistent with the operation of an energy-coupled transporter and probably a H+/hexose symporter. The K m of the symporter for glucose is 45 μM.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1026578506625
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  • 48
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    Spatial and temporal patterns of GUS expression directed by 5′ regions of the Arabidopsis thaliana farnesyl diphosphate synthase genes FPS1 and FPS2 (2000)
    Springer
    Plant molecular biology 44 (2000), S. 747-758 
    Details
    ISSN: 1573-5028
    Keywords: Arabidopsis thaliana ; β-glucuronidase (GUS) reporter gene ; farnesyl diphosphate synthase ; gene expression ; isoprenoids ; transgenic plants
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Farnesyl diphosphate synthase (FPS), the enzyme that catalyses the synthesis of farnesyl diphosphate (FPP) from isopentenyl diphosphate (IPP) and dimethylallyl diphosphate (DMAPP), is considered a regulatory enzyme of plant isoprenoid biosynthesis. The promoter regions of the FPS1 and FPS2 genes controlling the expression of isoforms FPS1S and FPS2, respectively, were fused to the β-glucuronidase (GUS) reporter gene and introduced into Arabidopsis thaliana plants. The FPS1S:GUS gene is widely expressed in all plant tissues throughout development, thus supporting a role for FPS1S in the synthesis of isoprenoids serving basic plant cell functions. In contrast, the FPS2:GUS gene shows a pattern of expression restricted to specific organs at particular stages of development. The highest levels of GUS activity are detected in flowers, especially in pollen grains, from the early stages of flower development. After pollination, much lower levels of GUS activity are detected in the rest of floral organs, with the exception of the ovary valves, which remain unstained throughout flower development. GUS activity is also detected in developing and mature seeds. In roots, GUS expression is primarily detected at sites of lateral root initiation and in junctions between primary and secondary roots. No GUS activity is detected in root apical meristems. GUS expression is also observed in junctions between primary and secondary stems. Overall, the pattern of expression of FPS2:GUS suggests a role for FPS2 in the synthesis of particular isoprenoids with specialized functions. Functional FPS2 gene promoter deletion analysis in transfected protoplasts and transgenic A. thaliana plants indicate that all the cis-acting elements required to establish the full pattern of expression of the FPS2 gene are contained in a short region extending from positions −111 to +65. The potential regulatory role of specific sequences within this region is discussed.
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    URL: http://dx.doi.org/10.1023/A:1026588708849
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  • 49
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    Electronic Resource
    Biosynthesis and secretion of recombinant human growth hormone in Pichia pastoris (2000)
    Springer
    Biotechnology letters 22 (2000), S. 109-114 
    Details
    ISSN: 1573-6776
    Keywords: alcohol oxidase ; gene expression ; human growth hormone ; Pichia pastoris
    Source: Springer Online Journal Archives 1860-2000
    Topics: Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract Mature human growth hormone (hGH) cDNA was cloned by homologous recombination into the yeast Pichia pastoris genome. The hGH gene expression was placed under the control of the methanol-inducible alcohol oxidase 1 (AOX1) gene promoter and the Saccharomyces cerevisiae α-factor signal sequence to direct the secretion of recombinant human growth hormone (rhGH) into the growth medium. O2-limited induction of recombinant yeast strains in shake tubes with 3 ml of culture medium produced up to 11 mg rhGH l−1, while high cell density cultures using a 2-l bioreactor produced about 49 mg rhGH l−1 achieving 40% of total protein of the culture medium supernatant.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1005675920451
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    PCR cloning of Pseudomonas resinovorans polyhydroxyalkanoate biosynthesis genes and expression in Escherichia coli (2000)
    Springer
    Biotechnology letters 22 (2000), S. 789-794 
    Details
    ISSN: 1573-6776
    Keywords: gene expression ; PCR cloning ; pha genes ; polyhydroxyalkanoates ; Pseudomonas resinovorans
    Source: Springer Online Journal Archives 1860-2000
    Topics: Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract A ca. 5.5-kb region of Pseudomonas resinovorans genome containing the polyhydroxyalkanoate (PHA) biosynthesis locus was sequenced. Three complete open-reading-frames (ORFs), i.e., phaC1 Pr, phaZ Pr, and phaC2 Pr, were identified. Using this sequence information, phaC1 Pr was PCR-cloned from P. resinovorans genomic DNA and expressed in E. coli as shown by a Nile Red plate assay and gas chromatography/mass spectrometric analysis.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1005614209342
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    Gene expression in allergenic pollen (2000)
    Springer
    Aerobiologia 16 (2000), S. 331-334 
    Details
    ISSN: 1573-3025
    Keywords: allergenic pollen ; gene expression ; molecular biology
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The molecular mechanism of gene expression for pollen specificity is not yet fully known. However, it is an exciting area with great potential and has a wide scope of application in the field of molecular biology, breeding systems, biotechnology etc. The main aim of this write-up is to review some of the interesting achievements made through studies like gene expression in allergic pollen and the research which will make a way towards practical application of pollen molecular biology in identifying and isolating the genes responsible for all allergic disorders reported among various individuals.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1026515820294
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    Comparison of RAP-PCR analysis of gene expression in fresh and immortalised rat hepatocyte cell lines (2000)
    Springer
    Cytotechnology 34 (2000), S. 159-163 
    Details
    ISSN: 1573-0778
    Keywords: gene expression ; immortalised hepatocytes ; RAP-PCR ; RT-PCR
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract Primary rat hepatocytes dedifferentiate rapidly losing theactivities of the drug metabolising enzymes involved in thedetoxification of xenobiotics in the liver. An alternativeapproach to using primary hepatocytes for toxicity testing isthe development of immortalised hepatocyte cell lines via thetransfection of primary hepatocytes with SV40 DNA. In order toassess the suitability of immortalised lines as an alternativeto primary cell cultures we have used RNA arbitrarily primedpolymerase chain reaction to compare gene expression inimmortalised rat hepatocyte cell lines with that in primary rathepatocytes. We have found that differences exist in the RNAtranscripts between fresh and immortalised hepatocyteshighlighting RNA arbitrarily primed polymerase chain reaction asa useful screening method for identifying immortalised lineswhich retain the most `normal' phenotype in relation to theprimary cells from which they originated.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1008159912634
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    Detection and cDNA cloning of H-strand mitochondrial regulatory region RNAs in cultured human cells and human tissues (2000)
    Springer
    Cytotechnology 33 (2000), S. 175-188 
    Details
    ISSN: 1573-0778
    Keywords: cDNA ; differentiation ; gene expression ; growth ; humancells ; human tissues ; mitochondrial DNA ; mitochondrial RNA ; polyadenylated RNA
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract In the mitochondrion, essential genetic elements for replication and transcription are mostly housed within a shortsegment of its DNA located between tRNAPhe and tRNAPro genes, which is called mitochondrial regulatoryregion (mrr). RNAs are known to be transcribed from mrr, thestructures and the functions of which are yet to be fullycharacterized.We detected ca. 1.3 kb H-strand transcripts of mrr (mrrH-RNAs),and 0.2 kb L-strand transcripts of mrr (mrrL-RNAs) in varioushuman cultured cells and tissues using double stranded mrrDNAprobes. The steady state levels of mrrL-RNAs were generally highin cultured cells, while they varied among tissues. On the otherhand, the levels of mrrH-RNAs varied among tissues and amongcultured cells. A tendency was observed in these cells andtissues that a high level of mrrL-RNA is associated with cellproliferation, and a high level of mrrH-RNA withdifferentiation. Several cDNA clones to 1.3 kb mrrH-RNA were obtained from humanskeletal muscle polyadenylated RNAs. The 5′ terminus of the 1.3 kb RNA was determined to be at nucleotide position 15953 whichis immediately downstream of tRNAThr sequence.Polyadenylation site for most of the clones was demonstrated tobe at nucleotide position 576 which is immediately upstream oftRNAPhe sequence. The longest cDNA insert obtained was 1177 bps long spanning from nucleotide positions 15969 to 576 which could code for a peptide of 76 amino acids. The cDNAs isolatedhere are the first cDNA clones reported to human mrrH-RNAs.These results, together with previous results, furthersubstantiate that polyadenylated mrrH- and mrrL-RNAs are commonly present at varying levels among human tissues andcells. The 3′ end sequences of the cloned mrrH-cDNA provideswith insights into the mechanisms of transcription termination.The cDNA clones will provide tools to further the study of thefunction of mrr RNAs.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1008154027997
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    Effect of Peptide Lys-Glu on Interleukin-2 Gene Expression in Lymphocytes (2000)
    Springer
    Bulletin of experimental biology and medicine 130 (2000), S. 898-899 
    Details
    ISSN: 1573-8221
    Keywords: peptides ; gene expression ; interleukin-2 ; lymphocytes
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract Lys-Glu in vitro stimulated interleukin-2 gene expression in mouse spleen lymphocytes. This effect depended on peptide concentration and duration of treatment. It is hypothesized that this peptide is the shortest regulatory fragment promoting the transport of trans-acting factors into the nucleus. It can not be excluded that Lys-Glu is a structural component of trans-acting factor active centers, which are necessary for the activation of interleukin-2 gene transcription in lymphocytes.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1015382732640
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    Regulation of Functional Activity of Bone Marrow Hemopoietic Stem Cells by Erythroid Cells in Mice (2000)
    Springer
    Bulletin of experimental biology and medicine 130 (2000), S. 1159-1161 
    Details
    ISSN: 1573-8221
    Keywords: erythroblasts ; cytokines ; gene expression ; hemopoietic stem cell
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract Transplantation of erythroid and bone marrow cells to irradiated mice stimulated exogenous colony formation. Pretreatment of erythroid cells with specific rabbit antiserum to erythroblasts abolished this effect. The reverse transcriptase polymerase chain reaction revealed the presence of mRNA for interleukin-1α, interleukin-1β, interleukin-3, interleukin-6, and granulocyte-macrophage colony-stimulating factor in erythroid cells. Granulocyte-macrophage colony-stimulating factor was found in the conditioned medium from erythroid cells. Thus, erythroid cells stimulated colony-forming activity of bone marrow cells, which was probably mediated via cytokine synthesis (e.g., granulocyte-macrophage colony-stimulating factor).
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1017523800090
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    Pollen Embryogenesis - The Stress Mediated Switch from Gametophytic to Sporophytic Development. Current Status and Future Prospects (2000)
    Springer
    Biologia plantarum 43 (2000), S. 481-489 
    Details
    ISSN: 1573-8264
    Keywords: cell symmetry ; gene expression ; heat shock proteins ; polarity
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Embryogenesis can be initiated directly from microspores or pollen grains. This is known as androgenesis and refers to the process of redirection of normal pollen development (gametophytic pathway) towards the embryo formation (sporophytic). This review mainly deals with the current knowledge of stress and developmental aspects of induction of androgenesis. The crucial role of stress inductive treatment together with changes in cell polarity are discussed in relation to other relevant biological systems. The intriguing speculations are made on the basis of these comparisons which may point out the direction of future investigations.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1002835330799
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    Isolation of RNA and Protein from Guard Cells of Nicotiana glauca (1999)
    Springer
    Plant molecular biology reporter 17 (1999), S. 371-383 
    Details
    ISSN: 1572-9818
    Keywords: epidermal peel ; extraction ; gene expression ; stomata ; tree tobacco
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Stomatal guard cells are critical for maintenance of plant homeostasis and represent an interesting cell type for studies of leaf cell differentiation and patterning. Here we describe techniques for the isolation of guard cell RNA and protein from blended epidermal peels of Nicotiana glauca. The RNA isolation procedure is a modification of the hot borate method, which is particularly well-suited for recalcitrant tissues. Protein was extracted by disrupting guard cell-enriched epidermis with a French® press. This system offers the following advantages: relatively high yield, low or no contamination by other cell types, fresh tissue as a source of RNA and protein rather than protoplasts, and a plant species that is readily transformable. These techniques will allow for cloning and analysis of genes expressed in guard cells, application of traditional biochemical techniques to guard cell proteins, as well as characterization of genetic manipulation of guard cell function in transgenic plants.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1007602017492
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    PPARγ, the ultimate thrifty gene (1999)
    Springer
    Diabetologia 42 (1999), S. 1033-1049 
    Details
    ISSN: 1432-0428
    Keywords: Keywords Adipogenesis ; adipose tissue ; cofactors ; gene expression ; fatty acids ; insulin resistance ; nuclear receptors ; prostaglandin ; thiazolidinediones ; Type II diabetes ; transcription.
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract The peroxisome proliferator-activated receptor gamma (PPARγ) quickly evolved over the last decade from a new orphan receptor to one of the best characterized nuclear receptors. This fast pace in PPARγ research was triggered by two main discoveries. Firstly, that PPARγ was shown to have a key role in adipogenesis and be a master controller of the “thrifty gene response” leading to efficient energy storage. Secondly, the discovery that its synthetic ligands, the thiazolidinediones, are promising insulin sensitizing drugs, which are currently being developed for the treatment of Type II (non-insulin-dependent) diabetes mellitus. More recently this nuclear receptor emerged from a role limited to metabolism (diabetes and obesity) to a power player in general transcriptional control of numerous cellular processes, with implications in cell cycle control, carcinogenesis, inflammation, atherosclerosis and immunomodulation. This widened role of PPARγ will certainly initiate a new flurry of research, which will not only refine our current often partial knowledge of PPARγ but more importantly also establish that this receptor has a definite role as a primary link adapting cellular, tissue and whole body homeostasis to energy stores. [Diabetologia (1999) 42: 1033–1049]
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/s001250051268
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    Uncoupling protein-3 gene expression: reduced skeletal muscle mRNA in obese humans during pronounced weight loss (1999)
    Springer
    Diabetologia 42 (1999), S. 302-309 
    Details
    ISSN: 1432-0428
    Keywords: Keywords Obesity ; genetics ; uncoupling protein-3 ; gene expression ; skeletal muscle.
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract Aims: Uncoupling protein-3 is a member of a protein family that serves to dissipate energy in the form of heat thereby modulating energy expenditure. Alternative processing of uncoupling protein-3 transcripts results in two mRNA species that encode a large and small protein, perhaps differing in functional activity. Since obesity is associated with disrupted energy homeostasis, we measured muscle mRNA expression in morbidly obese and lean subjects. Methods: The two uncoupling protein-3 mRNA species were quantified in muscle tissue using an RNase protection assay. Gene locus effects on mRNA expression were studied by quantitative allele-specific primer extension. Results: In both obese and lean subjects, the mRNA species encoding the small protein isoform was twice as abundant as the mRNA species encoding the large protein isoform. Neither the total uncoupling protein-3 mRNA expression nor the molar abundance ratios of the two mRNA species differed between obese and lean male or female subjects. Women who had lost 37 ± 22 kg of weight in response to dietary restriction and continued a hypocaloric diet displayed lower mRNA than obese (p 〈 0.005) or lean women (p 〈 0.05). Primer extension assays in lean and obese subjects showed similar allelic mRNA abundance in all but one subject studied. Conclusion: Muscle expression of the two uncoupling protein-3 mRNA species is similar in obese and lean people. In obese patients, prolonged hypocaloric diet downregulates uncoupling protein-3 mRNA expression in muscle and can thereby enhance its energy efficiency. Sequence substitutions at the gene locus may only be minor determinants of mRNA expression in muscle tissue. [Diabetologia (1999) 42: 302–309]
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/s001250051155
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    Regulation of UCP2 and UCP3 by muscle disuse and physical activity in tetraplegic subjects (1999)
    Springer
    Diabetologia 42 (1999), S. 826-830 
    Details
    ISSN: 1432-0428
    Keywords: Keywords Uncoupling proteins ; exercise ; tetraplegia ; skeletal muscle ; mRNA ; gene expression ; polymerase chain reaction.
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract Aims/hypothesis. The regulation of uncoupling protein 2 and uncoupling protein 3 gene expression in skeletal muscle has recently been the focus of intense interest. Our aim was to determine expression of uncoupling protein 2 and 3 in skeletal muscle from tetraplegic subjects, a condition representing profound muscle inactivity. Thereafter we determined whether exercise training would modify expression of these genes in skeletal muscle. Methods. mRNA expression of uncoupling protein 2 and 3 was determined using quantitative reverse transcription-polymerase chain-reaction. Results. Expression of uncoupling protein 2 and 3 mRNA was increased in skeletal muscle from tetraplegic compared with able-bodied subjects (3.7-fold p 〈 0.01 and 4.1-fold, p 〈 0.05, respectively). A subgroup of four tetraplegic subjects underwent an 8-week exercise programme consisting of electrically-stimulated leg cycling (ESLC, 7 ESLC sessions/week). This training protocol leads to increases in whole body insulin-stimulated glucose uptake and expression of genes involved in glucose metabolism in skeletal muscle from tetraplegic subjects. After ESLC training, uncoupling protein 2 expression was reduced by 62 % and was similar to that in able-bodied people. Similarly, ESLC training was associated with a reduction of uncoupling protein 3 expression in skeletal muscle from three of four tetraplegic subjects, however, post-exercise levels remained increased compared with able-bodied subjects. Conclusion/interpretation. Tetraplegia is associated with increased mRNA expression of uncoupling protein 2 and 3 in skeletal muscle. Exercise training leads to normalisation of uncoupling protein 2 expression in tetraplegic subjects. Muscle disuse and physical activity appear to be powerful regulators of uncoupling protein 2 and 3 expression in human skeletal muscle. [Diabetologia (1999) 42: 826–830]
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/s001250051233
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    Novel Technology for Detection of Genomic and Transcriptional Alterations in Pancreatic Cancer (1999)
    Springer
    Annals of oncology 10 (1999), S. 64-68 
    Details
    ISSN: 1569-8041
    Keywords: chromosomal aberrations ; gene expression ; oncogenes ; pancreatic cancer ; tumor suppressor genes
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract Aim: The present review summarizes our strategies aimed at identifying and characterizing genetic alterations occuring at the transcriptional and chromosomal level in pancreatic cancer. Methods: To study transcriptional alterations we have used a number of techniques including modified versions of differential hybridizations and cDNA-RDA (representational difference analysis). Comparative genomic hybridization (CGH) was used to study chromosomal aberrations occuring in pancreatic cancer tissues. Results: The study of transcriptional alterations led to the identification of more than 500 genes with differential expression in pancreatic cancer. The sum of these alterations represented the first expression profile characteristic for pancreatic tumors. The CGH analysis allowed the identification of a number of chromosomal regions containing putative tumor suppressor genes or oncogenes. These regions are presently being characterized at the molecular level. In a first approach the myb-oncogene was identified as the relevant oncogene of an amplification on 6q occurring in up to 10% of pancreatic cancer patients. Conclusions: Genes isolated in both approaches represent potential new disease genes for pancreatic cancer and are at present being characterized by individual or serial analysis.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1008392904359
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    Increased peroxisomal fatty acid β-oxidation and enhanced expression of peroxisome proliferator-activated receptor-α in diabetic rat liver (1999)
    Springer
    Molecular and cellular biochemistry 194 (1999), S. 227-234 
    Details
    ISSN: 1573-4919
    Keywords: microbodies ; diabetes mellitus ; steroid hormone receptor ; β-oxidation ; gene expression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract To determine whether the increased fatty acid β-oxidation in the peroxisomes of diabetic rat liver is mediated by a common peroxisome proliferation mechanism, we measured the activation of long-chain (LC) and very long chain (VLC) fatty acids catalyzed by palmitoyl CoA ligase (PAL) and lignoceryl CoA ligase and oxidation of LC (palmitic acid) and VLC (lignoceric acid) fatty acids by isotopic methods. Immunoblot analysis of acyl-CoA oxidase (ACO), and Northern blot analysis of peroxisome proliferator-activated receptor (PPAR-α), ACO, and PAL were also performed. The PAL activity increased in peroxisomes and mitochondria from the liver of diabetic rats by 2.6-fold and 2.1-fold, respectively. The lignoceroyl-CoA ligase activity increased by 2.6-fold in diabetic peroxisomes. Palmitic acid oxidation increased in the diabetic peroxisomes and mitochondria by 2.5-fold and 2.7-fold, respectively, while lignoceric acid oxidation increased by 2.0-fold in the peroxisomes. Immunoreactive ACO protein increased by 2-fold in the diabetic group. The mRNA levels for PPAR-α, ACO and PAL increased 2.9-, 2.8- and 1.6-fold, respectively, in the diabetic group. These results suggest that the increased supply of fatty acids to liver in diabetic state stimulates the expression of PPAR-α and its target genes responsible for the metabolism of fatty acids.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006930513476
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    Expression of calcium-binding protein regucalcin mRNA in the cloned rat hepatoma cells (Hr-II-E) is stimulated through Ca2+ signaling factors: Involvement of protein kinase C (1999)
    Springer
    Molecular and cellular biochemistry 198 (1999), S. 101-107 
    Details
    ISSN: 1573-4919
    Keywords: regucalcin ; Ca2+-binding protein ; protein kinase C ; Ca2+signaling ; gene expression ; H4-II-E hepatoma cells
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract The expression of hepatic Ca2+-binding protein regucalcin in the cloned rat hepatoma cells (H4-II-E) was investigated. The change in regucalcin mRNA levels was analyzed by Northern blotting using rat liver regucalcin complementary DNA (0.9 kb of open reading frame). Regucalcin mRNA was expressed in H4-II-E hepatoma cells. This expression was clearly stimulated in the presence of serum (10% fetal bovine serum). Bay K 8644 (2. 5 × 10-6 M), a Ca2+ channel agonist, significantly stimulated regucalcin mRNA expression in the absence or presence of 10% serum. Dibutyryl cyclic AMP (10-3 M) did not have a stimulatory effect on the regucalcin mRNA expression. The presence of phorbol 12-myristate 13-acetate (PMA; 10-6 M) or estrogen (10-8 M) caused a significant increase in regucalcin mRNA levels in the hepatoma cells cultured in serum-free medium, while insulin (5 × 10-9 M) or dexamethasone (10-6 M) had no effect. Bay K 8644-stimulated regucalcin mRNA expression in the hepatoma cells was completely blocked in the presence of trifluoperazine (10-5 M), an antagonist of calmodulin, or staurosporine (10-7 M), an inhibitor of protein kinase C. The stimulatory effect of PMA was clearly inhibited in the presence of stauroporine. The present study demonstrates that regucalcin mRNA is expressed in the transformed H4-II-E hepatoma cells, and that the expression is stimulated through Ca2+-dependent signaling factors.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006996506238
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    Posttranscriptional regulation of urokinase receptor gene expression in human lung carcinoma and mesothelioma cells in vitro (1999)
    Springer
    Molecular and cellular biochemistry 199 (1999), S. 189-200 
    Details
    ISSN: 1573-4919
    Keywords: lung ; cancer ; urokinase ; receptor ; gene expression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract The urokinase-type plasminogen activator (uPA) interacts with its receptor (uPAR) to promote proteolysis as well as cell proliferation and migration. These functions contribute to the pathogenesis of neoplastic growth and invasiveness. Expression of uPAR in tumor extracts also inversely correlates with prognosis in many forms of cancer. In this study, we sought to determine if differences in uPAR expression were distinguishable between cultured human lung carcinoma and malignant mesothelioma subtypes. We also sought to determine if, as in malignant mesothelioma cells, uPAR expression is regulated at the posttranscriptional level in cultured malignant lung carcinoma cells. Using 125I-uPA binding and ligand blotting techniques, uPAR was expressed by phenotypically diverse lung carcinoma cell lines, including the H460, H157 and H1395 non-small cell lines and the H146 small cell lung carcinoma line. Increased uPAR expression was also detected in spindle-shaped (M33K) and epithelioid (M9K and MS-1) malignant mesothelioma cells. Selected mediators, including TGF-β, TNF-α, LPS and PMA, uniformly enhanced uPAR expression in each of the tumor cell lines. Steady state uPAR mRNA expression was determined by RNase protection assay and correlated directly with the changes in cell surface uPAR expression. By gel mobility shift and UV-cross linking assays, a uPAR mRNA binding protein (uPAR mRNABp) implicated in the posttranscriptional control of message stability, was identified in each of the cell lines. Expression of uPAR and its message in cultured lung carcinoma and malignant mesothelioma cells is similarly influenced by effectors present in the tumor microenvironment. Regulation of the uPAR message occurs at the posttranscriptional level in cultured small and non-small cell lung carcinoma cells as well as spindle-shaped and fibrous malignant mesothelioma cell lines. Posttranscriptional regulation of uPAR in all these cells involves the interaction of the uPAR mRNABp with uPAR mRNA, which promotes uPAR mRNA destabilization.
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    URL: http://dx.doi.org/10.1023/A:1006914800447
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    Transcriptional modulation of the human complement factor I gene in Hep G2 cells by protein kinase C activation (1999)
    Springer
    Molecular and cellular biochemistry 201 (1999), S. 111-123 
    Details
    ISSN: 1573-4919
    Keywords: complement factor I ; TPA ; protein kinase C ; gene expression ; Hep G2 cells
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract This study examined the role of the protein kinase C (PKC) signalling pathway in the regulation of expression of human complement factor I (CFI) gene. The production of CFI by Hep G2 cells was enhanced in a dose- and time-dependent fashion by 12-O-tetradecanoyl-1,2-phorbol 13-acetate (TPA), a potent PKC activator. 4α-phorbol didecanoate, an inactive phorbol ester, had no effect on CFI synthesis. The TPA-dependent increase in CFI secretion was correlated with an increase in CFI mRNA levels. Forskolin, a cAMP-inducing agent, augmented the TPA response. W7, an inhibitor of protein kinase A and genistein, an inhibitor of protein tyrosine kinase(s) both did not prevent the increase in CFI expression mediated by TPA. However, calphostin C, a specific inhibitor of PKC, abolished the TPA-induced increase in CFI mRNA levels. Down regulation of intracellular PKC levels by prior exposure of Hep G2 cells to a high concentration of TPA also blocked the increase in CFI mRNA levels induced by TPA suggesting that the TPA effects were mediated via activation of PKC. mRNA decay studies indicated that the half-life of CFI mRNA in TPA-induced cells was not significantly different from control. Nuclear run-on transcriptional assays on the other hand demonstrated that whereas the CFI gene is transcribed under basal conditions in Hep G2 cells, TPA induced a 3-4 fold increase in the transcription rate of CFI gene in 24 h. The transcription rate of GAPDH gene did not change, indicating that the effects were not general on gene transcription. Transient transfections of Hep G2 cells with chloramphenicol acetyltransferase reporter gene (CAT) constructs containing a series of sequential 5′ deletions of the CFI promoter and CAT assays showed that the sequence between -136 and -130, containing an AP-1 consensus sequence (TGAGTCA) was required for the TPA response. This observation was substantiated by the finding that mutation of this AP-1 site to TttaTCA or TtAtcCA abolished the TPA responsiveness. The enhancement of the activity of transfected chimeric CAT constructs by TPA was abrogated by calphostin C and by pyrrolidine dithiocarbamate (an inhibitor of NF-κB and AP-1 transactivation). These results indicate that TPA regulation of CFI gene requires PKC signalling and is mediated by via a TPA response element (TRE) in the CFI promoter region located at -136/-130 and involves the transactivation of AP-1 and NF-κB transcription factors. We suggest that PKC may be one of the intracellular pathways that control CFI gene expression and that cellular processes (involving growth factors, hormones, cytokines etc.) that activate PKC may upregulate the expression of the CFI gene.
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    URL: http://dx.doi.org/10.1023/A:1007064602321
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    Expression of calcium-binding protein regucalcin and microsomal Ca2+-ATPase regulation in rat brain: Attenuation with increasing age (1999)
    Springer
    Molecular and cellular biochemistry 200 (1999), S. 43-49 
    Details
    ISSN: 1573-4919
    Keywords: regucalcin ; calcium-binding protein ; gene expression ; Ca2+-ATPase ; brain microsomes ; aging
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract The expression of calcium-binding protein regucalcin and its effect on the microsomal Ca2+-ATPase activity in rat brain tissues was investigated. The expression of regucalcin mRNA was demonstrated by reverse transcription-polymerase chain reaction (RT-PCR) analysis in brain tissues using rat regucalcin-specific primers. Regucalcin concentration in the brain tissues was about 5 × 10-9 M as measured using enzyme-linked immunoadsorbent assay (ELISA), and this level was lowered with increasing age (50 weeks old). The presence of regucalcin (10-9 to 10-7 M) in the enzyme reaction mixture caused a significant decrease in Ca2+-ATPase activity in the brain microsomes of young rats (5 weeks old). Meanwhile, the enzyme activity was not significantly altered by the addition of calmodulin (1 or 50 μg/ml), calbindin (1 or 10 μg/ml), and S-100 A protein (5 or 25 μg/ml), which are other Ca2+-binding proteins in rat brain. The effect of regucalcin to inhibit microsomal Ca2+-ATPase activity was weakened in the brain of rats with increasing age (50 weeks old). The present study demonstrates that regucalcin is expressed in the brain, and that it can uniquely inhibit Ca2+-ATPase activity in the brain microsomes of rats. The findings suggest that regucalcin plays a role in the regulation of microsomal Ca2+-ATPase activity in rat brain tissues.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006928402184
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    Electronic Resource
    Identification of stretch-responsive genes in pulmonary artery smooth muscle cells by a two arbitrary primer-based mRNA differential display approach (1999)
    Springer
    Molecular and cellular biochemistry 197 (1999), S. 87-96 
    Details
    ISSN: 1573-4919
    Keywords: mechanical stretch ; smooth muscle cells ; differential display ; gene expression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract Physical forces induce profound changes in cell phenotype, shape and behavior. These changes can occur in vascular structures as a result of pressure overload and their effects can be seen in atherosclerotic vessels in which smooth muscle cells have undergone hyperplastic and hypertrophic changes. At the molecular level, mechanical stimuli are converted into chemical ones and lead to modulation of gene expression and/or the activation of a new repertoire of genes whose encoded proteins help the cells to adapt to their microenvironment. In this study, we have used a two primer-based mRNA differential display technique to identify candidate mechano-responsive genes in pulmonary artery smooth muscle cells. As compared to the original method described by Liang and Pardee, this technique uses two arbitrary primers instead of an anchored oligo(dt) plus an arbitrary primer in the polymerase chain reaction. The chief advantages of these modifications are an increase in the efficiency of the amplification and in the identification of differentially expressed clones. Using this approach, we compared the pattern of expressed genes in cells cultured under static conditions with those in cells that were mechanically stretched (1 Hz) for 24 h in a well-defined in vitro mechanical system. Three candidate genes that showed reproducible differences were chosen for further characterization and cloning. One clone was under expressed in stretched cells and had a DNA sequence with 90% homology to the human fibronectin gene. Two other clones were highly expressed in stretched cells and had a 92% and a 83% sequence homology with human platelet-activating factor (PAF) receptor and rat insulin-like growth factor-I (IGF-I) genes respectively. Northern blot analysis confirmed low levels of fibronectin mRNA transcripts in stretched cells. In contrast, accumulation of PAF receptor mRNA occurred 30 min after mechanical stretch was initiated whereas IGF-I mRNA levels peaked at 8 h. Both mRNA levels were sustained for up to 24 h of mechanical stretching. These results demonstrate the usefulness of the two primer-based mRNA differential display that enabled us to identify and characterize alterations at the level of gene expression among matrix proteins, G-protein coupled receptors and growth factors, each of whose response to mechanical strain is different. A more complete understanding of these responses will provide further insight into the pathologic processes associated with hypertension and atherosclerosis.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006966530553
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  • 68
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    Electronic Resource
    Characterisation of genes isolated from a potato swellingstolon cDNA library (1999)
    Springer
    Potato research 42 (1999), S. 31-42 
    Details
    ISSN: 1871-4528
    Keywords: Solanum tuberosum L. ; tuberisation ; extensin ; acyl carrier protein thioesterase ; high mobility group protein ; gene expression ; plant development
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Summary In screening to isolate a full-length copy of a previously isolated cDNA clone, a further three cDNAs were also isolated from a library prepared from sub-apical swelling-stolon tissue of potato (Solanum tuberosum L.). Sequence analysis showed these clones to be similar to extensin-like protein genes, acyl carrier protein thioesterase genes and high mobility group protein genes, respectively. A further cDNA, isolated by subtractive hybridisation, was similar to a tomato cDNA previously isolated on the basis of its down-regulation following nematode infection. While all the newly isolated genes were expressed in swelling stolons, for most, maximal expression was seen to be in stem tissue. Possible roles for these genes in the development of potato plants are discussed, as is the significance of gene expression in stems and stolons to the process of tuberisation.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF02358389
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  • 69
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    Electronic Resource
    Introduction of a Proximal Stat5 Site in the Murine α-Lactalbumin Promoter Induces Prolactin Dependency In Vitro and Improves Expression Frequency In Vivo (1999)
    Springer
    Transgenic research 8 (1999), S. 23-31 
    Details
    ISSN: 1573-9368
    Keywords: transgenic mice ; prolactin ; mammary gland ; gene expression ; Stat5 ; β-globin insulator
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract In order to establish a possible correlation between in vitro prolactin induction and the transcriptional activity of mammary gene promoters in transgenic mice, a functional Stat5-binding site was created by means of site-directed mutagenesis at position −70 on a 560 bp murine α-lactalbumin promotor linked to a CAT reporter gene. Surprisingly, the wild-type promoter was constitutively active in vitro and could not be induced by prolactin. Introducing the proximal Stat5 site abolished this constitutive activity and resulted in prolactin dependence in both CHO-K1- and HC11-transfected cells. In transgenic mice, both the frequency of lines expressing the transgene and the prevalence of mid to late pregnancy expression were increased.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1008851802022
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  • 70
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    Electronic Resource
    Quantitative analysis of transcription and translation in gene amplified Chinese hamster ovary cells on the basis of a kinetic model (1999)
    Springer
    Cytotechnology 29 (1999), S. 93-102 
    Details
    ISSN: 1573-0778
    Keywords: CHO cells ; gene expression ; kinetic model ; protein secretion ; transcription ; translation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract The elevation of expression levels for secreted glycoproteins by gene amplification in mammalian cells shows a saturation behavior at high levels of gene amplification. At high expression levels a drop in the secretion efficiency for the recombinant protein occurs (Schröder and Friedl, 1997), coinciding with the appearance of misfolded protein in the cell. In this communication we investigated whether additional limitations exist at the levels of transcription and translation. Four Chinese hamster ovary (CHO) cell lines expressing different amounts of human antithrombin III (ATIII) were used as a model system. A tenfold increase in the ATIII cDNA copy number from the lowest to the highest producing cell line coincided with a 38-fold increase in ATIII mRNA levels, and an 80-fold increase in the amount of intracellular ATIII levels. The data was analyzed using a simple kinetic model. The following conclusions were derived: I. The transcriptional activity for the recombinant protein is not saturated. II. Translation itself is not saturated either, but may be downregulated as secretion efficiency drops. III. Two explanations for the previously reported drop in secretion efficiency for the recombinant protein with increasing expression level are possible: A. Protein degradation is an alternative fate for translated ATIII and the fraction of ATIII degraded after translation increases as expression level is increased. B. Translation is downregulated as the secretory apparatus becomes exhausted to maintain cell viability.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1008077603328
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  • 71
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    Electronic Resource
    Embryonic stem cell differentiation models: cardiogenesis, myogenesis, neurogenesis, epithelial and vascular smooth muscle cell differentiation in vitro (1999)
    Springer
    Cytotechnology 30 (1999), S. 211-226 
    Details
    ISSN: 1573-0778
    Keywords: cardiogenesis ; cell differentiation ; gene expression ; mouse embryonic stem cells ; myogenesis ; neurogenesis
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract Embryonic stem cells, totipotent cells of the early mouse embryo, were established as permanent cell lines of undifferentiated cells. ES cells provide an important cellular system in developmental biology for the manipulation of preselected genes in mice by using the gene targeting technology. Embryonic stem cells, when cultivated as embryo-like aggregates, so-called ‘embryoid bodies’, are able to differentiate in vitro into derivatives of all three primary germ layers, the endoderm, ectoderm and mesoderm. We established differentiation protocols for the in vitro development of undifferentiated embryonic stem cells into differentiated cardiomyocytes, skeletal muscle, neuronal, epithelial and vascular smooth muscle cells. During differentiation, tissue-specific genes, proteins, ion channels, receptors and action potentials were expressed in a developmentally controlled pattern. This pattern closely recapitulates the developmental pattern during embryogenesis in the living organism. In vitro, the controlled developmental pattern was found to be influenced by differentiation and growth factor molecules or by xenobiotics. Furthermore, the differentiation system has been used for genetic analyses by ‘gain of function’ and ‘loss of function’ approaches in vitro.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1008041420166
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  • 72
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    Electronic Resource
    Transient gene expression in mammalian cells grown in serum-free suspension culture (1999)
    Springer
    Cytotechnology 30 (1999), S. 71-83 
    Details
    ISSN: 1573-0778
    Keywords: gene expression ; HEK293(EBNA) cells ; serum-free ; transient transfection
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract In order to establish a simple and scaleable transfection system we have used the cationic polymer polyethylenimine (PEI) to study transient transfection in HEK293 and 293(EBNA) cells grown in serum-free suspension culture. The transfection complexes were made directly within the cell culture by consecutively adding plasmid and PEI (direct method). Alternatively, the DNA-PEI transfection complexes were prepared in fresh medium (1/10 culture volume) and then added to the cells (indirect method). The results of this study clearly show that the ratio of PEI nitrogen to DNA phosphate is very important for high expression levels. The precise ratio is dependent on the DNA concentration. For example, using 1 μg/ml DNA by the indirect method, the ratio of optimal PEI:DNA was about 10–13:1. However, the ratio increases to 33:1 for 0.1–0.2 μg/ml DNA. By testing several different molecular weights of the polycationic polymer we could show that the highest transfection efficiency was obtained with the PEI 25 kDa. Using PEI 25 kDa the indirect method is superior to the direct addition because significantly lower DNA concentrations are needed. The expression levels of the soluble human TNF receptor p55 are even higher at low DNA compared to 1 μg/ml plasmid. The EBV-based pREP vectors gave better transient gene expression when used in 293(EBNA) cells compared to HEK293 cells in suspension culture. No differences in expression levels in the two cell lines were observed when the pC1 (CMV)-TNFR was used. In conclusion, PEI is a low-toxic transfection agent which provides high levels of transient gene expression in 293(EBNA) cells grown in serum-free suspension culture. This system allows highly reproducible, cost-effective production of milligram amounts of recombinant proteins in 2–5 l spinner culture scale within 3–5 days. Fermentor scale experiments, however, are less efficient because the PEI-mediated transient tranfection is inhibited by conditioned medium.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1008000327766
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  • 73
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    Electronic Resource
    The role of the cell cycle in determining gene expression and productivity in CHO cells (1999)
    Springer
    Cytotechnology 30 (1999), S. 49-57 
    Details
    ISSN: 1573-0778
    Keywords: cell cycle ; CHO ; flow cytometry ; gene expression ; synchronisation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract Understanding the relationships between cell cycle and protein expression is critical to the optimisation of media and environmental conditions for successful commercial operation of animal cell culture processes. Using flow cytometry for the analysis of the early phases of synchronised batch cultures, the dependency of product expression on cell cycle related events has been evaluated in a recombinant CHO cell line. Although the production of recombinant protein is initially found to be cell cycle related, the maximum specific protein productivity is only achieved at a later stage of the exponential phase which also sees a maximum in the intracellular protein concentration. Subsequent work suggests that it is the batch phase/medium composition of cultures which is the major determinant of maximum specific productivity in this cell line. Furthermore the effect of the positive association between S phase and specific productivity is subordinate to the effect of batch phase/medium composition on the specific productivity of batch cultures.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1008093404237
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  • 74
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    Electronic Resource
    Differential expression of RAB5A in human lung adenocarcinoma cells with different metastasis potential (1999)
    Springer
    Clinical & experimental metastasis 17 (1999), S. 213-219 
    Details
    ISSN: 1573-7276
    Keywords: gene expression ; immunohistochemistry ; mRNA DD ; neoplasia metastasis ; RAB5A
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract For the sake of better understanding the molecular mechanism of neoplasia, we have used the mRNA differential display technique to analyze two human lung adenocarcinoma cell lines, AGZY83-a and Anip973. Anip973 was isolated from AGZY83-a, but manifested much higher metastatic potential than the parent line. We found that a significant differential cDNA fragment in Anip973 was over-expressed, then over-expressed cDNA fragment was cloned and sequenced. It showed that the over-expressed cDNA in Anip973 was RAB5A cDNA. And the RAB5A cDNA sequence was corresponding between the two cells. To determine whether RAB5A may be differentially expressed in the two human lung adenocarcinoma cells at protein level, we further detected RAB5A protein in the two cells by using immunofluorescent method. RAB5A protein was upregulated in highly metastatic Anip973. We also detected the difference in RAB5A gene expression at RNA level in human non-small cell lung carcinoma by RT-PCR. Using immunohistochemical staining, we also examined RAB5A change at protein level in 45 cases human non-small cell lung carcinoma paraffin sections. The results proved the evidence of upregulation of RAB5A in malignant tumor, indicated over-expression of RAB5A gene was correlated with the malignant degree and metastatic potential of lung cancer(χ2 test, p 〈0.01). The RAB5A gene is a member of RAS superfamily, which can transcribe GTP-binding protein that plays an important role in signal transduction of protein trafficking at the cell surface and GDP/GTP cycle in the regulation of endocytotic membrane traffic. Thus our results indicated that over-expression of the RAB5A gene was involved in the process of transformation from AGZY83-a to the higher metastatic cell line Anip973. The result may be a powerful experimental evidence that over-expression of RAB5A gene associated with neoplasia metastasis.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006617016451
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  • 75
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    Electronic Resource
    Effect of Portacaval Anastomosis on Glutamine Synthetase Protein and Gene Expression in Brain, Liver and Skeletal Muscle (1999)
    Springer
    Metabolic brain disease 14 (1999), S. 273-280 
    Details
    ISSN: 1573-7365
    Keywords: glutamine synthetase ; gene expression ; portacaval anastomosis ; hepatic encephalopathy ; liver ; skeletal muscle ; ammonia
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract The effects of chronic liver insufficiency resulting from end-to-side portacaval anastomosis (PCA) on glutamine synthetase (GS) activities, protein and gene expression were studied in brain, liver and skeletal muscle of male adult rats. Four weeks following PCA, activities of GS in cerebral cortex and cerebellum were reduced by 32% and 37% (p〈0.05) respectively whereas GS activities in muscle were increased by 52% (p〈0.05). GS activities in liver were decreased by up to 90% (p〈0.01), a finding which undoubtedly reflects the loss of GS-rich perivenous hepatocytes following portal-systemic shunting. Immunoblotting techniques revealed no change in GS protein content of brain regions or muscle but a significant loss in liver of PCA rats. GS mRNA determined by semi-quantitative RT-PCR was also significantly decreased in the livers of PCA rats compared to sham-operated controls. These findings demonstrate that PCA results in a loss of GS gene expression in the liver and that brain does not show a compensatory induction of enzyme activity, rendering it particularly sensitive to increases in ammonia in chronic liver failure. The finding of a post-translational increase of GS in muscle following portacaval shunting suggests that, in chronic liver failure, muscle becomes the major organ responsible for the removal of excess blood-borne ammonia.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1020741226752
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  • 76
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    Electronic Resource
    Differentiation Dependent Activation of the Myelin Genes In Purified Oligodendrocytes is Highly Resistant to Hypoglycemia (1999)
    Springer
    Metabolic brain disease 14 (1999), S. 189-195 
    Details
    ISSN: 1573-7365
    Keywords: Oligodendrocyte cultures ; glucose ; gene expression ; malic enzyme ; membrane synthesis ; myelination ; undernutrition
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract We have previously demonstrated that the developmental upregulation of myelin-specific genes in mixed glial cultures is strongly attenuated by hypoglycemia. The present study was designed to evaluate the effect of hypoglycemia on differentiation-dependent upregulation of myelin genes in purified oligodendrocyte cultures. The expression of major myelin protein genes, i.e., proteolipid protein (PLP), basic protein (BP) and myelin associated glycoprotein (MAG) were monitored by Northern blot analysis. In control cultures maintained at 6 mg/ml of glucose, the expression of all the genes upregulated rapidly, and plateaued at approximately day 4. A similar pattern of differentiation-dependent upregulation was observed for the gene encoding a lipogenic enzyme, i.e., malic enzyme (ME). In contrast to mixed glial cultures, however, this developmental gene upregulation was not significantly affected by severe hypoglycemia (approximately 0.02 mg/ml). The results indicate that the effect of glucose deprivation on oligodendrocyte genes observed in mixed glial cultures is mediated by other cells. The upregulation of the genes in differentiating oligodendrocytes was accompanied by the production of myelin-related membrane that was isolated by density gradient fractionation. In contrast to the effect on gene expression, this anabolic activity was highly dependent on glucose, as seen from a profound suppression by severe hypoglycemia.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1020614809546
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  • 77
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    Electronic Resource
    Genetic Variation in Spiroplasma citri (1999)
    Springer
    European journal of plant pathology 105 (1999), S. 519-533 
    Details
    ISSN: 1573-8469
    Keywords: genome ; gene expression ; mollicute ; recombination ; transposition ; virus
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract Spiroplasmas are members of the Class Mollicutes, wall-less prokaryotes having a high adenosine–thymidine content in their small genomes. Spiroplasma citri is a plant pathogen that inhabits phloem. Like other phytopathogenic spiroplasmas and the related phytoplasmas, it is transmitted from plant to plant by phloem-feeding leafhoppers that serve as alternate hosts for the spiroplasma as well as vectors. Genetic information in spiroplasmas is carried on a circular chromosome, on plasmids and/or in virus genomes. A picture emerging from recent research on the S. citri genome is one of frequent and often extensive variation, resulting from a number of different mechanisms. Expansion and contraction events must continually be occurring in about equal proportions so that the net genome size varies within defined boundaries. Particularly impressive are large changes in genome size that can occur in only a few generations. As with most organisms, genetic variation in S. citri results from variation in extrachromosomal DNA content, changes due to DNA replication and repair processes and changes due to recombination. The implied flux of genetic information into and out of the S. citri genome should be beneficial to the bacterium, allowing it, with its small genome size, to adapt to new environments.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1008757716803
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  • 78
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    Electronic Resource
    Evidence of Mucin Secretion in Human Lung Adenocarcinoma Cell Lines NCIH650 and NCIH2077 and Effect of Select Secretagogues on Mucin Secretion (1999)
    Springer
    Bioscience reports 19 (1999), S. 473-483 
    Details
    ISSN: 1573-4935
    Keywords: Mucin ; lung cancer ; gene expression ; secretion ; lung adenocarcinoma
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Abstract Mucins comprise an important class of tumor-associated antigens. The objectives of the present study were (a) to establish an in vitro model system using human non-small cell lung adenocarcinoma cell lines NCIH650 and NCIH2077 (b) provide evidence that these cell lines secrete mucin in culture conditions and (c) investigate the effects of select secretagogues on mucin secretion. The cell lines were established in ACL-4 medium containing several growth factors and retinoic acid and 5% fetal calf serum. The high molecular weight glycoconjugates secreted in the culture medium were purified by ammonium sulfate precipitation and Superose 6 and Superose 12 FPLC chromatography. The purified high molecular weight glycoconjugate fraction and the carcinoma cells were shown to have mucin by dot blot, Western blot and immunohistochemical analysis, respectively, using specific antibodies to purified major mucin, HTM-1. Also, incorporation experiments with mucin precursor 3H-glucosamine demonstrated that the cells indeed synthesize high molecular weight mucins. The effects of secretagogues such as, 8-bromocyclic AMP, ionomycin, phorbol-12-myristate-13-acetate and neutrophil elastase on mucin secretion were also investigated. Only 8-bromocyclic AMP and neutrophil elastase influenced mucin secretion. These studies provided strong evidence that the lung adenocarcinoma cell lines secrete high molecular weight mucins in culture conditions and only two of the four tested secretagogues significantly increased mucin secretion. Thus, this in vitro model system may be useful in determining alterations in mucin structure, if any, in lung adenocarcinomas as well as in studying the regulation of mucin gene expression.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1020224625088
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  • 79
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    Comparison of the Anticancer Effect of Free and HPMA Copolymer-Bound Adriamycin in Human Ovarian Carcinoma Cells (1999)
    Springer
    Pharmaceutical research 16 (1999), S. 986-996 
    Details
    ISSN: 1573-904X
    Keywords: adriamycin ; doxorubicin ; HPMA copolymer ; apoptosis, multidrug resistance ; gene expression ; signal transduction
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Abstract Purpose. To study peculiarities and the mechanism of the anticancer effect of free and HPMA copolymer-bound ADR in sensitive and resistant human ovarian carcinoma cells. Methods. Sensitive A2780 and ADR resistant A2780/AD cells were exposed to different doses of drugs during 12, 24, 36, 48, 60, and 72 hours. Cell viability, drug accumulation, apoptosis, cellular metabolism, lipid peroxidation, DNA content and gene expression were studied. Results. HPMA copolymer-bound ADR (P(GFLG)-ADR) possessed a comparable cytotoxicity to free ADR when comparison was based on intracellular concentrations. While free ADR up-regulated genes encoding ATP driven efflux pumps (MDR1, MRP), P(GFLG)-ADR overcame existing pumps and down regulated the MRP gene. Free ADR also activated cell metabolism and expression of genes responsible for detoxification and DNA repair. P(GFLG)-ADR down-regulated HSP-70, GSr-π, BUDP, Topo-IIα, β, and TK-1 genes. Apoptosis, lipid peroxidation and DNA damage were significantly higher after exposure to P(GFLG)-ADR, as reflected by simultaneous activation of p53, c-fos in A2780 cells) or c-jun (A2780/AD) signaling pathways and inhibition of the bcl-2 gene. Differences between free ADR and P(GFLG)-ADR increased with the time of incubation and drug concentration. Conclusions. P(GFLG)-ADR overcame drug efflux pumps, more significantly induced apoptosis and lipid peroxidation, inhibited DNA repair, replication, and biosynthesis when compared to free ADR.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1018959029186
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  • 80
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    Electronic Resource
    Enhanced expression and differential inducibility of soybean chalcone synthase genes by supplemental UV-B in dark-grown seedlings (1999)
    Springer
    Plant molecular biology 39 (1999), S. 785-795 
    Details
    ISSN: 1573-5028
    Keywords: UV-B ; soybean ; chalcone synthase ; gene expression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract By developing gene-specific RT-PCR and using filters to allow transmission down to 290 nm (UV-B+) or blocking all radiation below 320 nm (UV-B−), the effect of UV-B+ and UV-B− light on expression of each of the presently known seven members of soybean chalcone synthase (CHS) gene family in dark-grown seedlings was analyzed. Dark expression was detectable already in 18 h dark-germinating embryos, with progressive increases on successive days, suggesting that chs belongs to a class of genes expressed very early during germination, and that the expression at this stage is either constitutive or induced by non-light-dependent factors present in the seed or made available following imbibition. Exposure of 18 h dark-germinating embryos to UV-B− or to UV-B+ light did not lead to an increase in chs signal. However, the 24 h dark-germinating embryos showed a distinct effect of UV-B+, interestingly coinciding with the stage when the head of seedlings was in the process of being pushed up above ground by stem elongation, suggesting the possibility of a developmental switch modulating the appearance of UV-B response. The response to UV-B− was most prominent in chs1 and almost silent in chs2, while the up-regulation by UV-B+ was most prominent in chs5 and chs6 and much less so in chs2. Interestingly, chs2 was noted to be the only member of the Gmchs gene family devoid of H-box, raising the possibility that the H-box may be a good indicator of the photo-inducibility of a chs gene.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006124219945
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  • 81
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    Complexity and expression of the glutamine synthetase multigene family in the amphidiploid crop Brassica napus (1999)
    Springer
    Plant molecular biology 39 (1999), S. 395-405 
    Details
    ISSN: 1573-5028
    Keywords: amphidiploid genome structure ; gene expression ; glutamine synthetase ; multigene family ; nitrogen assimilation ; oilseed rape
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract In the amphidiploid genome of oilseed rape (Brassica napus) the diploid ancestral genomes of B. campestris and B. oleracea have been merged. As a result of this crossing event, all gene loci, gene families, or multigene families of the A and C genome types encoding a certain protein are now combined in one plant genome. In the case of the multigene family for glutamine synthetase, the key enzyme of nitrogen assimilation, six different cDNA sequences were isolated from leaf and root specific libraries. One sequence pair (BnGSL1/BnGSL2) was characterized by the presence of amino- terminal transit peptides, a typical feature of all nuclear encoded chloroplast proteins. Two other cDNA pairs (BnGSR1-1/BnGSR1-2 and BnGSR2-1/BnGSR2-2) with very high homology between each other were found in a root specific cDNA library and represent protein subunits for cytosolic glutamine synthetase isoforms. Comparative PCR amplifications of genomic DNA isolated from B. napus, B. campestris and B. oleracea followed by sequence–specific restriction analyses of the PCR products permitted the assignment of the cDNA sequences to either the A genome type (BnGSL1/BnGSR1- 1/BnGSR2-1) or the C genome type (BnGSL2/BnGSR1-2/BnGSR2-2). Consequently, the ancestral GS genes of B. campestris and B. oleracea are expressed simultaneously in oilseed rape. This result was also confirmed by RFLP (restriction fragment length polymorphism) analysis of RT-PCR products. In addition, the different GS genes showed tissue specific expression patterns which are correlated with the state of development of the plant material. Especially for the GS genes encoding the cytosolic GS isoform BnGSR2, a marked increase of expression could be observed after the onset of leaf senescence.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006193717093
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  • 82
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    Diverse range of gene activity during Arabidopsis thaliana leaf senescence includes pathogen-independent induction of defense-related genes (1999)
    Springer
    Plant molecular biology 40 (1999), S. 267-278 
    Details
    ISSN: 1573-5028
    Keywords: defense ; gene expression ; leaf senescence ; nitrilase ; pathogen-free ; salicylic acid
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract To determine the range of gene activities associated with leaf senescence, we have identified genes that show preferential transcript accumulation during this developmental stage. The mRNA levels of a diverse array of gene products increases during leaf senescence, including a protease, a ribosomal protein, two cinnamyl alcohol dehydrogenases, a nitrilase and glyoxalase II. Two of the genes identified are known to be pathogen-induced. The senescence specificity of each gene was determined by characterization of transcript accumulation during leaf development and in different tissues. The increased expression of nitrilase in senescent leaves is paralleled by an increase in free indole-3-acetic acid (IAA) levels. Additionally, we have demonstrated that the induction of defense-related genes during leaf senescence is pathogen-independent and that salicylic acid accumulation is not essential for this induction. Our data indicate that the induction of certain genes involved in plant defense responses is a component of the leaf senescence program.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006199932265
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  • 83
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    Isolation and molecular characterization of gibberellin-regulated H1 and H2B histone cDNAs in the leaf of the gibberellin-deficient tomato (1999)
    Springer
    Plant molecular biology 39 (1999), S. 883-890 
    Details
    ISSN: 1573-5028
    Keywords: gene expression ; gibberellin ; H1 histone ; H2B histones ; leaf ; Lycopersicon esculentum
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract After differential screening we isolated cDNA clones encoding a histone H1 (leH1) and three variants of histone H2B (leH2B-1, -2 and -3) from the gibberellin (GA)-deficient mutant of tomato (gib-1). The deduced polypeptide of leH1 is 271 amino acids long and exhibits the typical tripartite structure of histones H1. The full-length cDNA clone leH2B-1 encodes for a protein of 142 amino residues and shows the tripartite organization of histones H2B. The histones leH1 and leH2B, which show no tissue specificity, are developmentally expressed in the leaf. The mRNA accumulation was higher in organs which contain meristematic tissue and/or which have a high proportion of actively cycling cells. In the leaf of the gib-1 mutant we demonstrated GA-enhanced histone leH1 and leH2B expression which was not observed in the wild type. GAs of the early-13-hydroxylated pathway (GA1 and GA3) caused most enhanced transcription compared to GAs of the early-non-hydroxylation pathway (GA4 and GA9). Application of GA to the mutant increased histone expression that could correlate with enhanced DNA replication in leaf tissue. Increased chromosome replication may indicate that there is a higher rate of cell division and/or increase of endopolyploidy which both may be dependent on cell elongation induced by GAs.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006157718263
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    Differential expression of two spermidine synthase genes during early fruit development and in vegetative tissues of pea (1999)
    Springer
    Plant molecular biology 39 (1999), S. 933-943 
    Details
    ISSN: 1573-5028
    Keywords: cloning ; fruit development ; gene expression ; pea ; polyamine ; spermidine synthase
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Two cDNAs from young pea fruits coding for functional spermidine synthases (EC 2.5.1.16) were isolated. The corresponding genes were named psSPDSYN1 and psSPDSYN2. Both cDNAs complemented spe3Δ gene when introduced into the Y480 strain of Saccharomyces cerevisiae, which is a null mutant for the spermidine synthase gene. psSPDSYN1 and psSPDSYN2 are regulated differentially. psSPDSYN1 is up-regulated early after fruit set whereas psSPDSYN2 is expressed later. Spermidine synthase activity was detected in pea ovaries, and correlates with the pattern of expression of psSPDSYN1. In the pea plant, psSPDSYN1 is highly expressed in actively growing tissues, whereas the highest level of psSPDSYN2 mRNA was detected in fully elongated stem.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006158819849
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  • 85
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    Regulation of the chitinase gene expression in suspension-cultured rice cells by N-acetylchitooligosaccharides: differences in the signal transduction pathways leading to the activation of elicitor-responsive genes (1999)
    Springer
    Plant molecular biology 39 (1999), S. 907-914 
    Details
    ISSN: 1573-5028
    Keywords: chitin oligomer ; chitinase ; elicitor ; gene expression ; rice ; signal transduction
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Expression patterns of chitinase transcripts induced by N-acetylchitooligosaccharide elicitor were analyzed by northern blot hybridization in order to reveal a signal transduction pathway leading to the activation of class I chitinase genes (Cht-1 and Cht-3), which may play an important role in producing N-acetylchitooligosaccharide elicitor. The transcription level of both genes was enhanced in response to N-acetylchitooligosaccharides larger than pentaose at subnanomolar concentrations. These structure and dose dependencies were consistent not only with those for a 75 kDa high-affinity binding protein for N-acetylchitooligosaccharide elicitor in the plasma membrane, but also with other series of cellular responses including phytoalexin production and the expression of elicitor-responsive genes (EL2, EL3). Therefore, the elicitor signal to evoke these cellular responses including the activation of the chitinase genes could be common and transmitted into cells through the 75 kDa protein. However, the signal transduction pathway for the activation of the chitinase gene appeared to diverge from those for the other elicitor-responsive genes shortly after the signal perception. It was shown that the induction of chitinase expression by N-acetylchitooligosaccharide would require protein phosphorylation, but not de novo protein synthesis. The oxidative burst was demonstrated not to be necessary for transcriptional induction of the all four elicitor-responsive genes (Cht, PAL, EL2, EL3) by N-acetylchitooligosaccharide.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006161802334
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  • 86
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    Temporal and spatial gene expression of cytochrome B5 during flower and fruit development in olives (1999)
    Springer
    Plant molecular biology 40 (1999), S. 79-90 
    Details
    ISSN: 1573-5028
    Keywords: cytochrome b5 ; fruit and flower development ; gene expression ; in situ hybridisation ; olive
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract We report the characterisation of two cytochrome b5 genes and their spatial and temporal patterns of expression during development in olive, Olea europaea. A PCR-generated probe, based on a tobacco cytochrome b5 sequence, was used to isolate two full-length cDNA clones (cytochrome b5-15 and cytochrome b5-38) from a library derived from 13 WAF olive fruits. The cDNAs encoded proteins of 17.0 and 17.7 kDa, which contained all the characteristic motifs of cytochromes b5 from other organisms and exhibited 63% identity and 85% similarity with each other. The olive cytochrome b5-15 cDNA was then used as a probe for more detailed analysis. Southern blotting revealed a gene family of at least 4–6 members while northern blotting and in situ hybridisation showed a highly specific pattern of gene expression. Very low levels of cytochrome b5 mRNA were detected in tissues characterised by high rates of lipid accumulation, such as young expanding leaves, maturing seeds and ripening mesocarp. The cytochrome b5 genes were not induced at 6 °C and their response to ABA was relatively slow compared with fatty acid desaturase genes. In contrast, high levels of cytochrome b5 gene expression were found in young fruits at the pattern formation (globular/heart) stage of embryogenesis and in vascular and transmitting tissues of male and female reproductive organs. The data are consistent with a major role for cytochrome b5 in developmental processes related to plant reproduction in addition to being an electron donor to microsomal desaturases.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1026417710320
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  • 87
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    Cloning and characterization of a trypsin inhibitor cDNA from amaranth (Amaranthus hypochondriacus) seeds (1999)
    Springer
    Plant molecular biology 41 (1999), S. 15-23 
    Details
    ISSN: 1573-5028
    Keywords: Amaranthus hypochondriacus) ; gene expression ; trypsin inhibitor
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract We previously isolated and sequenced the major trypsin inhibitor from Amaranthus hypochondriacus seeds. This amaranth trypsin inhibitor (AmTI) is a 69 amino acid protein with high homology to members of the potato-1 inhibitor family. This paper describes the cloning and expression of a cDNA encoding this trypsin inhibitor in various vegetative tissues of the amaranth plant during seed development and imbibition, and investigates the possible induction of AmTI expression by wounding. We obtained a 393 bp cDNA sequence with an open reading frame corresponding to a polypeptide with 76 amino acid residues. With the exception of one residue (Ser-41), the polypeptide agrees with the amino acid sequence previously reported, plus 7 more residues at the N-terminus. These N-terminal residues are thought to be part of the signal used for intracellular sorting. The organ specificity of AmTI gene expression was investigated by northern analysis, showing that mRNA corresponding to AmTI genes was present in stems of plants growing under normal conditions. The kinetics of accumulation of the AmTI-mRNA, protein, and inhibitory activity during seed development and imbibition was determined. AmTI-mRNA accumulation reached a maximum at 14 days after anthesis (daa) and then gradually decreased, being barely detectable 36 daa. The AmTI protein accumulation followed the same profile as the inhibitory activity, both were delayed with respect to the mRNA. The maximum level was observed 22 daa, and then gradually decreased until a steady state was reached as seed maturation proceeded. Upon imbibition, a gradual decrease in AmTI protein and inhibitory activity was shown; however, an AmTI transcript was detected 24 h after imbibition. In contrast to representative members of the potato I family, this inhibitor was not inducible by wounding of leaves.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006262106267
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  • 88
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    Localization of peroxidase mRNAs in soybean seeds by in situ hybridization (1999)
    Springer
    Plant molecular biology 41 (1999), S. 57-63 
    Details
    ISSN: 1573-5028
    Keywords: embryo ; gene expression ; Glycine max ; oxidoreductase ; seed coat ; testa
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The soybean Ep gene encodes an anionic peroxidase enzyme that accumulates in large amounts in seed coat tissues. We have isolated a second peroxidase gene, Prx2, that is also highly expressed in developing seed coat tissues. Sequence analysis of Prx2 cDNA indicates that this transcript encodes a cationic peroxidase isozyme that is far removed from Ep in peroxidase phylogeny. To determine the expression patterns for these two peroxidases in developing seeds, the abundance and localization of the Ep and Prx2 transcripts were compared by in situ hybridization. Results show the expression of Ep begins in a small number of cells flanking the vascular bundle in the seed coat, spreads to encircle the seed, and then migrates to the hourglass cells as they develop. Expression of Prx2 occurs throughout development in all cell layers of the seed coat, and is also evident in the pericarp and embryo. Nonetheless, the Ep-encoded enzyme accounts for virtually all of the peroxidase activity detected in mature seed coats. The Prx2 enzyme is either insoluble in a catalytically inactive form, or is subject to degradation during seed maturation.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006244500951
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  • 89
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    Constitutive protein-DNA interactions on the abscisic acid-responsive element before and after developmental activation of the rab28 gene (1999)
    Springer
    Plant molecular biology 41 (1999), S. 529-536 
    Details
    ISSN: 1573-5028
    Keywords: ABRE ; embryogenesis ; G-box ; gene expression ; maize ; protein-DNA interaction
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Transcription of the rab28 gene from maize is induced in late embryo development and in response to abscisic acid. We have studied the regulation of the activity of the rab28 promoter in embryos. Two abscisic acid-responsive elements (ABREs) were necessary for expression in embryos of transgenic Arabidopsis and in transient transformation in maize embryos. In vivo footprinting showed that there was protein binding to the ABREs and to other cis elements in the promoter in young embryos before expression of rab28. This shows that the rab28 promoter is in an open chromatin structure before developmental activation. The ABREs are important for the induction and have protein binding in young embryos. Nuclear proteins extracted from embryos before activation of rab28 bound to the ABREs in band shift assays. A complex with different mobility was formed between nuclear proteins and the ABREs after induction of rab28 suggesting a modification of the ABRE-binding factor or an exchange of proteins. The footprints on the ABREs were unaltered by induction with abscisic acid or during developmental activation of rab28. These results indicate that constitutive binding of transcription factor(s) on the ABRE is central in embryonic regulation of the rab28 gene.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006345113637
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  • 90
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    Molecular characterization of a gene for alanine aminotransferase from rice (Oryza sativa) (1999)
    Springer
    Plant molecular biology 39 (1999), S. 149-159 
    Details
    ISSN: 1573-5028
    Keywords: alanine aminotransferase ; gene expression ; GUS expression ; promoter
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract A cDNA clone encoding alanine aminotransferase (AlaAT) has isolated from randomly sequenced clones derived from a cDNA library of maturing rice seeds by comparison to previously identified genes. The deduced amino acid sequence was 88% and 91% homologous to those of the enzymes from barley and broomcorn millet (Panicum miliaceum), respectively. Using this cDNA as a probe, we isolated and sequenced the corresponding genomic clone. Comparison of the sequences of the cDNA and the genomic gene revealed that the coding region of the gene was interrupted by 14 introns 66 to 1547 bp long. Northern and western blotting analyses showed that the gene was expressed at high levels in developing seeds. When the 5′-flanking region between −930 and +85 from the site of initiation of transcription was fused to a reporter gene for β-glucuronidase (GUS) and then introduced into the rice genome, histochemical staining revealed strong GUS activity in the inner endosperm tissue of developing seeds and weak activity in root tips. Similar tissue-specific expression was also detected by in situ hybridization. These results suggest that AlaAT is involved in nitrogen metabolism during the maturation of rice seed.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006156214716
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  • 91
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    Differential expression of expansin gene family members during growth and ripening of tomato fruit (1999)
    Springer
    Plant molecular biology 39 (1999), S. 161-169 
    Details
    ISSN: 1573-5028
    Keywords: expansin ; fruit growth ; fruit softening ; gene expression ; Lycopersicon esculentum
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract cDNA clones encoding homologues of expansins, a class of cell wall proteins involved in cell wall modification, were isolated from various stages of growing and ripening fruit of tomato (Lycopersicon esculentum). cDNAs derived from five unique expansin genes were obtained, termed tomato Exp3 to Exp7, in addition to the previously described ripening-specific tomato Exp1 (Rose et al. (1997) Proc Natl Acad Sci USA 94: 5955–5960). Deduced amino acid sequences of tomato Exp1, Exp4 and Exp6 were highly related, whereas Exp3, Exp5 and Exp7 were more divergent. Each of the five expansin genes showed a different and characteristic pattern of mRNA expression. mRNA of Exp3 was present throughout fruit growth and ripening, with highest accumulation in green expanding and maturing fruit, and lower, declining levels during ripening. Exp4 mRNA was present only in green expanding fruit, whereas Exp5 mRNA was present in expanding fruit but had highest levels in full-size maturing green fruit and declined during the early stages of ripening. mRNAs from each of these genes were also detected in leaves, stems and flowers but not in roots. Exp6 and Exp7 mRNAs were present at much lower levels than mRNAs of the other expansin genes, and were detected only in expanding or mature green fruit. The results indicate the presence of a large and complex expansin gene family in tomato, and suggest that while the expression of several expansin genes may contribute to green fruit development, only Exp1 mRNA is present at high levels during fruit ripening.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006130018931
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    Isolation and characterization of mRNAs differentially expressed during ripening of wild strawberry (Fragaria vesca L.) fruits (1999)
    Springer
    Plant molecular biology 39 (1999), S. 629-636 
    Details
    ISSN: 1573-5028
    Keywords: cDNA cloning ; fruit ripening ; gene expression ; non-climacteric fruit ; wild strawberry (Fragaria vesca L.)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Wild strawberry (Fragaria vesca L.) is an attactive model system for studying ripening in non-climacteric fruit, because of its small diploid genome, its short reproductive cycle, and its capacity for transformation. We have isolated eight ripening-induced cDNAs from this species after differential screening of a cDNA library. The predicted polypeptides of seven of the clones exhibit similarity to database protein sequences, including acyl carrier protein, caffeoyl- CoA 3-O-methyltransferase, sesquiterpene cyclase, major latex protein, cystathionine γ-synthase, dehydrin and an auxin- induced gene. A ninth cDNA clone that was constitutively expressed is predicted to encode a metallothionein-like protein. None of these proteins appear to be directly related to events generally associated with ripening such as cell wall metabolism or the accumulation of sugars and pigments, rather, their putative functions are indicative of the wide range of processes upregulated during fruit ripening.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006179928312
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    Enzymatic activity and gene expression under water stress of phospholipase D in two cultivars of Vigna unguiculata L.Walp. differing in drought tolerance (1999)
    Springer
    Plant molecular biology 39 (1999), S. 1257-1265 
    Details
    ISSN: 1573-5028
    Keywords: cowpea (Vigna unguiculata L.) ; drought ; gene expression ; lipid degradation ; phospholipase D
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Phospholipase D, a major lipid-degrading enzyme in plants, was studied in two cultivars of Vigna unguiculata L.Walp, differing in their tolerance to drought (cv. EPACE-1, drought-tolerant, and cv. 1183, drought-susceptible). Enzymatic activities, measured with 14C-PC as substrate, increased when plants were submitted to water stress, the increase being much higher in the drought-sensitive cultivar. A 2911 bp cDNA encoding a putative phospholipase D (VuPLD1) was isolated from a cDNA library prepared from V. unguiculata leaves. The deduced amino acid sequence (809 residues) shows 85.5% identity and 91.3% similarity to that of PLD from Ricinus communis. The expression of the VuPLD1 gene in the leaves is differently modulated by water deficit, depending on the intensity of stress and the tolerance or sensitivity of the plants. In the drought-susceptible V. unguiculata cv. 1183, it readily increased under water stress, reaching maximum values at mild water deficit (−1.5 MPa). In the drought-tolerant cv. EPACE-1, VuPLD1 mRNA remained low throughout the whole drought treatment. Dehydration of leaves led to a dramatic increase in transcript level in both cultivars. Changes in protein amounts semi-quantified by immunoblotting correlated well with variations in transcript steady-state level. Taken together, these results showed that phospholipase D in cowpea plants is essentially regulated at the transcriptional level, and that gene expression is strongly stimulated even by moderate water deficit in the drought-sensitive plant. On the contrary, the drought-tolerant plant presents a remarkable stability of PLD gene expression in conditions of water stress.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006165919928
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  • 94
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    Identification of genes specifically expressed in cauliflower reproductive meristems. Molecular characterization of BoREM1 (1999)
    Springer
    Plant molecular biology 39 (1999), S. 427-436 
    Details
    ISSN: 1573-5028
    Keywords: reproductive development ; gene expression ; subtractive hybridization ; cauliflower
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Using the meristems of the cauliflower curd as a source of tissue and a series of subtractive hybridizations and amplification reactions, we have constructed a cDNA library highly enriched in cDNAs expressed in reproductive meristems. The analysis of a sample of 250 clones from this library identified 22 cDNA clones corresponding to genes specifically expressed in these cauliflower meristems. Apart from two clones that corresponded to APETALA1, and two other ones showing similarity to different aminoacyl-tRNA synthetases, the remaining clones showed no similarity to any sequence in the databases and may correspond to novel genes. One of these clones, BoREM1, was further characterized and found to correspond to a gene encoding a protein with features of regulatory proteins that follows a expression pattern very similar to the LEAFY transcripts.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006130629100
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  • 95
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    A strong constitutive positive element is essential for the ammonium-regulated expression of a soybean gene encoding cytosolic glutamine synthetase (1999)
    Springer
    Plant molecular biology 39 (1999), S. 551-564 
    Details
    ISSN: 1573-5028
    Keywords: ammonium ; gene expression ; glutamine synthetase ; nodules ; positive element ; promoter
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract In order to identify important promoter elements controlling the ammonium-regulated expression of the soybean gene GS15 encoding cytosolic glutamine synthetase, a series of 5′ promoter deletions were fused to the GUS reporter gene. To allow the detection of positive and negative regulatory elements, a series of 3′ deletions were fused to a −90 CaMV 35S promoter fragment placed upstream of the GUS gene. Both types of construct were introduced into Lotus corniculatus plants and soybean roots via Agrobacterium rhizogenes-mediated transformation. Both spectrophotometric enzymatic analysis and histochemical localization of GUS activity in roots, root nodules and shoots of transgenic plants revealed that a strong constitutive positive element (SCPE) of 400 bp, located in the promoter distal region is indispensable for the ammonium- regulated expression of GS15. Interestingly, this SCPE was able to direct constitutive expression in both a legume and non- legume background to a level similar to that driven by the CaMV 35S full-length promoter. In addition, results showed that separate proximal elements, located in the first 727 bp relative to the transcription start site, are essential for root- and root nodule-specific expression. This proximal region contains an AAAGAT and two TATTTAT consensus sequences characteristic of nodulin or nodule-enhanced gene promoters. A putative silencer region containing the same TATTTAT consensus sequence was identified between the SCPE and the organ-specific elements. The presence of positive, negative and organ-specific elements together with the three TATTTAT consensus sequences within the promoter strongly suggest that these multiple promoter fragments act in a cooperative manner, depending on the spatial conformation of the DNA for trans-acting factor accessibility.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006169018296
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  • 96
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    Cloning and characterization of six embryogenesis-associated cDNAs from somatic embryos of Picea glauca and their comparative expression during zygotic embryogenesis (1999)
    Springer
    Plant molecular biology 39 (1999), S. 859-864 
    Details
    ISSN: 1573-5028
    Keywords: embryo-abundant cDNAs ; gene expression ; gymnosperm ; Picea glauca ; somatic embryogenesis
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Six somatic embryogenesis-associated cDNAs (PgEMB2, 6, 7, 8, 24 and 34) from white spruce (Picea glauca (Moench) Voss) somatic embryos have been characterized. Transcript accumulation during somatic embryo development and subsequent germination related to these genes, indicated that they were developmentally regulated. The transcripts related to clones PgEMB2, 6, 24 and 34 were also detected during zygotic embryo development, but transcripts of clones PgEMB7 and 8 were not. PgEMB24 had a similar gene expression pattern to spruce Em-like late embryo abundant (lea) gene, but other clones had no similarities in gene expression to either spruce lea-like or storage protein genes. Abscisic acid, a stimulator for spruce somatic embryo maturation, did not obviously affect gene expression corresponding to these cDNAs. The predicted proteins are distinguishable from known LEA proteins based on analyses of hydropathy plots, amino acid compositions and deduced protein structures. The similarities of the spruce cDNAs, and protein sequences predicted from these cDNAs, to other sequence data are described.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006146622614
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  • 97
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    Mutation of GT-1 binding sites in the Pr-1A promoter influences the level of inducible gene expression in vivo (1999)
    Springer
    Plant molecular biology 40 (1999), S. 387-396 
    Details
    ISSN: 1573-5028
    Keywords: gene expression ; GT-1 ; PR-1a ; PR proteins ; salicylic acid-induced ; transcription factors
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Infection of Nicotiana tabacum Samsun NN with tobacco mosaic virus (TMV) results in a hypersensitive plant response and leads to systemic acquired resistance (SAR). The induction of SAR is mediated by the plant hormone salicylic acid (SA) and is accompanied by the induced expression of a number of genes including the pathogenesis-related (PR) gene 1a. Previously, it has been found that TMV infection and SA treatment resulted in a reduction of binding of nuclear protein GT-1 to far-upstream regions (−902 to −656) of the PR-1a gene. To test if GT-1 is a negative regulator of PR-1a gene expression, the effects of mutations in the seven putative GT-1 binding sites in this region were studied in vitro using dimethyl sulfate interference footprinting and band shift assays. This showed that at least one of the seven sites is indeed a GT-1 binding site. However, when tested in transgenic plants, the mutations did not result in constitutive expression of the chimeric PR-1a/GUS transgene, while inducible expression after SA treatment was decreased. The results suggest that binding of GT-1-like proteins to far-upstream PR-1a promoter regions indeed influences gene expression. A possible model for GT-1's mode of action in PR-1a gene expression is discussed.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006144505121
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  • 98
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    Expression of two PIP genes in rapidly growing internodes of rice is not primarily controlled by meristem activity or cell expansion (1999)
    Springer
    Plant molecular biology 40 (1999), S. 985-995 
    Details
    ISSN: 1573-5028
    Keywords: aquaporin ; gene expression ; growth ; Oryza sativa ; plasma membrane intrinsic protein (PIP) ; rice
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Membrane intrinsic proteins facilitate movement of small molecules often times functioning as water channels. We have identified two genes from rice which encode proteins with characteristic features of plasma membrane intrinsic proteins (PIP). They possess six membrane-spanning domains, an NPA repeat, overall high sequence homologies and characteristic C- and N-terminal hallmark motifs which allowed assignment of OsPIP1a to the PIP1 subfamily and of OsPIP2a to the PIP2 subfamily. OsPIP1a and OsPIP2a showed similar but not identical expression patterns. The two genes were expressed at higher levels in seedlings than in adult plants and expression in the primary root was regulated by light. In internodes of deepwater rice plants which were induced to grow rapidly by submergence, transcript levels were slightly induced in the intercalary meristem (IM) and slightly reduced in the elongation zone (EZ) after 18 h. In internodes of GA-induced excised stem sections transcript levels transiently declined in the IM and EZ after 1 h and subsequently recovered to elevated levels after 18 h. GA also induced OsPIP expression in non-growing tissue after 18 h. In the IM of submergence-induced stem sections transcript levels remained constitutive. The different growth-promoting treatments showed no direct correlation between growth rate and OsPIP gene expression in dividing or expanding cells. In fact, treatment of excised stem sections with ABA or drought stress induced similar changes in OsPIP expression in the growing zone during the first 6 h as GA did. We conclude that regulation of OsPIP1a and OsPIP2a expression is not primarily controlled by growth. GA-induced growth may however change the water status of cells which in turn results in altered PIP abundance.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006265528015
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  • 99
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    Identification of two chilling-regulated 1-aminocyclopropane- 1-carboxylate synthase genes from citrus (Citrus sinensis Osbeck) fruit (1999)
    Springer
    Plant molecular biology 41 (1999), S. 587-600 
    Details
    ISSN: 1573-5028
    Keywords: ACC synthase ; chilling ; Citrus sinensis ; ethylene ; gene expression ; peel
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Diurnal change in the temperature below or above 12.5 °C hastens the degreening of citrus peel and elicits the phytohormone ethylene production in citrus fruit. Ethylene triggers the degradation of chlorophyll and synthesis of carotenoids in citrus peel. To investigate if ethylene is required for the degreening of citrus peel elicited by low temperatures, we studied the chilling-regulated gene expression of ACC synthase, one of the key enzymes catalyzing ethylene biosynthesis. We isolated and characterized a chilling-inducible 1-aminocyclopropane-1-carboxylate synthase (ACC synthase) gene, CS-ACS1, and a chilling-repressible gene, CS-ACS2, from citrus peel. The CS-ACS1 transcript 1.7 kb in length encodes a polypeptide of 483 amino acids (M r 54 115, pI 6.63), whereas the CS-ACS2 transcript of 1.8 kb encodes a polypeptide of 477 amino acids (M r 53 291, pI 6.72). Both genes showed a rapid but transient induction (within 2.4 h) of transcripts upon rewarming after the chilling (4 °C) treatment. After 24 h of incubation at room temperature, CS-ACS1 mRNA diminished to an undetectable level, whereas the CS-ACS2 mRNA regained its basal level of expression attained prior to the chilling treatment. Chilling-induced ethylene production and ACC accumulation were also observed upon rewarming. Both genes were also induced by the wound stress (excision). The protein synthesis inhibitor cycloheximide super-enhances the accumulation of both ACS transcripts at room temperature. Molecular analysis of the 3.3 kb genomic DNA of CS-ACS1 revealed that this gene consists of three introns and four exons. The intron 3 is exceptionally large (1.2 kb) and shares significant homology with mitochondrial DNA, supporting the intron-late theory.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006369016480
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    Electronic Resource
    Electronic Resource
    Occurrence of five different chromosomal HMG1 proteins in various maize tissues (1999)
    Springer
    Plant molecular biology 41 (1999), S. 351-361 
    Details
    ISSN: 1573-5028
    Keywords: chromatin ; gene expression ; high-mobility-group protein HMG1 ; HMGe ; protein stability ; Zea mays
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The nuclear HMG1 proteins of higher plants are small non-histone proteins that have DNA-bending activity and are considered architectural factors in chromatin. The occurrence of the chromosomal HMG1 proteins, HMGa, HMGc1/2 and HMGd, in various maize tissues was analyzed, and in the course of these studies a novel HMG1 protein, now termed HMGe, was identified. Purification and characterization of HMGe (Mr 13 655) and cloning of the corresponding cDNA revealed that it displays only moderate similarity to other members of the plant HMG1 protein family. The five maize HMG1 proteins could be detected in kernels, leaves, roots and suspension culture cells, indicating that these proteins can be expressed simultaneously and occur relatively ubiquitously. However, the various HMG1 proteins are present in significantly different quantities with HMGa and HMGc1/2 being the most abundant HMG1 proteins in all tissues tested. Furthermore, the relative amounts of the various HMG1 proteins differ among the tissues examined. The HMG1 proteins were found to be relatively stable proteins in vivo, with HMGc1/2, HMGd and HMGe having a half-life of ca. 50 h in cultured cells, while the half-life of the HMGa protein is ca. 65 h. Collectively, these findings are compatible with the concept that the different plant HMG1 proteins might act as general architectural proteins in concert with site-specific factors in the assembly of certain nucleoprotein structures involved in various biological processes.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006311121717
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