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101.
102.
Several cDNA clones encoding two different ADP-glucose pyrophosphorylase (AGPase, EC 2.7.7.27) polypeptides denoted VfAGPC and VfAGPP were isolated from a cotyledonary library of Vicia faba L. Both sequences are closely related to AGPase small-subunit sequences from other plants. Whereas mRNA levels of VfAGPP were equally high in developing cotyledons and leaves, the mRNA of VfAGPC was present in considerable amounts only in cotyledons. During development of cotyledons, both mRNAs accumulated until the beginning of the desiccation phase and disappeared afterwards. The increase of AGPase activity in cotyledons during the phase of storage-product synthesis was closely followed by the accumulation of starch. The AGPase activity in crude extracts of cotyledons was insensitive to 3-phosphoglycerate whereas the activity from leaves could be activated more than five-fold. Inorganic phosphate inhibited the enzyme from both tissues but was slightly more effective on the leaf enzyme. There was a correlation at the cellular level between the distribution of VfAGPP and VfAGPC mRNAs and the accumulation of starch, as studied by in-situ hybridisation and by histochemical staining in parallel tissue sections of developing seeds, respectively. During the early phase of seed development (12–15 days after fertilization) VfAGPase mRNA and accumulation of starch were detected transiently in the hypodermal, chlorenchymal and outer parenchymal cell layers of the seed coat but not in the embryo. At 25 days after fertilization both synthesis of VfAGPase mRNA and biosynthesis of starch had started in parenchyma cells of the inner adaxial zone of the cotyledons. During later stages, the expression of VfAGPase and synthesis of starch extended over most of the cotyledons but were absent from peripheral cells of the abaxial zone, provascular and procalyptral cells.Abbreviations AGPase
ADP-glucose pyrophosphorylase
- DAF
days after fertilization
- Glc1P
glucose-1-phosphate
- 3-PGA
3-phosphoglycerate
- VfAGPC
AGPase subunit of Vicia faba mainly expressed in cotyledons
- VfAGPP
AGPase subunit of Vicia faba mainly expressed in leaves and cotyledons
- pVfAGPC, pVfAGPP
plasmids containing VfAGPC and VfAGPP, respectively
This work was supported by the Bundesministerium für Forschung und Technologie BCT 0389, Molekular- und Zellbiologie von höheren Pflanzen und Pilzen. U.W acknowledges additional support by the Fonds der chemischen Industrie. We thank Elsa Fessel for excellent technical assistance. 相似文献
103.
The gibberellin (GA)-biosynthesis mutations, lh
i
, ls and Ie
5839
have been used to investigate the role(s) of the GAs in seed development of the garden pea (Pisum sativum L.). Seeds homozygous for lh
i
possess reduced GA levels, are more likely to abort during development, and weigh less at harvest, compared with wild-type seeds due to expression of the lh
i
mutation in the embryo and/ or endosperm. Compared with wild-type seeds, the lh
i
mutation reduces endogenous GA1 and gibberellic acid (GA3) levels in the embryo/endosperm a few days after anthesis and fertilizing lh
i
plants with wild-type pollen dramatically increases GA1 and GA3 levels in the embryo/ endosperm and restores normal seed development. By contrast, the ls and le
5839
mutations do not appear to reduce GA levels in the embryo/endosperm of seeds a few days after anthesis, and do not affect embryo or endosperm development. However, both the ls and lh
i
mutations substantially reduce endogenous GA levels in embryos at contact point (the first day the liquid endosperm disappears). Levels of GAs in seeds from crosses involving the ls and lh
i
mutations suggest that GAs are synthesised in both the embryo/endosperm and testa and that the expression of ls depends on the tissue and developmental stage examined. These results suggest that GAs (possibly GA1 and/or GA3) play an important role early in pea seed development by regulating the development of the embryo and/or endosperm. By contrast, the high GA levels found in wild-type seeds at contact point (and beyond) do not appear to have a physiological role in seed development.Abbreviations GAn
gibberellin An
- DAA
days after anthesis
- WT
wild-type
We thank Noel Davies, Katherine McPherson and Peter Bobbi for technical assistance, Professor L. Mander (ANU, Canberra) for dideuterated GA standards, and the Australian Research Council and Frontier Research Program, The Institute of Physical and Chemical Research (RIKEN, Japan), for financial support. 相似文献
104.
105.
Angel García-Gutiérrez Francisco R. Cantón Fernando Gallardo Francisca Sánchez-Jiménez Francisco M. Cánovas 《Plant molecular biology》1995,27(1):115-128
Pine seedlings are able to accumulate chlorophylls and develop green plastids in a light-independent manner. In this work, we have characterized ferredoxin-dependent glutamate synthase (EC 1.4.7.1; Fd-GOGAT), a key enzyme in nitrogen interconversion during this process. Fd-GOGAT has been purified about 170-fold from cotyledons of maritime pine (Pinus pinaster). As occurs in angiosperms, the native enzyme is a single polypeptide with an apparent molecular mass of 163–168 kDa that is confined to the chloroplast stroma. Polyclonal antibodies generated against the purified enzyme were used to immunoscreen a gt11 expression library from Scots pine (Pinus sylvestris) seedlings and partial cDNA clones were isolated and characterized. The clone with the longest cDNA insert (pGOP44) contained the codification for the C-terminal (550 amino acids) of the pine Fd-GOGAT polypeptide. Immunological cross-reactivity and comparative amino sequence analysis revealed that Fd-GOGAT is a well conserved protein in higher plants. Western blot analyses showed that protein was expressed in chloroplast-containing pine tissues and this expression pattern was not affected by exogenously supplied nitrogen. Fd-GOGAT mRNA, polypeptide and enzyme activity accumulated in substantial amounts in dark-grown pine seedlings. The presence of a functional Fd-GOGAT may be important to provide the required glutamate for the biosynthesis of nitrogen compounds during chloroplast biogenesis in the dark. 相似文献
106.
M. A. C. Demeulemeester W. Rademacher A. Van de Mierop M. P. De Proft 《Plant Growth Regulation》1995,17(1):47-52
Root explants of chicory (Cichorium intybus L.) were cultured in vitro under continuous light or darkness. On a standard medium (no plant growth regulators added), flowering-stems were initiated under continuous light while under continuous dark, vegetative-stems were formed. Different types of GA (gibberellin) biosynthesis inhibitors were added to the culture medium. Paclobutrazol and compounds belonging to the group of cyclohexanetriones clearly reduced flowering-stem growth under light conditions and vegetative-stem growth under dark conditions. Under light conditions, flower bud initiation was not affected. These and other results suggest that GA1 may be synthesized during the in vitro culture period and that it controls flowering-stem growth but not floral initiation.Abbreviations CCC
chlormequat chloride
- GA
gibberellin
- LAB 198 999
3,5-dioxo-4-butyryl-cyclohexane carboxylic acid ethyl ester
- BAS 111..W
1-phenoxy-3-(1H-1,2,4-triazol-1-yl)-4-hydroxy-5,5-dimethylhexane 相似文献
107.
Determination of substrate utilization patterns of soil microbial communities: An approach to assess population changes after hydrocarbon pollution 总被引:13,自引:0,他引:13
Abstract: Changes in hydrocarbon content in soils resulted in characteristic shifts of the substrate utilization patterns as tested with the Biolog system. The altered patterns of substrate utilization corresponded to similar changes in abundance of hydrocarbon-utilizing bacteria and the occurrence of specific bacterial groups in the soils. Substrate utilization patterns as recorded with the Biolog system are suitable for rapidly assessing dynamics of autochthonous soil communities and evaluating their biodegradative potential. 相似文献
108.
109.
Alterations in Phosphatidylcholine Metabolism of Stretch-Injured Cultured Rat Astrocytes 总被引:1,自引:1,他引:0
Robert G. Lamb Courtney C. Harper Jerry S. McKinney Beverly A. Rzigalinski Earl F. Ellis 《Journal of neurochemistry》1997,68(5):1904-1910
Abstract: The primary objective of this study was to determine the influence of stretch-induced cell injury on the metabolism of cellular phosphatidylcholine (PC). Neonatal rat astrocytes were grown to confluency in Silastic-bottomed tissue culture wells in medium that was usually supplemented with 10 µM unlabeled arachidonate. Cell injury was produced by stretching (5–10 mm) the Silastic membrane with a 50-ms pulse of compressed air. Stretch-induced cell injury increased the incorporation of [3H]choline into PC in an incubation time- and stretch magnitude-dependent manner. PC biosynthesis was increased three- to fourfold between 1.5 and 4.5 h after injury and returned to control levels by 24 h postinjury. Stretch-induced cell injury also increased the activity of several enzymes involved in the hydrolysis [phospholipase A2 (EC 3.1.1.4) and C (PLC; EC 3.1.4.3)] and biosynthesis [phosphocholine cytidylyltransferase (PCT; EC 2.7.7.15)] of PC. Stretch-induced increases in PC biosynthesis and PCT activity correlated well (r = 0.983) and were significantly reduced by pretrating (1 h) the cells with an iron chelator (deferoxamine) or scavengers of reactive oxygen species such as superoxide dismutase and catalase. The stretch-dependent increase in PC biosynthesis was also reduced by antioxidants (vitamin E, vitamin E succinate, vitamin E phosphate, melatonin, and n-acetylcysteine). Arachidonate-enriched cells were more susceptible to stretch-induced injury because lactate dehydrogenase release and PC biosynthesis were significantly less in non-arachidonate-enriched cells. In summary, the data suggest that stretch-induced cell injury is (a) a result of an increase in the cellular level of hydroxyl radicals produced by an iron-catalyzed Haber-Weiss reaction, (b) due in part to the interaction of oxyradicals with the polyunsaturated fatty acids of cellular phospholipids such as PC, and (c) reversible as long as the cell's membrane repair functions (PC hydrolysis and biosynthesis) are sufficient to repair injured membranes. These results suggest that stretch-induced cell injury in vitro may mimic in part experimental traumatic brain injury in vivo because alterations in cellular PC biosynthesis and PLC activity are similar in both models. Therefore, this in vitro model of stretch-induced injury may supplement or be a reasonable alternative to some in vivo models of brain injury for determining the mechanisms by which traumatic cell injury results in cell dysfunction. 相似文献
110.
Abstract: The subtilisin-like prohormone convertase SPC3 is likely to play a role in the biosynthesis of a variety of biologically active peptides. SPC3 undergoes a series of posttranslational processing events during its biosynthesis. Multiple forms have been identified that show varying degrees of truncation at the carboxyl terminus. In this study we show that the 86-kDa form of recombinant SPC3 with an intact carboxyl terminus can undergo rapid carboxyl-terminus truncation to produce a 64-kDa form. We have defined the optimal conditions for carboxyl-terminus truncation in vitro. The carboxyl-terminus truncation reaction was less calcium sensitive, active over a broader pH range, and showed differences in inhibitor sensitivity compared with the enzymatic activities of full-length and truncated forms of SPC3 toward a fluorescent peptide substrate. Increases in enzymatic activity of 86-kDa SPC3 were also measured over a time frame consistent with conversion to the 64-kDa form. However, similar specific activities for both forms of the enzyme suggest such activity increases may not be due to carboxyl-terminus truncation. The different enzymatic properties of the major molecular forms of SPC3 highlight the importance of understanding the molecular events regulating carboxyl-terminal processing of this endoprotease. 相似文献