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  • 1
    Electronic Resource
    Electronic Resource
    Springer
    Journal of insect behavior 3 (1990), S. 579-587 
    ISSN: 1572-8889
    Keywords: Genetics ; polymorphism ; reproductive isolation ; hovering behavior ; Tabanus nigrovittatus
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The salt marsh horse fly, Tabanus nigrovittatusMacquart, exhibits two nonoverlapping daily periods of hovering and mating activity, which are correlated with different environmental temperatures. Allelic and genotypic frequencies of hovering males collected during the two periods were compared by electrophoresis of three polymorphic enzyme loci. Approximately 26% of early-hovering males possessed a Pgmallozyme that was absent in our sample of late-hovering males. However, based on other allozyme loci, we found no evidence for reproductive isolation between early and late hoverers. All the genetic data are consistent with the hypothesis that the Pgmpolymorphism is associated with behaviorally and physiologically distinct groups of males that, by all other criteria, form a single Mendelian population.
    Type of Medium: Electronic Resource
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  • 2
    Electronic Resource
    Electronic Resource
    Springer
    Pediatric surgery international 5 (1990), S. 359-360 
    ISSN: 1437-9813
    Keywords: Hirschsprung's disease ; Genetics
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract Three male children with identical short-segment Hirschsprung's disease born to a young married couple are reported. There was no positive family history despite an extensive search. There were no associated abnormalities. Although sex-modified multifactorial inheritance, with males having a lower threshold of genes for expression of Hirschsprung's disease, is accepted, the identical expression of the disorder in the three siblings presented suggests a dominant, possibly X-linked gene with variable penetrance. Another possibility is that an identical micro-environmental factor was present prenatally resulting in all three boys having Hirschsprung's disease. This is the first report of three siblings with identical short-segment Hirschsprung's disease.
    Type of Medium: Electronic Resource
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  • 3
    Electronic Resource
    Electronic Resource
    Springer
    Pediatric nephrology 4 (1990), S. 533-541 
    ISSN: 1432-198X
    Keywords: Henoch-Schönlein purpura ; Pathogenesis ; Genetics
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract Laboratory studies of the pathophysiology of Henoch-Schönlein purpura (HSP) have become more numerous in recent years with the recognition of the disease's links with the mucosal immune system in general and IgA nephropathy in particular. There are weak genetic associations with C4 null phenotypes and with HLA B35 and DR4. Studies of plasma proteins in HSP patients show an increased IgA concentration, activation of the alternative pathway of complement and consumption of factor XIII. High molecular weight (polymeric) IgA has been detected in affected individuals, which some investigators have called “immune complexes”. Many patients synthesise an IgA rheumatoid factor in the acute phase, but other autoantibodies are largely absent. In vitro studies of lymphocytes from HSP patients have demonstrated an increased number of IgA-bearing and secreting B-cells, with altered T-cell regulation of antibody synthesis. While these observations point to immune dysregulation — primarily of IgA production — as a consistent feature of acute HSP, there is as yet insufficient information available to allow a consistent theory of pathogenesis to be formulated.
    Type of Medium: Electronic Resource
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  • 4
    Electronic Resource
    Electronic Resource
    Springer
    Sexual plant reproduction 3 (1990), S. 31-34 
    ISSN: 1432-2145
    Keywords: Mucoraceae ; Zygomycetes ; Homothallic ; Genetics ; Nutritional complementation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Auxotrophic strains of Mucor genevensis and Zygorhynchus exponens were crossed and the resulting zygospores germinated. The presence of a true sexual cycle in both species was demonstrated by the recovery of recombinant genotypes. Expected Mendelian ratios were not realized, however. The presence of selfed zygospores among those isolated makes this observation understandable. It was possible to demonstrate nutritional complementation when young mating mycelium was transferred to minimal medium and forced heterokaryons were recovered.
    Type of Medium: Electronic Resource
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  • 5
    ISSN: 1432-1076
    Keywords: Osteogenesis imperfecta ; Collagen type I ; Radiology, classification ; Genetics
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract Congenital osteogenesis imperfecta (OI) was diagnosed by ultrasound in a 31-week-old fetus, and the diagnosis confirmed after delivery by caesarean section at week 36. The baby survived the neonatal period, but failed to thrive, had recurrent respiratory infections and ultimately died at 8 months. Cultured fibroblasts synthesized both normal type I collagen and unstable type I collagen harbouring a structural defect in the α1(I) cyanogen bromide-derived peptide number 8 (CB8) region of the molecule, indicating a heterozygous dominant mutation. A+ birth, the radiological picture was that of the “thin bone”-type of congenital OI (OI type IIB/III in the Sillence classification); at the age of 12 weeks ribs and long bones had undergone a marked expansion giving a very different picture, that of the “thick bone”-type congenital OI (OI type IIA). The mechanism responsible for this change in bone structure is not known, but fractures and callus formation are unlikely to be the only factors. Caution is needed in the interpretation of radiographs of newborns with OI for prognostic or genetic purposes.
    Type of Medium: Electronic Resource
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  • 6
    Electronic Resource
    Electronic Resource
    Springer
    Journal of comparative physiology 166 (1990), S. 545-552 
    ISSN: 1432-1351
    Keywords: Honeybees ; Learning ; Classical conditioning ; Selection ; Genetics
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary Four strains of the honeybee (Apis mellifera capensis), which were selected for high (N=2) or low (N=2) performance levels in classic conditioning of olfactory and mechanosensory stimuli, were examined in two instrumental visual learning tasks. Bees were trained to coloured cardboards either at the hive entrance or at the feeding station. Positive correlations were detected between olfactory/mechanosensory conditioning and visual learning. Good and poor learners from strains selected for olfactory conditioning differed significantly in their visual learning values. These strain differences reflect genetic differences in a common learning system rather than task specific differences in sensory, motor or motivational components. Parameters that were influenced by activity of the colony (duration of stay at the feeding place, time between visits) also differed among selected strains. These effects were not due to selection. Instead, they reflect a specific genetic background produced in each strain independently of selection. The results indicate that associative learning has a genetic basis which is independent of the sensory stimuli associated with reward, the learning procedure (classical conditioning or instrumental learning) or the motor patterns used to execute the learned behavior (proboscis extension, control for flight behavior, open field orientation).
    Type of Medium: Electronic Resource
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  • 7
    Electronic Resource
    Electronic Resource
    Springer
    European archives of psychiatry and clinical neuroscience 239 (1990), S. 290-292 
    ISSN: 1433-8491
    Keywords: Schizophrenia ; Linkage ; Genetics
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Summary We analysed six multiplex pedigrees of schizophrenia for linkage to two DNA probes mapping to the chromosome 5q11–q13 region where linkage to a gene for schizophrenia was recently reported. Analyses were conducted using three penetrance models and considering the affected state to be schizophrenia, the schizophrenia spectrum, and all psychiatric diagnoses. All analyses gave consistently negative lod scores. Although the region was not formally excluded, no evidence for linkage was found.
    Type of Medium: Electronic Resource
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  • 8
    ISSN: 1432-1459
    Keywords: Spinocerebellar degeneration ; Friedreich's disease ; Diagnostic criteria ; Genetics ; Natural history
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Summary The clinical and genetic features of 80 patients with Friedreich's disease from 64 families are described. Diagnostic criteria were: no evidence of dominant inheritance, onset by the age of 20 years, progressive unremitting ataxia of limbs and gait, and absence of knee and ankle jerks. Furthermore, at least one of the following accessory signs was present: dysarthria, extensor plantar response and echocardiographic evidence of hypertrophic cardiomyopathy. Two peaks of onset age were evident at 6–9 and 12–15 years. Analysis of intrafamily variation of onset age and absence of clustering of cardiomyopathy and diabetes did not suggest genetic heterogeneity. Peripheral nerve impairment was an early finding and showed slight further progression, whereas involvement of the cerebellar and corticospinal pathways appeared later and mainly accounted for the progressive worsening of the disease.
    Type of Medium: Electronic Resource
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  • 9
    ISSN: 1432-1343
    Keywords: Analysis of variance ; Choropleth map ; Ecology ; Genetics ; Geography ; Permutation test ; Spatial autocorrelation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Mathematics
    Description / Table of Contents: Résumé Cet article présente une solution au problème de l'analyse de variance, pour certains cas où la variable à analyser est spatialement autocorr élée alors que le critère de classification représente des sous-régions connexes du territoire à l'étude. On sait que les méthodes classiques d'analyse de variance ne sont pas applicables dans ce type de situation puisque la condition d'indépendance des échantillons n'est pas respectée; l'autocorrélation positive réduit la variabilité intragroupe, si bien que la quantité relative de variabilité intergroupe s'en trouve artificiellement augmentée. Cette situation correspond en réalité à une vaste catégorie de problèmes en génétique des populations, en écologie et dans d'autres branches de la biologie, ainsi qu'en épidémiologie, en géographie, en géologie, en science économique, en science politique et en sociologie. Ce nouveau test appartient à la famille des tests par permutation. Nous calculons la somme des dispersions intragroupes et testons contre une distribution de référence obtenue en permutant les régions géographiques un grand nombre de fois sur la carte. La véritable difficulté de ce test est d'ordre algorithmique, puisqu'il n'est pas facile de permuter des régions sur une carte, de façon à ce que chaque groupe demeure connexe, et que la carte permutée occupe le même espace total que la carte d'origine. Cet article présente la théorie, les algorithmes, ainsi que des résultats obtenus par cette méthode. Un programme écrit en PASCAL est disponible.
    Notes: Abstract The classical method for analysis of variance of data divided in geographic regions is impaired if the data are spatially autocorrelated within regions, because the condition of independence of the observations is not met. Positive autocorrelation reduces within-group variability, thus artificially increasing the relative amount of among-group variance. Negative autocorrelation may produce the opposite effect. This difficulty can be viewed as a loss of an unknown number of degrees of freedom. Such problems can be found in population genetics, in ecology and in other branches of biology, as well as in economics, epidemiology, geography, geology, marketing, political science, and sociology. A computer-intensive method has been developed to overcome this problem in certain cases. It is based on the computation of pooled within-group sums of squares for sampled permutations of internally connected areas on a map. The paper presents the theory, the algorithms, and results obtained using this method. A computer program, written in PASCAL, is available.
    Type of Medium: Electronic Resource
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  • 10
    ISSN: 1432-2242
    Keywords: Genetics ; Symbiosis ; Nitrogen fixation ; Coevolution
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary To determine the relationship between nodulation restriction by the Rj4 allele of soybean, rhizobitoxine-induced chlorosis, and taxonomic grouping of bradyrhizobia, 119 bradyrhizobial isolates were tested in Leonard jar culture for nodulation response and chlorosis induction. In addition to strain USDA 61, the strain originally reported as defining the Rj4 response, eight other isolates (i.e., USDA 62, 83, 94, 238, 252, 259, 260, and 340) were discovered to elicit the nodulation interdiction of the Rj4 allele. Only 16% of all the bradyrhizobial strains tested induced chlorosis, but seven of the nine strains (78%) interdicted by the Rj4 allele were chlorosis-inducing strains. Furthermore, in tests for antibiotic resistance profile, eight of the nine interdicted strains (89%) were classed in DNA homology group II. This evidence suggests that the Rj4 allele has a positive value to the host plant in shielding it from nodulation by certain chlorosis-inducing bradyrhizobia of a DNA homology group with impaired efficiency of nitrogen fixation with soybean.
    Type of Medium: Electronic Resource
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  • 11
    ISSN: 1432-2242
    Keywords: Beta vulgaris ; Sugar beet ; Isozymes ; Genetics ; Linkage ; Pollen fertility restorer
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The NADP-specific malate dehydrogenase isozymes were controlled by multiple gene systems. Three genes coding for dimeric enzymes segregated in a dependent fashion (NADP-Mdh 1, NADP-Mdh 2, NADP-Mdh 3). A fourth gene (NADP-Mdh 4), also coded for dimers, but was not polymorphic in B. vulgaris. A fifth gene (NADP-Me 1) coded for enzymes active as monomers. Two genes were found to control the main zone of NAD-specific malate dehydrogenase: one coded for dimers (Mdh 1), while a second (Mdh 2) was not polymorphic in the assessions studied. 6-P-Gluconate dehydrogenase was not polymorphic in B. vulgaris; the two types detected on SGE1 electrophoresis were due to developmental expression of the different systems. No genetical segregations could be detected in progeny of crosses of the distinct phenotypes. A shikimate dehydrogenase gene (Skdh 1) that coded for monomers was identified. The diaphorase system was rather complex, but one gene (Dia 1) coding for monomeric enzymes could be identified. Aconitase was found to be controlled by two independent genes (Aco 1, Aco 2), both polymorphic and coding for proteins active as monomers. Tight linkage was found between the genes NADP-Mdh 1, NADP-Mdh 2 and NADP-Mdh 3. Linkage was also found between a pollen fertility restorer (Z) and the Mdh 1 gene. The identification of linkage with Aco 1 needs further investigation. R segregated independently from Mdh 1, Aco 1 and Dia 1. Independent segregations were scored for isozyme genes Pgm 2, Icd 1, Ak 1, Gpi 1, Aco 1 and Dia 1.
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  • 12
    Electronic Resource
    Electronic Resource
    Springer
    Biology and philosophy 5 (1990), S. 349-371 
    ISSN: 1572-8404
    Keywords: Genetics ; gene structure ; hereditary unit
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Philosophy
    Notes: Abstract Definitions of the term ‘gene’ typically superimpose molecular genetics onto Mendelism. What emerges are persistent attempts to regard the gene as a ‘unit’ of structure and/or function, language that creates multiple meanings for the term and fails to acknowledge the diversity of gene architecture. I argue that coherence at the molecular level requires abandonment of the classical unit concept and recognition that a gene is constructed from an assemblage of domains. Hence, a domain set (1) conforms more closely to empirical evidence for genetic organization of DNA regions capable of transcription and (2) has ontological properties lacking in the traditional unit definition.
    Type of Medium: Electronic Resource
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  • 13
    ISSN: 1573-5133
    Keywords: Demography ; Genetics ; Geographic variation ; Stochasticity ; Fluctuating environments ; Allele frequencies ; River ecology
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Synopsis The purpose of this study was to determine the effects of unpredictable environmental fluctuations on the demographic and genetic structure of Fundulus zebrinus populations. Collections of F. zebrinus were taken from three rivers in the Arkansas River basin: the Arkansas, Chikaskia, and Ninnescah. Fish were sampled from three sites on each river on nine collection dates throughout 1984 and 1985. Totals of 2100 fish and 6000 fish were included in electrophoretic and demographic analyses, respectively. The results of the study indicate that within a limited geographic region (i.e. within rivers) spatial differences and temporal changes in both demographic and genetic population characteristics occur frequently and are primarily stochastic. However, on a larger spatial scale (i.e. across rivers), general trends emerge for demographic and especially for genetic population characteristics. These results illustrate the importance of sampling scale for conclusions of life-history evolution in fluctuating environments. In addition, it was found that regulation of Fundulus zebrinus populations includes an important density-independent component. Stochastic demographic differences across space and changes through time and spatially and temporally heterogeneous allele frequencies, are both indicative of density-independent regulation. Variation in population parameters, both demographic and genetic, was observed between populations sampled from each river. These population differences were attributed to differences between the rivers themselves.
    Type of Medium: Electronic Resource
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  • 14
    Electronic Resource
    Electronic Resource
    Springer
    Journal of chemical ecology 16 (1990), S. 2935-2946 
    ISSN: 1573-1561
    Keywords: Genetics ; sex pheromone ; Lepidoptera ; Noctuidae ; Trichoplusia ni ; cabbage looper moth ; reproductive isolation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Abstract The genetic basis of interpopulational differences in the pheromone blend emitted by the cabbage looper moth,Trichoplusia ni (Hübner), was examined by crossing individuals from a field-derived population (P1) with individuals from a long-maintained laboratory colony (P2). These colonies differed in the emission rate and relative proportions of four of the five known minor pheromone components, but not in the emission rate of the major component, (Z)-7-dodecenyl acetate (Z7-12∶Ac). These differences in pheromone blend were quantitatively small but biologically significant, because in the field, males responded preferentially to traps baited with a pheromone blend that is similar to that emitted by P1 females relative to a blend similar to that emitted by P2 females. In initial crosses, variation in the quantity and quality of pheromone blends among families of P1, P2, and F1 hybrid females was examined. In F1 females the relative proportions (quantity relative to the major component) of (Z)-5-dodecenyl acetate (Z5-12∶Ac) and (Z)-7-tetradecenyl acetate (Z7-14∶Ac) were intermediate to parental lines. In a second more extensive set of crosses, analyses included P1, P2, F1, F2, and selected backcrosses. The relative proportion of Z5-12∶Ac, Z7-14∶Ac, and Z9-14∶Ac emitted by F1 females were intermediate to parental lines. The frequency distributions of relative proportions of these components emitted by females were not consistent with those expected under a single autosomal or sex-linked gene hypothesis, suggesting that more than one gene is involved in the quantitative differences in the pheromone blend.
    Type of Medium: Electronic Resource
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  • 15
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Molecular Reproduction and Development 27 (1990), S. 168-172 
    ISSN: 1040-452X
    Keywords: Motility ; Genetics ; Sex chromosome ratio ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: In this study, we address the relationship between motility and genetic content of mouse sperm. The chromosome complements of highly motile mouse sperm, selected using the swim-up technique, were analyzed after in vitro fertilization, at the first cleavage state. They were compared to those of unselected sperm. Identification of male and female chromosome sets was possible because of their differential condensation at the first mitotic division. In vitro fertilization, swim-up separation, chromosome preparation, and staining were carried out using standard techniques. The results indicate that highly motile mouse sperm did not differ in types and frequencies of chromosomal abnormalities from those not selected for motility. Moreover, separation of motile sperm does not deviate the sex ratio from the theoretical 1:1.
    Additional Material: 1 Ill.
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  • 16
    Electronic Resource
    Electronic Resource
    Chichester [u.a.] : Wiley-Blackwell
    Developmental Genetics 11 (1990), S. 88-96 
    ISSN: 0192-253X
    Keywords: Pattern formation ; segmentation ; gap genes ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: L(1)giant is a zygotic lethal mutation which affects the embryonic development of both the labial/thoracic segments and a subset of posterior abdominal segments. Using antibodies specific for proteins encoded by several Drosophila genes to identify the compartmental origin of the defects, we show that the requirement of giant activity is different in these two embryonic domains. Anteriorly, the posterior compartment of the labial segment is missing at the blastoderm stage. Posteriorly, cells are specifically deleted by cell death within the anterior compartments of abdominal segments 5-7 during germ band elongation. In mature embryos, posterior compartment structures of the peripheral nervous system of A5-7 are fused. In addition to a different pattern of defect in the two parts of the embryo, the kind of action appears different. Anteriorly, giant resembles a gap mutation in that a particular region is missing from the blastoderm fate map, whereas in the abdominal domain, giant affects the development of anterior compartment-specific structures.
    Additional Material: 4 Ill.
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  • 17
    Electronic Resource
    Electronic Resource
    Chichester [u.a.] : Wiley-Blackwell
    Developmental Genetics 11 (1990), S. 123-123 
    ISSN: 0192-253X
    Keywords: Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Type of Medium: Electronic Resource
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  • 18
    Electronic Resource
    Electronic Resource
    Chichester [u.a.] : Wiley-Blackwell
    Developmental Genetics 11 (1990) 
    ISSN: 0192-253X
    Keywords: Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Type of Medium: Electronic Resource
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  • 19
    ISSN: 0192-253X
    Keywords: Heat shock promoters ; HSP70-CAT ; microinjection ; linker-scanner mutations ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Microinjected human HSP70 promoter-chloramphenicol acetyl transferase (CAT) chimeric genes are constitutively expressed immediately after the midblastula transition of Xenopus embryogenesis. Analysis of a series of 5′-deletion mutants in the HSP70 promoter revealed that sequences within 74 bases of the transcriptional start site were sufficient for strong basal activity. We investigated the role of specific sequences in the basal promoter by injecting HSP70-CAT vectors containing linker-scanner mutations in the basal elements (CCAAT, purine-rich element, GC-element, ATF/AP1, and 1ATA). Our data reveal that deletion of any of these cis-acting elements in the basal promoter prevents expression after the midblastula stage of development. Furthermore, we have identified specific binding activities in embryonic nuclear extracts that complex with basal promoter elements (CCAAT, ATF, and GC) of the heterologous HSP70 promoter. These trans-acting factors are detectable in nuclear extracts of early blastula embryos, and their respective binding activity increases dramatically after the midblastula transition. The expression of the human HSP70 gene after the midblastula transition of Xenopus embryogenesis requires an array of cisacting elements, which interact with specific Xenopus transcription factors.
    Additional Material: 9 Ill.
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  • 20
    Electronic Resource
    Electronic Resource
    Chichester [u.a.] : Wiley-Blackwell
    Developmental Genetics 11 (1990), S. 110-122 
    ISSN: 0192-253X
    Keywords: Embryonic cell surface ; glycoconjugates ; monoclonal antibodies ; developmental expression of glycoconjugates ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Molecular markers for specific cell lineages would be useful in studies of cellular differentiation. To isolate such markers monoclonal antibodies (MoABs) were raised against plasma membranes isolated from gastrulating Xenopus embryos. Those antibodies that recognized subsets of cells within the embryo were selected by indirect immunofluorescence. The analysis of eight such MoAbs is presented. Western blot analysis showed that all but one MoAb recognized a complex pattern of glycoconjugates associated with glycoproteins. All the antigens recognized by the MoAbs were maternal in origin and displayed similar spatial patterns of pregastrular expression. This pattern of immunoreactivity at the apical surface was inherited passively during cleavage by the resulting superficial blastomeres suggesting that ectodermal specific markers of maternal origin are pre-localized to the cortical ooplasm in mature oocytes. We suggest that these maternal components may be specific glycosyl transferases. Three different patterns of expression were observed during gastrulation as exemplified by MoAbs 1F10C1, 3A4D1, and 6F10B6. MoAb 6F10B6 was specific for both neural and non-neural epithelium. MoAb 3A4D1 was specific for non-neural epidermis. MoAb 1F10C1 appeared to recognize a protein epitope on an extracellular component expressed by the superificial and involuting epithelial cells. The pattern of expression for the 1F10C1 antigen suggests that it may play a role in facilitating the movement of the involuting cells during gastrulation.
    Additional Material: 7 Ill.
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  • 21
    Electronic Resource
    Electronic Resource
    Chichester [u.a.] : Wiley-Blackwell
    Developmental Genetics 11 (1990), S. 125-132 
    ISSN: 0192-253X
    Keywords: Flow cytometry ; polytenization ; C value ; fluorescence ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Multiparametric flow cytometry was used to analyze the development of the endosperm in Zea mays L. during the period from 8 to 20 days after pollination (dap). Nuclear size, DNA content per nucleus, and frequencies of nuclei with varying properties were measured in preparations that included all of the endosperm nuclei of single kernels of the inbred strain Al88. Characteristics of nuclear populations from different kernels on the same ear showed minimal variation. The dynamic changes of non-mitotic cells involved in endosperm development consisted of alternating periods of DNA replication with non-replication. Seven rounds of DNA replication had occurred in some nuclei in the later developmental stages with the rate averaging approximately one round per 24-hour period. Analysis of the DNA levels in the nuclei showed an exact doubling pattern indicating an endoreduplication process, that is, replication of the entire genome during each round. The loosely organized polytenization of the chromatin occurred to varying extents among the nuclei within an endosperm. A weak positive correlation existed between DNA content and size of nuclei suggesting that DNA increases and nuclear growth may not be highly coordinated in this tissue. Increased proportions of the larger nuclei occurred in the later stages of endosperm development. Considering the entire endosperm, the average DNA content per nucleus at the 15-dap peak level was approximately 12.8 C constituting a 2.7-fold overall increase from 8 dap.
    Additional Material: 8 Ill.
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  • 22
    Electronic Resource
    Electronic Resource
    Chichester [u.a.] : Wiley-Blackwell
    Developmental Genetics 11 (1990), S. 176-181 
    ISSN: 0192-253X
    Keywords: Direct gene transfer ; transgenic plants ; expression of transgenes ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Direct gene transfer to protoplasts is one of several methods developed for the production of transgenic plants. This method utilizes the efficient uptake of DNA from the surrounding medium by protoplasts (cell wall-less plant cells). Where a suitable protoplast system exists large numbers of transformant clones can be efficiently produced and often regenerated to normal fertile plants. This review concentrates on the fate of the DNA which is taken up into the protoplasts. Particular emphasis is given to the factors which can influence the integration and form of the transferred DNA, the expression of transferred genes, and the inheritance in further generations of those genes. The information available suggests (1) that DNA is taken up by a large proportion of the cells in a transformation mixture, (2) that this DNA forms complexes sometimes involving carrier DNA, (3) that fewer cells actually take up DNA into the nucleus, and (4) that the complex may be rearranged and/or amplified and then integrated into the genome. If the DNA is arranged in such a way that a gene can be expressed it does so in a normal manner and is stably inherited both mitotically and meiotically.
    Type of Medium: Electronic Resource
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  • 23
    Electronic Resource
    Electronic Resource
    Chichester [u.a.] : Wiley-Blackwell
    Developmental Genetics 11 (1990), S. 197-204 
    ISSN: 0192-253X
    Keywords: Light-regulated genes ; transgenic plants ; enhancer ; silencer ; regulatory elements ; trans-acting factors ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Transgenic plants have been particularly useful in studying nuclear genes encoding for photosynthetic functions. The expression of these genes and their chimeric constructs in transgenic plants faithfully mimics their natural counterparts. The use of sensitive chimeric reporter genes has enabled localizing the activity of genes encoding photosynthetic proteins to individual cells. Cab and rbcS transgenes have been shown to retain sensitivity to light quality, which is modulated by phytochrome. Conditional light activation under the influence of a circadian rhythm has been shown for Cab transgenes. Transgenic plants containing truncated promoters have helped delineate cis-regulatory positive and negative elements involved in light-mediated transcriptional induction and tissue specificity.
    Additional Material: 1 Ill.
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  • 24
    ISSN: 0192-253X
    Keywords: nodulins ; leghemoglobin ; glutamine synthetase ; Enod2 ; cis-acting elements ; transacting factors ; Agrobacterium turnefaciens ; A. rhizogenes ; binary vectors ; plant transformation ; chimeric genes ; chloramphenicol acetyltransferase ; glucuronidase ; cytokinin induction ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Nodulin genes are plant genes specifically activated during the formation of nitrogen-fixing nodules on leguminous plants. These genes are interesting to study since they are not only induced in a specific developmental fashion by signals coming directly or indirectly from the rhizobial symbiont, but are also expressed in a tissue-specific manner. By examining the expression of chimeric nodulin-reporter genes in transgenic legume plants it has been shown that nodule specific expression is mediated by DNA sequences present in the 5′upstream region of several nodulin genes. Here we summarize the available data on these cis-acting elements and the trans-acting factors interacting with them. We also review experiments designed to identify rhizobial “signals” which may play a role in nodule specific gene expression.
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  • 25
    Electronic Resource
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    Chichester [u.a.] : Wiley-Blackwell
    Developmental Genetics 11 (1990), S. 205-213 
    ISSN: 0192-253X
    Keywords: cauliflower mosaic virus 35S RNA ; cell cycle gene expression ; cis-acting element ; wheat histone H3 gene ; trnas-acting factor ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: A majority of histone genes are expressed in the S phase during the cell cycle. Using the gene expression system of transformed sunflower cells into which wheat histone H3 gene was introduced by the Ti-plasmid gene transfer technique, we determined three cis-acting control sequences (hexameric, octameric, and nonameric motifs) which seemed to confer the S-phase-specific transcription of wheat histone genes. Furthermore, as candidates for regulatory transcription factors, three nuclear DNA-binding proteins HBP-la, HBP-lb, and HBP-2 that interact with the hexameric and nonameric motifs were identified. The structural analysis of the cDNA of HBP-la revealed that a nuclear protein has the leucine-zipper structure and a DNA-binding motif. The hexameric motif in the H3 gene was also seen in cauliflower mosaic virus 35S (CaMV 35S) promoter and shown to function as a regulatory element of this promoter. The wheat HBP-1b can interact with the hexameric motif of the CaMV 35S promoter. Much attention has been paid to the significance of the hexameric sequences within the H3 and CaMV 35S promoters and the DNA-binding proteins HBP-la and HBP-lb.
    Additional Material: 4 Ill.
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  • 26
    Electronic Resource
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    Chichester [u.a.] : Wiley-Blackwell
    Developmental Genetics 11 (1990) 
    ISSN: 0192-253X
    Keywords: Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Type of Medium: Electronic Resource
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  • 27
    ISSN: 0192-253X
    Keywords: lux ; luc reporter genes ; light emission ; gene expression ; single photon imaging in vivo ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Genes encoding light-emitting luciferase were recently isolated from luminous marine bacteria and fireflies. Expression of luciferase genes in diverse organisms is a unique way for studying gene expression by simple and sensitive measurement of light. Recent advances in application of luciferase reporter genes are reviewed and documented by examples of in vivo visualization of their expression in transgenic plants.
    Additional Material: 2 Ill.
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  • 28
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    Chichester [u.a.] : Wiley-Blackwell
    Developmental Genetics 11 (1990), S. 214-223 
    ISSN: 0192-253X
    Keywords: Gene interactions ; cytosine methylation ; epigenetic variation ; transgenic plants ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Unusual gene interactions were observed in several doubly transformed tobacco plants which were obtained following sequential transformation steps using two T-DNAs encoding different selection and screening markers. The expression of T-DNA-I, which encoded kanamycin resistance (Kanr) and nopaline synthase (NOS), was suppressed in some, but not all, of the double transformants after the introduction of T-DNA-II, which encoded hygromycin resistance (Hygr) and octopine synthase (OCS). Double transformants in which T-DNA-I had been inactivated could produce KanrNOS+ progeny, but these were shown to lack T-DNA-II, thus establishing the role of this T-DNA in the suppression of T-DNA-I. Reversible cytosine methylation of the promoters of T-DNA-I genes was shown to correlate with their activation/inactivation cycle. In this paper we pursue further the questions of the mechanism of suppression of T-DNA-I genes by T-DNA-II, and also the timing and extent of demethylation of T-DNA-I promoters in Kanr progeny following the loss of T-DNA-II. We propose that the suppression is due to the competition between homologous regions on each T-DNA for binding to nuclear sites with fixed locations. We further suggest that incomplete demethylation patterns of T-DNA-I promoters in Kanr progeny reflect the existence in the shoot apex meristem of two cell populations, which have either methylated or unmethylated T-DNA-I promoters, respectively. Thus, Kanr progeny are epigenetic chimeras with respect to the expression of T-DNA-I genes.
    Additional Material: 7 Ill.
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  • 29
    Electronic Resource
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    Chichester [u.a.] : Wiley-Blackwell
    Developmental Genetics 11 (1990), S. 233-247 
    ISSN: 0192-253X
    Keywords: Chromatin structure ; foreign gene expression ; transgenic plants ; Nicotiana tabacum ; position effect ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: The chromatin structure of foreign genes in transgenic tobacco plants was investigated by digestion of nuclei with DNase I and micrococcal nuclease, respectively, followed by restriction and Southern analysis of the digestion products. The results were compared to the differential expression of the different transgenes. Two model systems were used: plants harbouring vector DNA derived from the disarmed vector pGV 3850 and plants harbouring the light-regulated and organ-specifically expressed potato ST-LS1 gene and the cotransferred ncpaline synthase (nos) reporter gene. Our results show that transferred genes are located in DNase l-sensitive domains in all transformants. Slight variations of DNase l-sensitivity of the transferred ST-LS1 constructs in different transformants neither reflected the between-transformant variability of expression nor the organ-specific activity of the transgenes. A deletion event was found responsible for silencing the ST-LS1 gene but not the nos gene in one of the transformants. Whereas no DNase l-hypersensitive sites were found within the 3850-T-DNA and the ST-LS1 gene, one prominent site was mapped to the nos promoter within the ST-LS1 construct in all transformants. Digestion of chromatin harbouring 3850-T-DNA with micrococcal nuclease resulted in a blurred nucleosomal pattern as compared to nucleolar and bulk chromatin, the extent of blurring being independent of the expression of transferred genes. The present results favour the “permissive domain” hypothesis which capitalizes on the chromatin surrounding the integration site as the determining factor for the chromatin structure of incoming alien genes. However, between-transformant variability of expression is not reflected by differential sensitivity to DNase I. Hence, other factors than chromatin structure must be involved in creating “position effects”.
    Additional Material: 9 Ill.
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  • 30
    Electronic Resource
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    Chichester [u.a.] : Wiley-Blackwell
    Developmental Genetics 11 (1990), S. 249-253 
    ISSN: 0192-253X
    Keywords: Drosophila ; embryonic development ; sex differentiation ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: We report the first practical application of a genetic scheme devised for the purpose of obtaining large quantities of embryos of a specific sex. The scheme, which is based on the meiotic drive system Segregation Distorter, results in the production of populations of zygotes that are almost exclusively of one sex. We have used this scheme to determine that the steady-state levels of transcripts of X-linked genes are the same in early male and female embryos, establishing that these genes are dosage compensated.
    Additional Material: 2 Ill.
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  • 31
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    Chichester [u.a.] : Wiley-Blackwell
    Developmental Genetics 11 (1990), S. 263-269 
    ISSN: 0192-253X
    Keywords: Penetrance ; reciprocal effects ; suppressor genes ; developmental cycle in mammals ; initialization ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: The phenomenon of gametic imprinting in mammals has raised developmentally relevant questions concerning the manifestation and inheritance of genes with variable penetrance. The dominant fused (Fu) gene located on chromosome 17 is one of the few good cases demonstrating the phenomenon in mice. The Fu mutation has a maternal effect.We have previously shown that the † 12 haplotype significantly lowers the penetrance of Fuin ♀ ♀ Fu/†12 offspring. Results of recipiocal matings of the heterozygotes for Fu indicated that the Fu of maternal origin has a lowered level of penetrance. The dominant suppressors locotad outside chromosome 17, in contrast to †12 residing in it, had stronger effects on the manifestation of Fu, decreasing its penetrance to 8-17%. Experimental evidence is presented that the pathway via which Fu passes to the zygote nucleus during gametogenesis through successive generations has a marked effect on its penetrance. Based on this evidence, patterns of genetic imprinting are described. A survey of genetic imprinting allowed us to distinguish two developmental phases, gametic and zygotic. The hypothesis for the gametic phase of the development of multicellular organisms suggests that it proceeds from initialization, a process thought to ensure the freeing of chromosomes from redundant epigenetic information and their preparation for the consecutive developmental cycle.
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  • 32
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    Chichester [u.a.] : Wiley-Blackwell
    Developmental Genetics 11 (1990), S. 318-325 
    ISSN: 0192-253X
    Keywords: Mice ; housekeeping genes ; liver ; tissue specificity ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: In situ hybridization using nucleic acid probes was used to detect cell- and tissue-specific transcript(s) of embryonic genes during development and differentiation. This highly sensitive technique has the potential to provide valuable information on the regulation of low-abundance housekeeping genes during development. We have determined the experimental conditions required to detect the catalase message in adult mouse liver. Catalase effects the breakdown of H2O2 to O2 and H2O and offers protection against the toxic effects of oxygen radicals. We used a cloned 550 bp BamHI-Pstl fragment from a mouse catalase cDNA (pMCT-1) to generate 35S-labeled sense and antisense riboprobes. The experimental conditions used were sensitive enough to quantitate the abundance of silver grains generated by the antisense riboprobe on the adult liver, a tissue known to be positive for this message. The hybridization protocol was applied to serial sections of 13- and 18-day-old mouse embryos. The results suggest that the catalase expression in the liver and brain begins with somite formation and increases with development and differentiation. On the other hand, this message appears to be absent in mesenchyme, particularly in day 13 embryos. The message in positive tissues appears evenly distributed throughout the cell. The observed expression of the catalase message in the adult liver is approximately six times that in the embryonic liver. It is compatible with the enzyme activity results and emphasizes the sensitivity of the in situ hybridization method (over northern blot, etc.) used in this study.
    Additional Material: 5 Ill.
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  • 33
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    Chichester [u.a.] : Wiley-Blackwell
    Developmental Genetics 11 (1990), S. 328-332 
    ISSN: 0192-253X
    Keywords: Cellular slime molds ; cytoskeleton ; actin-binding proteins ; review ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: The cellular slime mold Dictyostelium discoideum is becoming the premier system for the explication of the biochemical and cellular events that occur during motile processes. Proteins associated with the actin cytoskeleton, in particular, appear to play key roles in cellular responses to many external stimuli. This review summarizes our present understanding of the actin-associated proteins in Dictyostelium, including their in vitro activities and their structural and/or functional analogues in mammalian cells.
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  • 34
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    Chichester [u.a.] : Wiley-Blackwell
    Developmental Genetics 11 (1990), S. 333-340 
    ISSN: 0192-253X
    Keywords: Dictyostelium discoideum ; actin ; ABP-120 ; elongation factor 1 alphc ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: In this work we evaluate the cartical expansion model for amoeboid chemo-taxis with regard to new information about molecular events in the cytoskeleton following chemo-tactic stimulation of Dictyostelium amoebae. A rapid upshift in the concentration of chemoattrac-tant can be used to synchronize the motile behavior of a large population of cells. This synchrony presents an opportunity to study the biochemical basis of morphological changes such as pseudopod extension that are required for amoeboid chemotaxis. Changes in the composition and activity of the cytoskeleton following stimulation can be measured with precision and correlated with important morphological changes. Such studies demonstrate that activation of actin nucleation is one of the first and most crucial events in the actin cytoskeleton following stimulation. This activation is followed by incorporation of specific actin cross-linking proteins into the cytoskeleton, which are implicated in the extension of pseudopods and filopods. These results, as well as those from studies with mutants deficient in myosin, indicate that cortical expansion, driven by focal actin polymerization, cross-linking and gel osmotic swelling, is an important force for pseudopod extension.It is concluded that whereas three forces, frontal sliding, tail contraction, and cortical expansion may cooperate to produce amoeboid movement, the cortical expansion model offers the simplest explanation of how focal stimulation with a chemoat-tractant causes polarized pseudopod extension.
    Additional Material: 6 Ill.
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  • 35
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    Chichester [u.a.] : Wiley-Blackwell
    Developmental Genetics 11 (1990), S. 362-368 
    ISSN: 0192-253X
    Keywords: Dictyostelium ; filopodia ; actin binding proteins ; calcium ; cell motility ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: The Dictyostelium discoideum 30,000 dalton actin-binding protein is an actin cross-linking protein that organizes formation of parallel bundles of actin filaments in vitro, and is present in filopodia in living cells. This protein binds calcium directly and exhibits a decreased affinity for actin filaments in the presence of micromolar calcium. In this work, the existence of antigenic homologs of the 30,000 dalton protein in Physarum polycephalum, Schistosoma mansoni, Chara carolina, and Drosophila melanogaster is detected by use of affinity purified antibody and electrophoretic blotting methods. The expression of this protein during development of Dictyostelium is also analyzed, revealing a progressive 3-fold decrease in the level of this protein in amoebae between the vegetative and slug stages. A highly ordered structure of bundles of actin and the 30,000 dalton protein formed in vitro is inferred from the presence of transverse striations on the bundles with a minor periodicity at 11.4 nm and a major periodicity at 33.9 nm. Finally, we propose a working model of the interaction of this actin cross-linking protein with actin filaments to form bundles.
    Additional Material: 5 Ill.
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  • 36
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    Chichester [u.a.] : Wiley-Blackwell
    Developmental Genetics 11 (1990), S. 369-376 
    ISSN: 0192-253X
    Keywords: Cytoskeleton ; capping proteins ; genamic structure ; Dictyostelium ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: The two subunits of the het-erodimeric protein cop32/34, an actin-binding protein, are encoded by separate single-copy genes. We have established the genomic structure of both genes. A sequence comparison of cap32/34 with capZ from chicken skeletal muscle and two partially known sequences from Saccharomyces cerevisiae and Xenopus laevis show that heterodimeric capping proteins belong to a highly conserved group of actin-binding proteins. This conclusion is supported by the cross-reaction of polyclonal antibodies against cap32 and cap34 with proteins from lower and higher eukaryotes. In addition, a system is presented that allows the expression of truncated cap34 polypeptides under the control of the cap34 promoter.
    Additional Material: 5 Ill.
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  • 37
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    Chichester [u.a.] : Wiley-Blackwell
    Developmental Genetics 11 (1990), S. 388-390 
    ISSN: 0192-253X
    Keywords: Transformation ; electroporotion ; gene targetting ; antisense RNA ; complementation ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: DNA mediated transformation is a critical technique for uniting genetic, molecular genetic and reverse genetic approaches to a wide range of problems in cell and molecular biology. In Dictyostelium, there is now the capability not only to manipulate DNA sequences in vitro and put them back into the cell, but also to alter the sequences of endogenous chromosomal genes through high frequency homologous recombination. This means that the range of gene manipulation techniques that have made yeast such a useful system should now be applicable in Dictyostelium. For studying problems such as cellular motility, morphogenesis, and gene regulation, few organisms have the combination of features offered by Dictyostelium.
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  • 38
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    Chichester [u.a.] : Wiley-Blackwell
    Developmental Genetics 11 (1990), S. 391-395 
    ISSN: 0192-253X
    Keywords: Lipofectin ; slime mould ; transformation ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: We have studied the transient expression, in Dictyostelium cells growing on a bacterial food source, of a construct containing the coding region of the firefly luciferase gene inserted downstream of a Dictyosteliumactin promoter. The fusion gene is not detectably expressed when DNA is introduced by calcium phosphate precipitation or by electroporation, but it is expressed when introduced using cationic liposomes (lipofectin). Using this latter procedure, we are able to transform cells with a G418 resistance vector and select stable, drug-resistant transformants at a relatively low, but workable, efficiency. This technique will allow molecular genetics to be applied to the many important nonaxenic Dictyostelium strains.
    Additional Material: 2 Ill.
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  • 39
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    Chichester [u.a.] : Wiley-Blackwell
    Developmental Genetics 11 (1990), S. 377-387 
    ISSN: 0192-253X
    Keywords: Cell-cell ; adhesion molecule ; cell binding site ; cell aggregation ; development ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: During development of Dictyostelium discoideum, cells acquire EDTA-resistant cell-cell adhesion at the aggregation stage. The EDTA-resistant cell binding activity is associated with a cell surface glycoprotein of Mr 80,000 (gp80), which mediates cell-cell binding via ho-mophilic interaction. Analysis of the structure of gp80 deduced from cDNA sequence reveals the presence of three internally homologous segments in the NH2-terminal domain, which also contains regions with homology to the neural cell adhesion molecule. Secondary structure predictions show an abundance of β-structures and very few α-helices. This is confirmed by circular dichroism measurements. It is likely that the homologous segments are organized into globular structures, extended from the cell surface by a Pro-rich stalk domain. The cell binding activity of gp80 resides within the first globular repeat of the NH2-terminal domain and has been mapped to a 51 amino acid region between Val123 and Leu 173. Synthetic oligopeptides corresponding to sequences within this region have been prepared and assayed for their ability to bind to cell surface gp80. Results lead to identification of the homophilic binding site to an octapeptide sequence within this region. Synthetic peptides containing this octapeptide sequence and univalent antibodies directed against this site block the formation of organized cell streams during aggregation. Although cell aggregates are eventually formed, most fail to undergo further development to give rise to slugs and fruiting bodies, indicating that cell-cell adhesion involving gp80 is an important step in normal morphogenesis.
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  • 40
    ISSN: 0192-253X
    Keywords: Transformation ; Dictyostelium discoideum ; UMP-synthase/ura-complementation ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Whereas transformation ofDictyostelium discoideum now can be done routinely and reliably, there is increasing demand for a system that allows selection of cells that have gone through repeated transformation cycles. Such a system is presented here. Selection is based on resistance to 5-fluoro-orotic-acid (5-FOA). D. discoi-deum is highly sensitive to this drug, and cells can survive only in the presence of 5-FOA if they carry a defective UMP-synthase gene. After transformation with a plasmid carrying a cloned version of the UMP-synthase gene, 5-FOA resistant cells are obtained at high frequency. Because 5-FOA-resistant cells depend on exogenously added uracil, these cells can be taken through a second round of transformation. A plasmid-borne UMP-synthase gene renders 5-FOA-resistant cells phenotypically ura +. Finally, cells can be further transformed using the standard G418 selection system.
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  • 41
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    Chichester [u.a.] : Wiley-Blackwell
    Developmental Genetics 11 (1990), S. 425-426 
    ISSN: 0192-253X
    Keywords: Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
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  • 42
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    Chichester [u.a.] : Wiley-Blackwell
    Developmental Genetics 11 (1990), S. 403-409 
    ISSN: 0192-253X
    Keywords: Electroporation ; soft-agarose ; G418 ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: A new method for clonal growth of Dictyostlium axenic amoebae has been developed. Cells are plated in growth medium containing 1% ultra-low gelling temperature agarose. Cells grow normally in the agarose and form colonies up to several millimeters in diameter. When the colonies have grown to a sufficient size, they begin multicellular development. Pseudoplasmodia are formed, migrate to the surface of the agar, and then undergo fruiting body formation. Cells can be removed from the soft agarose colonies with a toothpick or by picking spores from the fruiting bodies. This method should be useful for drug, auxotrophic, and temperature selections where clonal maintenance of axenic colonies is important.This method has been used in combination with a selection for resistance to G418 to isolate independent colonies following DNA-mediated transformation. Several parameters in the calcium phosphate and electroporation transformation protocols have been optimized and the transformation frequency qualified. Independent transformed colonies are obtained at a frequency of 1 in 104 to 1 in 105 cells when integrating plasmids are introduced using calcium phosphate coprecipitation. The frequency is about tenfold higher when extrachromosomal shuttle vectors are introduced into cells.
    Additional Material: 5 Ill.
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  • 43
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    Chichester [u.a.] : Wiley-Blackwell
    Developmental Genetics 11 (1990), S. 410-417 
    ISSN: 0192-253X
    Keywords: Suppressor †RNA genes ; yeast ; lacZ expression in D. discoideum ; †RNATrp genes ; †RNAGlu genes ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: We describe the generation of Dictyostelium discoideum cell lines that carry different suppressor †RNA genes. These genes were constructed by primer-directed mutagenesis changing a †RNATrp(CCA) gene from D. discoideum to a †RNATrp(amber) gene and changing a †RNAGlu(UUC) gene from D. discoideum to a †RNAGlu(ochre) as well as a †RNAGlu(amber) gene. These genes were stably integrated into the D. discoideum genome together with a reporter gene. An actin 6::lacZ gene fusion carrying corresponding translational stop signals served as a reported. Active β-galactosidase is expressed only in D. discoideum strains that contain, in addition to the reporter, a functional suppressor †RNA. Both amber suppressors are active in D. discoideum without interfering significantly with cell growth and development. We failed, however, to establish cell lines containing a functional †RNAGlu(ochre) suppressor. This may be due to the fact that nearly every message from D. discoideum known so far terminates with UAA. Therefore a †RNA capable of reading this termination codon may not be compatible with cell growth.
    Additional Material: 7 Ill.
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  • 44
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    Chichester [u.a.] : Wiley-Blackwell
    Developmental Genetics 11 (1990), S. 418-424 
    ISSN: 0192-253X
    Keywords: Transcription ; development ; cAMP ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Genetic studies have revealed a network of three unlinked regulatory genes that control the developmental expression of the family of endogenous lectins in Dictyostelium discoideum. Mutations in the disA and disB loci have a null phenotype and do not express the discoidin I or II lectins. The third mutation, drsA, is a second-site suppressor of the disB mutation, which restores expression of all lectin species. Cells carrying this mutation express the discoidin lectins during growth, which is in contrast to wild-type cells in which lectin synthesis is developmentally regulated. In addition to this basic level of genetic control, the conditions of growth dramatically influence the patterns of discoidin expression. Growth-phase wild-type cells do not express lectin if the cells are grown on plates in association with bacteria. However, wild-type cells growing in bacterial suspensions express high levels of lectin during growth. Synthesis of the discoidin lectins in growing cells is sensitive to the levels of extracellular cyclic adenosine monophosphate (cAMP) and folic acid. These results suggest that the drsA mutation renders cells insensitive to cAMP and/or folate and thus allows expression of lectin during growth.
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  • 45
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    Chichester [u.a.] : Wiley-Blackwell
    Developmental Genetics 11 (1990), S. 439-441 
    ISSN: 0192-253X
    Keywords: Cellular slime molds ; timing of differentiation ; differentiation ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Vital dyes have been extensively used in many laboratories to distinguish the prestalk and prespore zones of a migrating slug in Dictyostelium discoideum. Here we present evidence that the two zones do not form immediately following aggregation, as is sometimes assumed, but only after 1-4.5 hr of migration. When the transition to the two-toned state occurs, it changes very quickly, in a matter of approximately 10 min. Furthermore, it can be rapidly induced by submerging the slug in mineral oil. The time of appearance of the zones is correlated with previously reported changes in the distribution of cyclic AMP secretion and with changes in regulation properties along the axis of the slug.
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  • 46
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    Chichester [u.a.] : Wiley-Blackwell
    Developmental Genetics 11 (1990), S. 442-446 
    ISSN: 0192-253X
    Keywords: Prestalk cells ; prespore cells ; slugs ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Ammonia appears to be an important regulatory signal for several aspects of the Dictyostelium life cycle. The postulated role of ammonia in the determination of the prespore pathway in cells of the slug stage has led us to examine the effect of ammonia on the prestalk/prespore ratio of migrating slugs. In the presence of 10-3 M ammonium chloride, the volume of the prestalk region decreases by 40.8%. The kinetics of the process make it unlikely that this is due to a shift in the differentiation pathway. A test of the hypothesis that the decrease in volume of the prestalk region is due to the conversion of prestalk cells to anterior-like cells shows that the percent of anterior-like cells in the posterior region increases by the amount predicted by the hypothesis. This suggests that ammonia may be the molecular signal, produced by the tip, that prevents anterior-like cells from chemotactically migrating to the tip and thereby becoming anterior cells. The effect of enzymatic removal of ammonia from vitally stained migrating slugs is the appearance of a series of dark stripes beginning at the posterior end and progressing forward. We interpret this as a result of progressive removal of anterior-like cells from tip dominance and essentially as the formation of new potential tips. Indeed, in a few cases one or even two of the stripes separate from the posterior of the cell mass and form small fruiting bodies. We consider the phenomenon of stripe formation further evidence that the tip acts on anterior-like cells through ammonia.
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  • 47
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    Chichester [u.a.] : Wiley-Blackwell
    Developmental Genetics 11 (1990), S. 427-438 
    ISSN: 0192-253X
    Keywords: Cellular slime molds ; patterning ; development ; immunoblotting ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: A series of monoclonal antibodies were previously raised against developing Polysphondylium pallidum cells. In this work, six of these antibodies have been used as probes to identify and characterize antigens regulated during development. Soluble and membrane fractions of P. pallidum cells at six stages of development or three stages of cyclic adenosine monophosphate (cAMP)-induced development were run in sodium dodecyl sulfate (SDS)-polyacrylamide gels and subjected to Western blot analysis. Three of the monoclonals, anti-Tp200, anti-Tp423, and antiPg 101, stain sorogen tips. Tp423 and Tp200 are membrane-associated antigens; both are stable to urea extraction, and Tp200 remains in the membrane after NaOH extraction. Tp423 is present in starved cells but is more prominent in sorogens and particularly in cAMP-developed cells. In contrast, Tp200 is first detected in early to mid-aggregation and is more abundant late in development. Pg101, which is expressed as a gradient with its highest concentration in tips, first appears in tight aggregates but is much more abundant in sorogens; unlike the Tp antigens, Pg101 is not greatly induced in cAMP-developed cells. All three of these antigens undergo changes in apparent molecular weight at the tight aggregate or sorogen stage: The gel mobilities of Tp200 and Pg101 increase, whereas that of Tp423 decreases. In addition to the tip-specific monoclonals, two monoclonals that stain all but the tips of sorogens have been used for analysis. One of these, anti-3D 10Pnk stains most cells within secondary tips, whereas anti-3D 10Dif does not. 3D 10Dif is membrane associated; it is present very early in development, increasing two- to threefold through the sorogen stage and diminishing in late cAMP-developed cells. 3D 10Pnk is a mostly soluble species first detected in late streaming. Anti-1c3, a sixth monoclonal, which stains nuclei uniformly throughout sorogens, is also developmentally expressed. 1c3 is mainly membrane associated and is expressed from late streaming through the sorogen stage.
    Additional Material: 6 Ill.
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  • 48
    Electronic Resource
    Electronic Resource
    Chichester [u.a.] : Wiley-Blackwell
    Developmental Genetics 11 (1990), S. 453-453 
    ISSN: 0192-253X
    Keywords: Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
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  • 49
    Electronic Resource
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    Chichester [u.a.] : Wiley-Blackwell
    Developmental Genetics 11 (1990), S. 447-452 
    ISSN: 0192-253X
    Keywords: Pattern regulation ; cell sorting ; transplantation ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: In classical transplantation experiments, we have attempted to distinguish between cell sorting and position-dependent mechanisms of pattern formation during the regulation of amputated prespore regions of Dictyostelium slugs. Host and transplanted tissue were distinguished by prior labelling with 14C and 3H, and their distributions in the manipulated slugs were determined by double-label scintillation counting of serial sections of the slugs. First, we showed that the regenerated prestalk region is formed predominantly from the anterior of a prespore isolate. This disproves extreme cell sorting models, which hold that the new prestalk cells arise at random in the prespore mass, before sorting to the prestalk zone. Second, we showed that a proportion of posterior cells, which are not normally recruited into the new prestalk region, can be recruited into it if they are transplanted into the anterior of the prespore isolate. Recruitment of these cells occurs only during the first hour of regulation. We believe that these experiments prove a position-dependent mechanism of pattern formation during slug regulation. This in turn implies the existence of localized inductive signals in the regulating fragments. Although cell sorting occurs efficiently when cells are misplaced in the slug, it does not appear to play a major role in reforming the prestalk/prespore pattern.
    Additional Material: 4 Ill.
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  • 50
    ISSN: 0749-503X
    Keywords: Hansenula polymorpha ; methanol ; glycerol ; dihydroxyacetone ; xylose ; alcohol oxidase ; continuous culture ; regulation ; mutants ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: The physiological responses of Hansenula polymorpha wild-type and mutant strains 17B (dihydroxyacetone kinase-negative) and 17BG51 (dihydroxyacetone kinase- and glycerol kinase-negative) to growth on mixtures of xylose and methanol in chemostats were investigated. Increasing methanol concentrations (0-110 mM) in the feed of the wild-type culture resulted in increasing cell densities and a gradual switch towards methanol metabolism. At the lower methanol feed concentrations the mutant cultures used methanol and xylose to completion and changes in enzyme patterns comparable to the wild type were observed. This was not reflected in significant changes in cell densities. Instead, formaldehyde assimilation resulted in dihydroxyacetone (DHA) production, which was proportional to the amount of methanol added. At intermediate methanol concentration the cultures showed a strong variation in DHA levels and cell densities. Further increased in the methanol feed concentrations resulted in a drop in DHA accumulation rates, repression of alcohol oxidase synthesis and accumulation of residual methanol. The phenomena were studied in more detail in transition experiments and with gradients of methanol. The results indicate that xylulose-5-phosphate (Xu5P) generated in xylose metabolism served as acceptor molecule for formaldehyde assimilation by the peroxisomal enzyme DHA synthase. Accumulation of DHA in the mutant cultures, however, further diminished the availability of carbon for growth. The data suggest that with increasing methanol concentrations Xu5P eventually became growth rate limiting. This resulted in an unstable situation but wash-out of the culture did not occur to a significant extent. Instead, DHA accumulation ceased and cell densities, and enzymes specifically involved in xylose metabolism increase, indicating that the organism resumed its xylose metabolism. The molecular mechanisms controlling the partitioning of Xu5P over xylose (pentose phosphate pathway) and methanol (peroxisome) metabolism under these conditions remain to be elucidated.
    Additional Material: 4 Ill.
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  • 51
    ISSN: 0749-503X
    Keywords: Hansenula polymorpha ; methanol ; dihydroxyacetone ; xylose ; mutants ; transketolase ; formaldehyde ; continuous culture ; peroxisome ; regulation ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Contrary to expectation, a mutant of Hansenula polymorpha blocked in dihydroxyacetone (DHA) synthase was able to assimilate methanol-carbon when grown in chemostat culture on mixtures of xylose and methanol. Incubation of a DHA synthase- and DHA kinase-negative double mutant resulted in DHA accumulation, indicating that a DHA synthase-type of reaction was involved. Low residual DHA synthase activity subsequently was shown to be present when using an assay with improved sensitivity. This activity was not associated with the (mutated) DHA synthase protein, which was still present in the peroxisomes, but with the enzyme transketolase. Transketolase from methanol grown cells was purified (525-fold) to homogeneity in 9% yield. The native enzyme was dimeric, as has been reported fro other transketolases, with a subunit molecular weight of 74000. The affinity of the purified enzyme for formaldehyde was low (Km = 5 mM), but high for xylulose-5-phosphate (ca. 10 μM). The in vivo functioning of transketolase in formaldehyde assimilation, and the influence of the hydration state of formaldehyde is discussed.
    Additional Material: 4 Ill.
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  • 52
    ISSN: 0749-503X
    Keywords: Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
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  • 53
    ISSN: 0749-503X
    Keywords: linear plasmids ; killer ; stability ; mating type locus control ; a1-α2 repressor ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: The linear killer plasmids, pGKL1 and pGKL2, from Kluyveromyces lactis stably replicated in mitochondria1 DNA-deficient (ρ0) MATa or MATα haploids of Saccharomyces cerevisiae, but were unstable and frequently lost in ρ° MATa/MATα diploids, suggesting that the replication of pGKL plasmids was under the control of the MAT locus. In MATa/MATα cells of S. cerevisiae, the MATα gene product (α2) is combined with the MATa gene product (a1) and the resultant protein, a1-α2, acts to repress the expression of haploid specific genes. Experiments showed that the K. lactis linear plasmids were stably maintained in ρ0 mata1/MATα diploids, indicating that the a1-α2 repressor interfered with the stability of pGKL2. It was revealed by computer analysis that the consensus sequence homologous to the a1-α2 repressor binding site occurred within the coding regions of pGKL2 genes which were presumed to be essential for the plasmid replication. Since the plasmids were stably maintained in diploids of K. lactis, the mating type control must not be working there.
    Additional Material: 2 Ill.
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  • 54
    Electronic Resource
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    New York, NY [u.a.] : Wiley-Blackwell
    Yeast 6 (1990), S. 455-460 
    ISSN: 0749-503X
    Keywords: Ornithine decarboxylase ; putrescine ; SPE1 gene ; spe10 mutation ; chromosome XI ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: The gene for ornithine decarboxylase in Saccharomyces cerevisiae, SPE1, has been assigned to chromosome XI by the technique of transverse alternating pulsed field electrophoresis and DNA-DNA hybridization. Genetic mapping by tetrad analysis shows that the SPE1 gene is located on the left arm of chromosome XI, 6 cM from the LAP1 gene and 43 cM from the TRP3 gene. The spe10 mutation previously isolated in this laboratory is mapped to the N-terminal region of the SPE1 gene, and therefore should be designated as a spe1 allele.
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  • 55
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    New York, NY [u.a.] : Wiley-Blackwell
    Yeast 6 (1990) 
    ISSN: 0749-503X
    Keywords: Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
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  • 56
    ISSN: 0749-503X
    Keywords: Omnipotent suppressor ; gene structure ; evolutionary conservation ; codon bias analysis ; Pichia pinus ; yeast ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: SUP2(SUP35) is an omnipotent suppressor gene, coding for an EF-1α-like protein factor, intimately involved in the control of translational accuracy in yeast Saccharomyces cerevisiae.In the present study a SUP2 gene analogue from yeast Pichia pinus was isolated by complementation of the temperature-sensitive sup2 mutation of S. cerevisiae.The nucleotide sequence of the SUP2 gene of P. pinus codes for a protein of 82·4 kDa, exceeding the Sup2 protein of S. cerevisiae by 6 kDa. Like the SUP2 gene product of S. cerevisiae, the Sup2 protein of P. pinus represents a fusion of a unique N-terminal part of a region homologous to EF-1α. The comparison of amino acid sequences of the Sup2 proteins reveals high conservations (76%) of the C-terminal region and low conservation (36%) of the N-terminal part where, in addition, the homologous correspondence is ambiguous.Proteins related to the Sup2 of S. cerevisiae where found in P. pinus and some other yeast species by the immunoblotting technique.The relation between the evolutionary conservation of different regions of the Sup2 protein and their functional significance is discussed.
    Additional Material: 7 Ill.
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  • 57
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    New York, NY [u.a.] : Wiley-Blackwell
    Yeast 6 (1990), S. 535-536 
    ISSN: 0749-503X
    Keywords: Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
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  • 58
    ISSN: 0749-503X
    Keywords: Cell wall porosity ; permeability ; mannan ; cell wall composition ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: The cell porosity of batch-grown Saccharomyces cerevisiae was maximal in the early exponential phase and fell off rapidly to lower levels in later growth phases.Treatment of stationary-phase cells with alpha-mannosidase restored wall porosity to the level of cells in early exponential phase. When cells in the early exponential phase were treated with alpha-mannosidase, or tunicamycin, an inhibitor of N-glycosylation, even higher porosities were obtained. Mutants with truncated mannan side-chains in their wall proteins also had very porous walls. The importance of the mannan side-chains for wall porosity was also seen during sexual induction. Treatment with alpha pheromone, which leads to the formation of wall proteins with shorter mannan side-chains, enhanced wall porosity.Disulphide bridges also affect cell wall porosity. They were predominantly found in the glucanase-soluble wall proteins. Because the main part of the mannan side-chains is also found in this family of wall proteins, our results demonstrate that the glucanase-soluble mannoproteins limit cell wall porosity in yeast.
    Additional Material: 1 Ill.
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  • 59
    ISSN: 0749-503X
    Keywords: Hansenula polymorpha ; peroxisomes ; peroxisome-deficient mutants ; amine oxidase ; D-amino acid oxidase ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: We have studied growth of two peroxisome-deficient mutant strains of Hansenula polymorpha on glucose in the presence of different organic nitrogen sources (methylamine, ethylamine and D-alanine), the metabolism of which is mediated by peroxisome-borne oxidases in wild-type (WT) cells. Both strains grew well on each of these substrates with growth rates comparable to WT cells. Growth on both methylamine and ethylamine was associated with enhanced levels of catalase and amine oxidase in the cells; in D-alanine-grown cells D-amino acid oxidase activity and increased. In WT cells of H-polymorpha the activities of these enzymes were confined to the peroxisomal matrix; however, in both peroxisome-deficient strains their activities were localized in the cytosol. Electron microscopy indicated that, dependent on the stage of growth, the enzymes may form large protein aggregates.The molecular masses of both amine oxidase and D-amino acid oxidase in the mutant strains were identical to their respective counterparts in WT cells, indicating that both proteins were correctly assembled and active in the cytosol.
    Additional Material: 9 Ill.
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  • 60
    Electronic Resource
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    New York, NY [u.a.] : Wiley-Blackwell
    Yeast 6 (1990), S. 1-29 
    ISSN: 0749-503X
    Keywords: Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Additional Material: 10 Ill.
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  • 61
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    New York, NY [u.a.] : Wiley-Blackwell
    Yeast 6 (1990), S. 35-43 
    ISSN: 0749-503X
    Keywords: Hanseula polymorpha ; Candida boidinii ; peroxisomes ; peroxisomal membrane proteins ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: We have the substructure and polypeptide composition of the peroxisomal membranes in two methylotrophic yeasts in relation to different growth conditions. The results obtained that no significant ultrastructural differences existed between the membranes of variously grown cells.The presence of specific peroxisomal membrane proteins (PMPs) was studied biochemically. On sodium dodecyl sulphate-polyacrylamide gels of purified microbody membranes isolated from methanol-grown Hansenula polymorpha, prominent proteins bands were observed at 22, 31, 35, 42, 49 and 51 kD. These proteins were also present when the cells were grown in media containing ethanol and/or ethylamine. Apart from these, several other PMPs were specifically induced under these conditions, namely 24, 29, 37 and 62 kD proteins. The polypeptide composition of peroxisomal membranes from H. polymorpha was compared with that of another methylotroph. Candida biodinii. In the latter organism a specific PMP with a molecular weight of 23 kD was induced during growth on D-alanine instead of ammonium sulphate as the nitrogen source.
    Additional Material: 8 Ill.
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  • 62
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    New York, NY [u.a.] : Wiley-Blackwell
    Yeast 6 (1990) 
    ISSN: 0749-503X
    Keywords: Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
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  • 63
    ISSN: 0749-503X
    Keywords: ARS elements ; shuttle vectors ; DNA replication ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: We have undertaken a search for autonomously replicating (ARSs) from Kluvermyces lactis chromosomal DNA able to sustain plasmid replication in K. lactis and in Saccharomyces cerevisiae. The discovery of such sequences might be interesting for the comparison of ARSs from different sources and possibly useful for the construction of multivalent vectors.HindIII fragments from K. lactis chromosomal DNA were inserted in the YIp5 plasmid (lacking an origin of replication) and the resulting chimaeric plasmids were selected for the ability to transform S. cerevisiae. Four plasmids were identified and further analysed. Two contained the same 1.8 kb K. lactis fragment and transformed both K. lactis and S. cerevisiae with the same efficiency and stability, whereas the third transformed only S. cerevisiae and the fourth transformed K. lactis with a higher efficiency than S. cerevisiae. A detailed study was performed on the 1.8kb fragment which exhibited ARS function in both yeasts. The fragment was subcloned using different restriction enzymes and Bal31 exonuclease. Subclones were tested for ARS function. ARS activities in the two yeasts were localized in the same 100 bp region. Sequencing demonstrated the presence in this region of the dodecanucleotide 5′ATTTATTGTTTT 3′ differing from the ARS core consensus of S. cerevisiae only by a T insertion.A similar nucleotide sequence of S. cerevisiae only in the putative replication origin of the 2 μ-like plasmid pkD1 which stably replicates in K. lactis. Homologies with ARSs from S. cerevisiae were also found in the regions flanking the above-mentioned dodecanucleotide.
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  • 64
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    New York, NY [u.a.] : Wiley-Blackwell
    Yeast 6 (1990), S. 139-139 
    ISSN: 0749-503X
    Keywords: Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Additional Material: 1 Ill.
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  • 65
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    New York, NY [u.a.] : Wiley-Blackwell
    Yeast 6 (1990), S. 141-148 
    ISSN: 0749-503X
    Keywords: Pachysolen ; ornithine carbamoyltransferase ; xylose ; yeast ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: A fragment of DNA from a yeast Pachysolen tannophilus, bearing the ornithine carbamoyltransferase gene (OCTase, EC 2.1.3.3) has been cloned from a genomic library by functional complementation of the Escherichia coli OCT-negative mutant. The gene was located within the cloned segment of DNA and its coding sequence identified by DNA sequencing. This has indicated that P. tannophilus OCT gene encodes a 347 amino acid polypeptide, which shows 60% identity to the homologous Saccharomyces cerevisiae protein. The amino acid composition of its N-terminus indicates that this protein is translocated across the mitochondrial membrane. The gene can be expressed in E. coli as well as in S. cerevisiae. Comparison with other OCTases confirms a high degree of conservation among these proteins.
    Additional Material: 3 Ill.
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  • 66
    ISSN: 0749-503X
    Keywords: Maltose transport ; α-glucosidase ; yeast ; chemostat ; cell death ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: When Saccharomyces cerevisiae CBS 8066 was grown under maltose limitation, two enzymes specific for maltose utilization were present: a maltose carrier, and the maltose-hydrolysing α-glucosidase. The role of these two enzymes in the physiology of S. cerevisiae was investigated in a comparative study in which Candida utilis CBS 621 was used as a reference organism.Maltose pulses to a maltose-limited chemostat culture of S. cerevisiae resulted in ‘substrate-accelerated death’. This was evident from: (1) enhanced protein release from cells: (2) excretion of glucose into the medium; (3) decreased viability. These effects were specific with respect to both substrate and organism: pulses of glucose to maltose-limited cultures of S. cerevisiae did not result in cell death, neither did maltose pulses to maltose-limited cultures of C. utilis. The maltose-accelerated death of S. cerevisiae is most likely explained in terms of an uncontrolled uptake of maltose into the cell, resulting in an osmotic burst. Our results also provide evidence that the aerobic alcoholic fermentation that occurs after pulsing sugars to sugar-limited cultures of S. cerevisiae (short-term Crabtree effect) cannot solely be explained in terms of the mechanism of sugar transport. Both glucose and maltose pulses to maltose-limited cultures triggered aerobic alcohol formation. However, glucose transport by S. cerevisiae occurs via facilitated diffusion, whereas maltose entry into this yeast is mediated by a maltose/proton symport system.
    Additional Material: 8 Ill.
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  • 67
    Electronic Resource
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    New York, NY [u.a.] : Wiley-Blackwell
    Yeast 6 (1990) 
    ISSN: 0749-503X
    Keywords: Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
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  • 68
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    New York, NY [u.a.] : Wiley-Blackwell
    Yeast 6 (1990), S. 193-204 
    ISSN: 0749-503X
    Keywords: Kluyveromyces lactis ; alcohol dehydrogenase ; gene regulation ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: We have studied the alcohol dehydrogenase (ADH) system in the yeast Kluyvefromyces lactis. Southern hybridization to the Saccharomyces cerevisiae ADH2 gene indicates four probable structural ADH genes in K. lactis. Two of these genes have been isolated from a genomic bank by hybridization to ADH2. The nucleotide sequence of one of these genes shows 80% and 50% sequence identity to the ADH genes of S. cerevisiae and Schizosaccharomyces pombe respectively. One K. lactis ADH gene is preferentially expressed in glucose-grown cells and, in analogy to S. cerevisiae, was named K1ADH1. The other gene, homologous to K1ADH1 in sequence, shows an amino-terminal extension which displays all of the characteristics of a mitochondrial targeting presequence. We named this gene K1ADH3. The two genes have been localized on different chromosomes by Southern hybridization to an orthogonal-field-alternation gel electrophoresis-resolved K. lactis genome. ADH activities resolved by gel electrophoresis revealed several ADH isozymes which are differently expressed in K. lactis cells depending on the carbon source.
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  • 69
    ISSN: 0749-503X
    Keywords: Saccharomyces cerevisiae ; cell cycle regulation ; CDC28 gene ; rho- mutation ; chromosome loss ; plasmid maintenance ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: The cdc28-srm mutation in Saccheromyces cerevisiae decreases spontaneous and induced mitochondrial rhomutability and the mitotic stability of native chromosomes and recombinant circular minichromosomes. The effects of cdc28-srm on the genetic stability of cells support the hypothesis that links cell cycle regulation in yeast to changes in chromatin organization dependent on the start gene CDC28 (Hayles and Nurse, 1986).
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  • 70
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    New York, NY [u.a.] : Wiley-Blackwell
    Yeast 6 (1990) 
    ISSN: 0749-503X
    Keywords: Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
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  • 71
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    New York, NY [u.a.] : Wiley-Blackwell
    Yeast 6 (1990), S. 271-297 
    ISSN: 0749-503X
    Keywords: Yeast ; DNA-binding proteins ; transcription regulation ; RNA polymerase B ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
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  • 72
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    Yeast 6 (1990), S. 319-330 
    ISSN: 0749-503X
    Keywords: δ-Aminolevulinate dehydratase purification ; Saccharomyces cerevisiae HEM2 transformants ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Saccharomyces cerevisiae transformed with a multicopy plasmid carrying the yeast structural gene HEM2, which codes for δ-aminolevulinate dehydratase, was enriched 20-fold in the enzyme. Beginning with cell-free extracts of transformed cells, the dehydratase was purified 193-fold to near-homogeneity. This represents a 3900-fold purification relative to the enzyme activity in normal, untransformed yeast cells. The specific activity of the purified enzyme was 16·2 μmol h-1 per mg protein at pH 9·4 and 37·5°C. In most respects the yeast enzyme resembles mammalian enzymes. It is a homo-octamer with an apparent Mr, of 275 000, as determined by centrifugation in glycerol density gradients, and under denaturing conditions behaved as a single subunit of Mr ≃ 37 000. The enzyme requires reduced thiol compounds to maintain full activity, and maximum activity was obtained in the presence of 1·0 mM-Zn2+. It is sensitive to inhibition by the heavy metal ions Pb2+ and Cu2+. The enzyme exhibits Michaelis-Menten kinetics and has an apparent Km of 0·359 mM. Like dehydratases from animal tissues, the yeast enzyme is rather thermostable. During the purification process an enhancement in total δ-aminolevulinate dehydratase activity suggested the possibility that removal of an inhibitor of the enzyme could be occurring.
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  • 73
    ISSN: 0749-503X
    Keywords: Saccharomyces cerevisiae ; chromosome III ; ordered clone bank ; physical map ; transcripts ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Using λ phage vector EMBL4, we isolated 344 clones containing segments of chromosome III of Saccharomyces cerevisiae, analysed their physical structure with eight restriction enzymes and sorted the data in contiguous groups with computer programmes. Furthermore, we performed Southern hybridizations between the sorted contiguous clone groups and interrelated them into larger groups. In this way, we constructed an ordered clone bank that covers almost the whole of chromosome III with a single gap of several kilobases in length. The consensus physical map thus obtained totals 334·6 kb, which is in good agreement with the size of this chromosome estimated by pulsed-field gel electrophoresis. Southern hybridization analysis with the DNA probes containing telomere-specific sequences showed that the bank contained a telomere at a position corresponding to the right arm terminus of chromosome III. Also, five Ty elements were found to be present. To estimate the number of genes on this chromosome and to analyse their levels of expression, we performed a series of Northern hybridization experiments using total poly(A)+ RNA from vegetatively growing cells and appropriate restriction enzyme fragments from the bank. Thus, we identified a total of 156 transcripts on chromosome III, indicating, on an average, one gene in every 2 kb on this chromosome. The transcripts were visually categorized into five groups according to their apparent levels of expression. It was found that the genes located near both termini are expressed only at low levels and that highly expressed genes are rather scattered over the chromosome.
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  • 74
    ISSN: 0749-503X
    Keywords: Protein sorting ; membranes ; phospholipid synthesis ; yeast ; Saccharomces cerevisiae ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: The product of the yeast CHO1 gene, phosphatidylserine synthase (PSS), is an integral membrane protein that catalyses a central step in cellular phospholipid biosynthesis. A 1·2 kb fragment containing the regulatory and structural components of the CHO1 gene was sequenced. Transcription initiation in wild-type cells was found to occur between -1 and -15 relative to the first ATG of a large open reading frame capable of encoding a 30 804 molecular weight protein. This translation initiation site was active in vivo and in vivo in a hetrologous system. In both cases it supported production of a protein of approximately 30 000 molecular weight. A second potential translation initiation site was detected 225 or 228 bases dowstream from the first ATG. This second site was active in vitro where it supported production of a protein of 22 400 molecular weight. A subclone, lacking the 5′ regulatory region and the subsequence encoding the first 12 amino acids of the large open reading frame, allowed translation in vivo starting at the second ATG. The resulting protein was 22 000 moleculare weight, lacked the 74 N-terminal amino acids and was capable of complementing the choline auxotroy of a cho1 null-mutant. In transformants carrying this construct, PSS activity and 22 kDa protein was found to be associated with membrane fractions corresponding to mitochondria and endoplasmic reticulum. However, most of the truncated PSS protein accumulated in the cytosol in an inactive form. A hybrd-protein containing the 63 N-terminal amino acids of PSS protein accumulated in the cytosol in a active form. A hybrid-protein containing the 63 N-terminal amino acids of PSS fused to mouse dihydrofolate reductase was found exclusively in the cytosol when expressed in wild-type yeast. Thus, the hydrophilic, highly acidic N-terminus of PSS is required for efficient membrane insertion but does not appear to contain sequences required for a targeting to the membrane compartment.
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  • 75
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    Yeast 6 (1990), S. 53-60 
    ISSN: 0749-503X
    Keywords: Cyclic AMP ; Cell Cycle ; Saccharomyces cerevisiae ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Using the techniques of centrifugal elutriation it was demonstrated that during the cell division cycle of the budding yeast Saccharomyces cerevisiae there are stage-specific fluctuations in the intracellular concentration of adenosine 3′,5′-cyclic monophosphate (cAMP). Results shown here indicate that the intracellular concentration of cAMP is at its highest during the division cycle, and its lowest immediately prior to and just after cell sepraration. Results also show the extrusion of extracellular cAMP into the medium by Saccharomyces cerevisiae, extracellular cAMP levels being ten to one hundred times higher than intracellular levels. During the cell of Saccharomyces cerevisiae the extracellular level of cAMP does not fluctuate.
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  • 76
    ISSN: 0749-503X
    Keywords: Cytochrome P-450 ; Lanosterol 14α-demethylase ; alkane hydroxylase ; Immunological Analysis ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: The occurrence of cytochrome P-45014DM(lanosterol 14α-demethylase) and cytochrome P-450alk (long-chain alkane terminal hydroxylase) in various yeast strain was determined with immunological procedures. Cytochrome P-45014DM, which is a constitutive or housekeeping enzyme playing an essential role in ergosterol biogenesis, was found in all yeast strains so far tested. Cytochromes P-45014DM from different species of yeast were immunologically different, although they may have a few common antigenic sites. In contrast, cytochrome P-450alk was detected only in the alkane-assimilating yeasts.
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  • 77
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    Yeast 6 (1990), S. 87-97 
    ISSN: 0749-503X
    Keywords: Hansenula polymorpha ; methylotrophic yeast ; microbodies ; peroxisome-deficient mutants ; alcohol oxidase ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: As a first step in a genetic approach towards understanding peroxisome biogenesis and function, we have sought to isolate mutants of the methylotrophic yeast Hansenula polymorpha which are deficient in peroxisomes. A collection of 260 methanol-utilization-defective strains was isolated and screened for the ability to utilize a second compound, ethanol, the metabolism of which involves peroxisomes. Electron microscopical investigations of ultrathin sections of selected pleiotropic mutants revealed two strains which were completely devoid of peroxisomes. In both, different peroxisomal matrix enzymes were active but located in the cytosol; these included catalase, alcohol oxidase, malate synthase and isocitrate lyase.Subsequent backcrossing experiments revealed that for all crosses involving both strains, the methanol- and ethanol utilizing-deficient phenotypes segregated independently of each other, indicating that different gene mutations were responsible for these phenotypes. The phenotype of the backcrossed peroxisome-deficient derivates was identical: defective in the ability to utilize methanol but capable of growth on other carbon sources, including ethanol.The mutations complemented and therefore were recessive mutations in different genes.
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  • 78
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    Yeast 6 (1990), S. 127-137 
    ISSN: 0749-503X
    Keywords: Saccharomyces cerevisiae ; precursor processing ; aspartyl endopeptidase ; nucleotide sequence ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Mutants of Saccharomyces cerevisiae which lack the KEX2-encoded endopeptidase are unable to process proteolytically the mating factor alpha (MFα) propheromone produced from the chromosomal MFα1 and MFα2 genes (Julius et al., 1983). Overproduction of pheromone precursor from multiple, plasmid-borne MFα genes did, however, lead to the production of active MFα peptides in the absence of the KEX2 gene product. S. cerevisiae therefore must possess an alternative processing enzyme. The cleavage site of this enzyme appeared identical to that of the KEX2-encoded endopeptidase. To identify the gene responsible for the alternative processing, we have isolated clones which allowed production of mature MFα in a kex2-disrupted strain even from the chromosomal MFα genes. The gene isolated in this way was shown also to be essential for the KEX2-independent processing of propheromone overproduced from plasmid-borne MFα1. The amino acid sequence deduced from the gene shows extensive homology to a number of aspartyl proteases including the PEP4 and BARI gene products from S. cerevisiae. In contrast to the BARI gene product, the novel aspartyl protease (YAP3 for Yeast Aspartyl Protease 3) contains a C-terminal serine/threonine-rich sequence and potential transmembrane domain similar to those found in the KEX2 gene product. The corresponding gene YAP3 was located to chromosome XII. The normal physiological role of the YAP3 gene product is not known. Strains disrupted in YAP3 are both viable and able to process the mating factor a precursor.
    Additional Material: 5 Ill.
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  • 79
    ISSN: 0749-503X
    Keywords: dsRNA ; yeast ; killer yeast ; neutral yeast ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: K2 neutral strain Saccharomyces cerevisiae USM12 was identified and characterized. This strain carried an M double-stranded RNA (dsRNA) genome encoding for resistance to K2 toxin. The M dsRNA was larger than the K2 killer yeast M dsRNA and homoduplex analysis of denatured and reannealed K2 neutral M dsRNA revealed an inverted duplication. Heteroduplex analysis showed that two thirds of the K2 M genome had homology with K2 neutral M genome. Hybridization showed that the USM12 M dsRNA has significant homology with the K2M dsRNA. Protein profiles of extracellular proteins from USMA12 and cured strain indicated that USM12 did not secrete any toxin. This is the first time that a K2 neutral yeast strain has been characterized.
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  • 80
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    Yeast 6 (1990), S. 177-177 
    ISSN: 0749-503X
    Keywords: Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
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  • 81
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    Yeast 6 (1990), S. 205-212 
    ISSN: 0749-503X
    Keywords: Cell fusion ; Saccharomyces cerevisiae ; fragile mutants ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Fragile mutants of Saccharomyces cerevisiae are defective in the structure of the cell wall and plasma membrane. The mutant cells lyse in hypotonic solutions but grow exponentially when osmotic stabilizer is induced in the medium. These mutants display a general increase in the permeability of the plasma membrane. We show here that fragile yeast cells of the same mating type can fuse without protoplast formation. The frequency of cell × cell fusion is lower than that observed for protoplast × protoplast fusion and can be significantly increased if the cells of one partner are converted to protoplasts. Microscopic observations and genetic analysis demonstrate that the hybrids obtained are fusion products. The fusion between fragile cells is explained in terms of the existence of local defects on their surface where the cell wall is thinner (or even missing), thus allowing a direct contact of cells by means of their plasma membranes.
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  • 82
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    Yeast 6 (1990), S. 221-229 
    ISSN: 0749-503X
    Keywords: Gene expression ; yeast ; vector ; gene fusion ; acid phosphatase ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: The expression of acid phosphatase (Apase) from PHO5 and MFα-PHO5 hybrid genes is regulated by inorganic phosphate and mating type locus respectively, as well as the PHO4 and MATα1 gene products respectively. When PHO5 and MFα-PHO5 hybrid genes were cloned in the BamHI site of the pBR322 sequence of the yeast shuttle vectors (YRp7 or YEp9T), in one orientation they were regulated normally but in the other orientation their expression was not regulated but expressed constitutively. The pBR322 sequences present upstream of the inserted genes are responsible for the constitutive expression. By replacing the PHO5 upstream activating sequences (UAS) element with pBR322 fragments, we have identified three pBR322 sequences, rom nucleotides 376 to 650, 2068 to 2116 and 2136 to 2247, which were able to promote expression of APase. A comparison of these three pBR322 fragments revealed 5′ ATCGCGCGAG 3′ and 5′ CGGTGATGNCGG 3′ to be the common sequences likely to act as USAs in Saccharomyces cerevisiae. By using synthetic oligonucleotides, it was found that both sequences are required for maximum expression of APase activity.
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  • 83
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    Yeast 6 (1990), S. 255-261 
    ISSN: 0749-503X
    Keywords: Saccharomyces cerevisiae ; glycolysis ; oscillation ; carbon dioxide ; mass spectrometry ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: The rate of formation of carbon dioxide in cytoplasmic yeast extracts in an open system with continuous infusion of glucose was measured by membrane inlet mass spectrometry during glycolytic oscillations. The rate of CO2 production rose in the first third of each cycle to a maximum of about 100 μmol per ml yeast extract per hour and subsequently diminished to a final level of about 50 μmol per h. Measurements of the NADH light absorption under the same conditions revealed oscillations of relaxations type. The phase of high CO2 production could be related to the phase of the high NADH level, giving evidence that the flux in glycolysis is increased during the phase of high NADH concentration. Only half of the amount of injected glucose was metabolized to CO2 during the sustained oscillation, although free glucose did not accumulate.
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  • 84
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    Yeast 6 (1990), S. 213-219 
    ISSN: 0749-503X
    Keywords: Zygosaccharomyces ; food spoilage yeasts ; DNA relatedness ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: The extent of nuclear DNA complementarity was determined for members of the genus Zygosaccharomyces. From these comparisons, nine species have been identified: Z. baillii, Z. bisporus, Z. cidri, Z. fermentati, Z. florentinus, Z. mellis, Z. microellipsoides, Z. mrakii, and Z. riuxii. Candida mogii, the proposed anamorph of Z. rouxii, showed low relatedness to all nine species. The recently described Saccharomyces astigiensis and S. albasitensis were conspecific with Z. fermentati, and S. placentae showed high relatedness with Z. rouxii. Growth tests were defined that allow recognition of all Zygosaccharomyces species.
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  • 85
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    Yeast 6 (1990), S. 245-254 
    ISSN: 0749-503X
    Keywords: malate synthase gene ; nucleotide sequence ; topogenic signal ; yeast microbody ; Hansenula polymorpha ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: We have cloned the MAS gene, encoding the microbody matrix enzyme malate synthase (EC 4.1.3.2.) from the methylotrophic yeast Hensenula polymorphia. The gene was isolated by screening of a genomic library with a mixed-sequence probe, based on the partial amino acid sequence of the purified enzyme. The nucleotide sequence of a 2·4-kilobase stretch of DNA covering the MAS gene was determined. The gene contains an open reading frame 555 amino acids, amounting to a calculated molecular mass of 63 254 for the encoded protein. Comparison of the amino acid sequence with the malate synthase sequences of Escherichia coli, Brassica napus L. and Cucumis sativus L. Clearly establishes the homology of all four proteins. Compared to the soluble enzyme from E. coli, the malate synthases from H. polymorpha and both plant species, which are located in the microbodies, have a short carboxy-terminal extension. In the plant malate synthases, the extension is probably involved in routing to the microbodies, since it contains the potential peroxisomal targeting signal, Ser-Arg/Lys-Leu, at the carboxy terminus. The H. polymorpha enzyme terminates with similar amino acids, but their sequence, Ser-Leu-Lys, does not conform to any of the known peroxisomal targeting signals.
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  • 86
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    Yeast 6 (1990), S. 263-270 
    ISSN: 0749-503X
    Keywords: GABA Transport ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Gamma-aminobutyrate (GABA) accumulation in growing cultures of Saccharomyces cerevisiae was shown to occur by means of an active transport system that is inhibited by proton ionophores, azide, fluoride and arsenate ions. Transport occurred maximally at pH 5·0 and exhibited apparent Km values of 12 μM and 0·1 mM. Accumulated GABA did not efflux upon treatment with proton ionophores and exchanged with extracellular material only very slowly. However, release was complete upon treatment with nystatin. These observations raise the possibility that a major portion of intracellular GABA is sequestered in the vacuole. The response of GABA uptake to growth on various nitrogen sources suggested that uptake may be subject to several types of regulation.
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  • 87
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    Yeast 6 (1990), S. 363-366 
    ISSN: 0749-503X
    Keywords: Saccharomyces cerevisiae ; Selectable markers ; Plasmids ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: A set of plasmids was constructed that contain the yeast selectable markers HIS3, LEU2, TRP1 or URA3 embedded in the multiple cloning site of pUC18.
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  • 88
    ISSN: 0749-503X
    Keywords: Kluveromyces lactis ; budding yeast ; mitochondrial DNA ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: The nucleotide sequence of the cytochrome oxidase subunit 2 (cox2) and val-tRNA genes and surrounding regions from Kluyveromyces lactis mitochondrial DNA is reported. Analysis of the coding regions shows that the codons CUN (Thr), CGN (Arg) and AUA (Met) are absent in this gene. A single sequence, ATATAAGTAA, identical to the baker's yeast mtRNA polymerase recognition site, was detected upstream of val-tRNA. This sequence is absent from regions between val-tRNA-cox2 and cox2-cox1. In addition a sequence AATAATATTCTT, identical to the mRNA processing site in other yeast mitochondrial genomes is present 32-43 bp downstream to the TAA stop codon for the cox-2 gene. Another short conserved sequence of 5 bp, TCTAA, is present upstream of the coding regions of cox2 genes in several yeasts, including K. lactis, but is not present upstream of other genes. Comparison of cox2 sequences from other organisms indicates that the mitochondrial DNA of K. lactis. is closely related to that of Saccharomyces cerevisiae.
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  • 89
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    Yeast 6 (1990), S. 441-450 
    ISSN: 0749-503X
    Keywords: Saccharomyces cerevisiae ; novobiocin-resistance ; SUP45 ; nucleic acid synthesis ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Yeast (Saccharomyces cerevisiae) strains sensitive to a variety drugs were used to select for novobiocin-resistant mutants that were simultaneously temperature-sensitive. The mutants remained as sensitive as the parent strains to a wide range of drugs other than novobiocin, and did not exhibit any suppression of suppressible auxotrophic markers. At the non-permissive temperature, the mutant cells arrested mainly as unbudded cells, and were instantly defective in DNA and RNA synthesis, but not protein synthesis. The cloned wild-type gene was identified as SUP45, which has been previously implicated in the translation process. Our results suggest that SUP45 may have a function in addition to, or different from, the one that has been assigned to it previously.
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  • 90
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    Yeast 6 (1990), S. 473-482 
    ISSN: 0749-503X
    Keywords: Leader ; phosphoglycerate kinase ; recombinant DNA ; Saccharomyces cerevisiae ; trailer ; translation ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: In order to determine the effect of nucleotide composition of the 5′-untranslated (leader) region on the translational efficiency of mRNA in yeast, we replaced a large part of the leader region of the phosphoglycerate kinase (PGK) gene by various deoxyoligonucleotides of defined sequence. All mutations left the context of the transcription initiations site and AUG start codon intact. The mutant genes were introduced into yeast cells on a multicopy vector and the ratio of the steady-state levels of PGKmRNA and protein were determined.We found the translational efficiency to be unaffected by the presence of either an 18 nucleotides (nt) long poly A or poly C tract or by sequences consisting of mixtures of A and C residues in any proportion. In contrast, a polyU tract, as well as mixtures of U and C residues, reduced translational efficiency by a factor of two to three, presumably by long-rang base -pairing between the leader and sequences elsewhere in the coding or 3′-non-coding regions of the messenger. In agreement with this hypothesis, a five-fold reduction in translational efficiency was found for an mRNA carrying a polyC tract in the leader as well as a polyG tract in the trailer, neither of which had any effect on translational efficiency by itself. Therefore, we conclude that the leader and trailer regions (including the polyA tail) of PGK mRNA are sufficiently close to base-pair when containing complementary sequences. The resulting secondary structure evidently constitutes a barrier for incoming 40S subunits on their way to the AUG start codon.The presence of an 18 nt long polyG tract in the leader completely abolished translation of the PGK mRNA in accordance with earlier observations. However, we found the leaders containing up to 40% G residues interspersed with either A or U, still allow highly efficient translation. This value is about four times as high as the average G content of leader sequence in naturally occurring yeast mRNAS.Finally, neither deletion of about 40% of the trailer sequence of PGK mRNA, not replacement of this sequence by homopolymer tracts had any effect on translational efficiency.
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  • 91
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    Yeast 6 (1990) 
    ISSN: 0749-503X
    Keywords: Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
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  • 92
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    Yeast 6 (1990), S. 31-34 
    ISSN: 0749-503X
    Keywords: Yeast ; transcription ; recombinant DNA ; in situ hybridization ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Recently a convenient method for detection of specific RNA sequences in bacteria has been developed (Ivanov and Gigova, 1986) but the original protocol was inapplicable to microorganisms with a rigid cell wall. Here we report a modification of the RNA colony hybridization for use with yeast. The modified method includes the following consecutive procedures: (a) treatment of the yeast colonies on the membrane filter with 10% SDS at 65°C for 30 min; (b) treatment of the same filter with 3 × SSC, 10% formaldehyde at 65°C for 30 min; (c) hybridization with 32 P-labelled oligonucleotide or (DNA) specific for the RNA sequence of interest. The intensity of the radioactive signals thus obtained is comparable with that of the E. colsi colonies.
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  • 93
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    Yeast 6 (1990), S. 45-51 
    ISSN: 0749-503X
    Keywords: Yeasts ; peroxisomes ; ATPase ; freeze-fracturing immunocytochemistry ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: The presence of an ATpase on yeast peroxisomal membranes was studied by immunological methods. Western blot analysis of purified peroxisomal membranes from several yeasts revealed distinct cross-reaction with specific antibodies against the F1-part or the β-subunit of the mitochondrial ATpase of Saccharomyces cerevisiae. This was not due to mitochondrial contamination as was demonstrated by analytical sucrose gradient centrifugation.Protein A-gold labelling carried out on Lowicryl-embedded methanol-grown Hansenula polymorpha using these antibodies did not result in significant staining. However, when organelles isolated from this yeast were successively incubated with antibodies and protein A-gold prior to embedding, specific labelling was observed on both the peroxisomal membrane and the membrane of damaged mitochondria but not on intact mitochondira. Specific labelling of the peroxisomal membrane was confirmed by freeze-fracture immunocytochemistry. In addition to the peroxisomal membrane, the mitochondrial membrane was also labelled in these experiments. Freeze-fracture immunocytochemistry was also successful for the localization of peroxisomal matrix proteins, e.g. alcohol oxidase and dihydroxyacetone synthase and of mitochondrial membrane proteins, e.g. cytochrome c oxidase.
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  • 94
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    Yeast 6 (1990), S. 77-86 
    ISSN: 0749-503X
    Keywords: Flocculation ; yeast ; salt inhibition ; pH value ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Yeast flocculation was inhibited by high concentrations of a number of salts. Calcium and magnesiun salts were potent inhibitors and cesium salts were least effective. Partial inhibitors by different salts additive and were completely reversible by salt removal. Inhibition by salts was time dependent; prolonged incubation increased the degree of inhibition. Salt inhibition was partly caused by the action of salts lowering the buffer pH value, and partly caused by chaotropic inhibition of proteins on the surfaces of flocculent cells. Flocculation receptors on non-flocculent cells were ubaffected by high salt concentration. At sub-inhibitory salt concentrations, there was an enhancement of flocculation in both rate and extent.
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  • 95
    ISSN: 0749-503X
    Keywords: Hansenula polymorpha ; peroxisome biogenesis ; low pH-induced membrane fusion ; alcohol oxidase ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Low pH-induced fusion between liposomes and protoplasts of the yeast Hansenula polymorpha was demonstrated using a fluorescent assay and freeze-etch techniques. By this method foreign proteins could be introduced into the cytosol of the protoplast. For instance, after fusion of glucose oxidase-containing liposomes with protoplasts, activity of this enzyme was demonstrated cytochemically in the cytosol of the resulting protoplasts. Similar results were obtained when ferritin-containing liposomes were used. Incorporation of foreign proteins into liposomes was not a prerequisite for their introduction into the cytosol of protoplasts. In experiments where ferritin was added to protoplast suspensions together with empty liposomes, this protein was also delivered to the protoplast cytosol. However, in the absence of liposomes no uptake of proteins occurred.We tested the potential of this system in our studies on peroxisome biogenesis. Protoplasts of glucose-grown H. polymorpha remained stable for prolonged periods in osmotically stabilized cultivation media. Peroxisomes in such protoplasts were capable of protein import and assembly of matrix proteins as was demonstrated by the 20-fold increase in catalase activity after incubation with methanol or ethanol. However, mature alcohol oxidase purified from H. polymorpha introduced into protoplasts of glucose-grown cells of this organism was not targeted to peroxisomes as demonstrated with (immuno)cytochemical techniques. The enzyme remained present in the cytosol while its activity gradually decreased. Therefore mature alcohol oxidase probably does not expose the right topogenic signal(s) for recognition by its target organelle.
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  • 96
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    Yeast 6 (1990), S. 173-176 
    ISSN: 0749-503X
    Keywords: Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
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  • 97
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    Yeast 6 (1990), S. 179-186 
    ISSN: 0749-503X
    Keywords: DNA replication ; replication origin ; eukaryotic chromosome ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Replication origins in Saccharomyces cerevisiae have been identified through the clonning of autonomous replication sequence (ARS) elements that allow the extrachromosonal maintenance of plasmid molecules. ARS activity requires a close matcht to an 11 bp consensus sequence and A+T-rich flanking DNA. ARS elements with a wide range of capacities for promoting plasmid maintenance have been described. We determined the ARS activity of plasmid with inserts consisting of repetitions of a 64 bp 100% A+T sequence that has sequence similarities to known ARS elements. An insert with approximately four repeats did not yield transformants, but inserts with either eight or eleven repeats did. The cooperative of ARS activity did not require a contiguous arrangement since a plasmid containing two inserts of four repeats each, separated by about 1 kb, was functional. Our results show that a charge from non-function to function can be accomplished by the cumulative action of individually inactive sequences. We conclude that the probability of replication initiation is too low with only four repeats to allow plasmid maintenance, but the overall probability is increased by further sequence iteration to provide origin activity. We suggest that chromosomes may contain streches with dispersed, weak origin elements, each undetected by the conventional ARS assay, that in sum provide origin function.
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  • 98
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    Yeast 6 (1990), S. 187-191 
    ISSN: 0749-503X
    Keywords: Halotolerance ; osmoregulation ; glycerol-sodium symport ; glycerol-potassium symport ; sodium-proton exchange ; Debaryomyces hansenii ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Several authors have shown that the halotolerant yeast Debaryomyces hansenii, when growing exponentially in glucose medium in the presence of sodium chloride, maintains osmotic balance by establishing sodium and glycerol gradients of opposite signs across the plasma membrane. Evidence is presented here that the two gradients are linked through a sodium-glycerol symport that uses the sodium gradient as a driving force for maintaining the glycerol gradient. The symporter also accepts potassium ions as co-substrate. The kinetic parameters at 25°C, pH 5·0 were the following: Vmax, decreasing from over 500 to less than 40 μmol g-1 per h over a concentration range of 0-3 M extracellular sodium chloride; Km (glycerol) 0·40-0·6 mM over the same range; Km (sodium ions) 16·0 ± 3·21μM; Km, (potassium ions) 10·4 ± 3·6μM. Furthermore, it was observed that glycerol uptake was accompanied by proton uptake when extracellular sodium chloride was present and that the protonophore carbonylcyanide-M-chlorophenylhydrazone induced collapse of the glycerol gradient, supporting earlier proposals by others that the sodium gradient is maintained by an active sodium-proton exchange mechanism.
    Additional Material: 5 Ill.
    Type of Medium: Electronic Resource
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  • 99
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Yeast 6 (1990), S. 299-310 
    ISSN: 0749-503X
    Keywords: Dekkera ; Brettanomyces ; Eeniella ; DNA ; enzymes ; systematics ; yeasts ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: The taxonomic status of various species of Dekkera, Brettanomyces and Eeniella was examined by electrophoretic comparison of enzymes, by deoxyribonucleic acid homology and by physiological characterization. These studies demonstrated that two teleomorphic Dekkera species, D. anomala and D. bruxellensis (Synonym. D. intermedia), and four anamorphic Brettanomyces species, B. anomalus (synonym B. claussenii), B. bruxellensis (synonym B. abstinens, B. custersii, B. Intermedius, B. lambicus), B. custersianus and B. naardenesis, can be recognized. The anamorphic genus Eeniella remained as a separate, monotypic taxon.
    Additional Material: 1 Ill.
    Type of Medium: Electronic Resource
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  • 100
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Yeast 6 (1990), S. 345-352 
    ISSN: 0749-503X
    Keywords: RNA processing ; divergent transcripts ; temperature-sensitive mutants ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: ORF2 is an essential gene immediately upstream of PRP4 (formeryl RNA4), a gene involved in nuclear mRNA processing in Saccharomyces cerevisiae. The two genes are arranged head-to-head. An 8 base-pair conserved sequences element is found upstream of both genes, as well as upstream of certain other genes that are known to be involved in pre-mRNA processing. Through deletion analysis we have found that both of the conserved sequence elements are important for transcription of both genes. We have cloned ORF2 and have isolated temperature-sensitive orf2 mutants. The phenotype of these mutants does not suggest a role for ORF2 in mRNA processing. The deduced amino acid sequence of ORF2 indicates significant similarity to DPR1, a gene encoding a protein that is involved in the carboxy-terminal processing of G-protein.
    Additional Material: 3 Ill.
    Type of Medium: Electronic Resource
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