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  • 1
    ISSN: 1399-3054
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: The effects of impaired carotenogenesis on plastid membrane organization, functionality and stability were studied in etiolated barley plants grown at 20 and 30°C. The plants were treated with norflurazon or amitrole, two herbicides affecting phytoene desaturation and lycopene cyclization, respectively. At 20°C, the amitrole-treated etioplasts, which accumulated lycopene in their inner membranes, exhibited disorganized prolamellar bodies, containing a prevalent form of non-phototransformable protochlorophyllide (Pchlide). They also showed a certain difficulty in reducing the phototransformable pigment to chlorophyllide when exposed to light, and were unable to reform the active ternary complex [protochlorophyllide–oxidoreductase (POR)–Pchlide–NADPH] when placed back in darkness. No ultrastructural alterations were found in norflurazon-treated etioplasts, with carotenogenesis inhibited at the phytoene desaturation step. In these latter organelles, Pchlide, whose forms were comparable with those of the control etioplasts, was photoreduced quickly after illumination and the ternary complex was reformed during a subsequent dark period. Thus, the impaired carotenogenesis leading to the accumulation of lycopene showed greater interference with the etioplast membrane arrangement and functionality than did the earlier interruption of the biosynthetic pathway at the phytoene level. This might be due to the different interactions of the distinct carotenoid precursors with other membrane components. However, in etioplasts of norflurazon-treated plants, a rise in growth temperature caused a partial demolition of prolamellar bodies, showing a lowered thermostability of the carotenoid-deficient membranes. This latter effect strengthens the concept that a correct and complete carotenogenesis pathway, leading to the synthesis of polar carotenoids (i.e. xanthophylls), is required for the maintenance of stable plastid membranes.
    Type of Medium: Electronic Resource
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  • 2
    ISSN: 1399-3054
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: The pigments of etiolated leaves of barley (Hordeum vulgare L.) were analysed during dark periods after flash illumination, and the results were compared with in vivo spectroscopy of the leaves. Pretreatment of the leaves with kinetin slightly stimulated and pretreatment with NaF and anaerobiosis inhibited the esterification of chlorophyllide a (Chlide) at 10–40 min after the flash, whereas the rapid esterification within 30 s after the flash remained unchanged. Irrespective of pretreatment, the amount of esterified pigment was, at any time, identical with the amount of pigment that had shifted its absorption from 684 to 672 nm (Shibata shift). Cycloheximide (CHI) had only a small inhibitory effect on esterification, but drastically inhibited the hydrogenation of geranylgeraniol to phytol, bound to Chlide. The regeneration of long-wavelength protochlorophyllide a (Pchlide650) was stimulated by kinetin and inhibited by CHI and NaF. During the rapid phase (0–30 s after the flash), the esterification was faster than the regeneration of Pchlide650, and this, in turn, was faster than the formation of photoactive Pchlide. The kinetics changed after pretreatment with 5-aminolaevulinic acid: regeneration of Pchlide650 was the fastest reaction and the Shibata shift preceded the esterification of Chlide. The results are discussed as pigment exchange reactions at NADPH:protochlorophyllide oxidoreductase (POR; EC 1.6.99.1).
    Type of Medium: Electronic Resource
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  • 3
    ISSN: 1399-3054
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: Dark-green cress seedlings were treated with 5-aminolevulinate (ALA) or the metal complexing agents 2,2′-bipyridyl and 8-hydroxyquinoline. The subsequent induction of cab-mRNA coding for the light-harvesting chlorophyll a/b protein (LHCP) by white or far-red light was investigated. Pretreatment with ALA 48 h after sowing yielded increased levels, the same pretreatment 72 or 96 h after sowing decreased levels of cab-mRNA. The strongest inhibition of light induction of cab-mRNA was found by pretreatment with 2,2′-bipyridyl. Less inhibition was obtained by pretreatment with 8-hydroxyquinoline. Steady-state levels of transcripts which are not light-regulated (actin, psbA) were only slightly decreased by pretreatment with ALA or the metal chelators, whereas no decrease was found for mRNA coding for NADPH: protochlorophyllide oxidoreductase. Run-off transcription with isolated nuclei showed that the transcription rate was reduced by pretreatment of intact plants with the metal chelator 2,2′-bipyridyl. The results are discussed in the context of current proposals for the role of transition metals. ALA, and chlorophyll precursors in light-induction of gene expression.
    Type of Medium: Electronic Resource
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  • 4
    ISSN: 1399-3054
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: A reduced rate of greening after continuous illumination was observed in dark-grown cress seedlings (Lepidium sativum L.) incubated with 5-aminolevulinate (ALA) or the complexing agents 2,2′-bipyridyl, 8-hydroxyquinoline or 1,10-phenanthroline. This effect cannot be explained merely by photodynamic damage caused by chlorophyll precursors which are accumulated in the dark under these conditions. Flash light experiments revealed that photoconversion of protochlorophyll(ide) to chlorophyllide was not influenced by chelator treatment. The next step in the chlorophyll pathway, the esterification of chlorophyllide, however, was inhibited. Simultaneously applicated ALA and complexing agents did not result in a synergistic reponse; on the contrary, ALA seemed to render cress plants less susceptible to the treatment with complexing agents upon subsequent irradiation. Ultrastructural studies demonstrated that grana formation in light was inhibited after pretreatment with ALA or complexing agents.
    Type of Medium: Electronic Resource
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  • 5
    ISSN: 1399-3054
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: Membrane fractions containing intact etioplasts, etioplast inner membranes, prolamellar bodies or prothylakoids from wheat (Triticum aestivum L. cv. Walde) were assayed for chlorophyll synthetase activity. Calculated on a protein basis, the etioplast inner membrane fraction showed a higher activity than the intact etioplasts. The activity was higher in the prolamellar body fraction than in the prothylakoid fraction. However, when the fractions were incubated in isolation medium with 50% (w/w) sucrose and 0.3 mM NADPH, chlorophyll synthetase activity could not be detected in the prolamellar body fraction, while the prothylakoid fraction maintained a high activity. The spectral shift to a shorter wavelength of the newly formed endogenous chlorophyllide was very rapid in the prothylakoid fraction but slow in the prolamellar body fraction. The relation between the spectral shift of chlorophyllide and the esterification activity in the fractions is discussed. Even exogenous short-wavelength chlorophyllide could not be esterified in well preserved prolamellar bodies. This indicates that chlorophyll synthetase is present in an inactive state in the prolamellar body structure. A large-scale method for the synthesis of geranylgeranylpyrophosphate, one of the substrates of the chlorophyll synthetase reaction, is also presented.
    Type of Medium: Electronic Resource
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  • 6
    ISSN: 1432-2048
    Keywords: Avena ; Chlorophyll biosynthesis ; Etioplasts ; Geranylgeranyldiphosphate ; Protoplasts
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The uptake of [1-3H]geranylgeranyl diphosphate (GGPP) into protoplasts and intact etioplasts and the metabolic interconversion therein was studied after a 2 min pulse of white light. The chlorophyll synthetase reaction, Chlide+GGPP→ChlGG, was taken as a natural probe for the etioplast compartment. This reaction yields labeled ChLGG and, by hydrogenation, labeled ChlP, when [1-3H]GGPP receives access to the etioplast stroma. It was found that penetration across the plastid envelope was rapid and that penetration across the plasma membrane of protoplasts, however, was slow. A cellular pool of soluble GGPP was detected. This pool was lost, in part, during preparation of the protoplasts and almost completely during preparation of the etioplasts. The membrane-bound phytol pool of etioplasts could not be replaced by exogenous [3H]GG. The endogenous GG and phytol pools of protoplasts, which were larger than those of etioplasts, could be replaced in part by exogenous [3H]GGPP. That part of this pool exists as soluble GGPP or as a direct precursor in the cytoplasm is discussed.
    Type of Medium: Electronic Resource
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  • 7
    ISSN: 1432-2048
    Keywords: Avena (phytochrome) ; Conformational change (phytochrome) ; Monoclonal antibody (phytochrome) ; Zea (phytochrome)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Monoclonal antibodies to defined locations on six regions of the phytochrome molecule (from Avena sativa L. or Zea mays L.) were each found to have a different affinity toward the farred-absorbing form of phytochrome (Pfr) and the red-absorbing form (Pr). The differences were small, but were consistently shown by antibodies which bind to the vicinity of the aminoterminus, the carboxylterminus and to sequences in between. It seems that the conformational differences between Pr and Pfr extend over the whole molecule in as far as it is represented by these regions and the antibodies binding to them.
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  • 8
    ISSN: 1432-2048
    Keywords: Avena ; Calmodulin ; Etiolated seedling ; Phytochrome ; Protein kinase ; (ATP-dependent)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract A protein-kinase activity which is co-purified with phytochrome from etiolated oat seedlings was investigated in some detail. Whereas phytochrome was always phosphorylated in solution (together with some contaminating protein bands), radioactive phosphate was not found in the phytochrome band after native gel electrophoresis and incubation of the entire gel with labeled ATP. Since protein kinases are usually autophosphorylated under these conditions, the result shows that the kinase activity does not reside in the phytochrome molecule itself. Radioactivity was exclusively detected in a band with the apparent molecular weight 450 kDa; sodium-dodecyl-sulfate gel electrophoresis revealed an apparent molecular weight of 60 kDa for the phosphorylated subunit. The N-terminal amino-acid sequence A L E S A G K Q L V P W was determined for this subunit which is a potential candidate for the protein kinase. The optimum conditions (pH, metal ion concentration) and kinetics of the phosphorylation reaction were determined. The presumed connection between proteinkinase activity and the signal chain leading from the far-red-absorbing form of phytochrome to physiological responses still awaits elucidation.
    Type of Medium: Electronic Resource
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  • 9
    ISSN: 1432-2048
    Keywords: Avena ; Photoreceptor ; Phototropism ; Plasma membrane ; Protein kinase ; Protein phosphorylation (blue-light-dependent)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Blue light induced the phosphorylation of a 116-kDa plasma-membrane-associated protein in dark-grown seedlings from Avena sativa L. The response was restricted to the phototropically sensitive tissue of the coleoptile tip. Surprisingly, this protein showed different properties in membrane preparations from plants that were grown for 3 d than in those from 5-d-old seedlings. In contrast to the younger coleoptiles, in 5-d-old seedlings phosphorylation of the 116-kDa protein depended strictly on the addition of Triton X-100 or other non-ionic detergents and was not abolished when the membranes were pretreated with trypsin. These latter membranes were also characterized by the appearance of two additional bluelight-regulated phosphoproteins of slightly lower molecular masses, exhibiting properties similar to the 116-kDa protein from 3-d-old plants. The data, together with solubilization studies, indicate that the 116-kDa protein is strongly membrane-bound only at the very beginning of seedling development and becomes more loosely associated in the course of coleoptile growth. In addition, we demonstrate that the capacity of the light-activated photoreceptor to recover photosensitivity in the dark also can occur under in-vitro conditions.
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  • 10
    Electronic Resource
    Electronic Resource
    Springer
    Planta 209 (1999), S. 364-370 
    ISSN: 1432-2048
    Keywords: Key words: Chlorophyllase ; Chlorophyll b reductase ; Hordeum ; 71-Hydroxy-chlorophyll a ; Leaf senescence
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract. During senescence of flowering plants, only breakdown products derived from chlorophyll a were detected although  b disappears, too (Matile et al., 1996, Plant Physiol 112: 1403–1409). We investigated the possibility of chlorophyll b reduction during dark-induced senescence of barley (Hordeum vulgare L.) leaves. Plastids isolated from senescing leaves were lysed and incubated with NADPH. We found 71-hydroxy-chlorophyll a, 71-hydroxy-chlorophyllide a, and, after incubation with Zn-pheophorbide b, also Zn-71-hydroxy-pheophorbide a, indicating activity of chlorophyll(ide) b reductase. The highest activity was found at day 2 of senescence when chlorophyll breakdown reached its highest rate. Chlorophyllase reached its highest activity under the same conditions only at days 4–6 of senescence. Based on the chlorophyll b reductase activity of plastids at day 2.5 of senescence (=100%), the bulk of activity (83%) was found in the thylakoids and only traces (5%) in the envelope fraction. Chlorophyll b reduction is considered to be an early and obligatory step of chlorophyll b breakdown.
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