| Project/Area Number |
07456053
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| Research Category |
Grant-in-Aid for Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
応用微生物学・応用生物化学
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| Research Institution | Okazaki National Rsearch Institutes |
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Principal Investigator |
NISHIMURA Mikio Okazaki National Rsearch Institute, National Institute for Basic Biology, Professor, 基礎生物学研究所, 教授 (80093061)
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| Co-Investigator(Kenkyū-buntansha) |
HAYASHI Makoto Okazaki National Research Institute, National Institute for Basic Biology, Resea, 基礎生物学研究所, 助手 (50212155)
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| Project Period (FY) |
1995 – 1996
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| Project Status |
Completed (Fiscal Year 1996)
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| Budget Amount *help |
¥7,700,000 (Direct Cost: ¥7,700,000)
Fiscal Year 1996: ¥1,700,000 (Direct Cost: ¥1,700,000)
Fiscal Year 1995: ¥6,000,000 (Direct Cost: ¥6,000,000)
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| Keywords | microbody / targeting signal / transgenic plant / functional transformation / glyoxysomes / leaf-peroxisomes / alternative splicing / Alternative splicing / アスコルビン酸ペルオキシダーゼ / ヒドロキシルピルビン酸レダクターゼ |
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| Research Abstract |
1. In germinating fatty seedlings, microbodies are functionally transformed to leaf peroxisomes from glyoxysomes, and during greening to glyoxysomes from leaf peroxisomes during senesecence.Immunocytochemical studies have shown that glyoxysomes can exchange directly into leaf peroxisomes during greening and leaf peroxisomes are once again converted directly to glyoxysomes during senescence. To crarify the regulation underlying the microbody tranformation, transgenic Arabidopsis plants that expressed a fusion protein composed of the N-terminal region of and glyoxysomal citrate synthase (gCS) and beta-glucuromidase (GUS), were generated and their localization and processing were characterized by immunological and immunocytochemical methods. The fusion protein was transported into functionally different microbodies, such as glyoxysomes, leaf peroxisomes and unspecialized microbodies and was subsequently processed. These observations indicated that the transport of gCS is mediated by its a
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minoterminal presequence and that the transport system is functional in all plant microbodies. Therefore, it is unlikely that glyoxysomes and leaf peroxisomes possess different targeting machineries.
2. Hydroxpyruvate reductase belongs to leaf-peroxisome specific enzymes and is accumulated in microbodies during greening. Two different hydroxypyruvate redutase (HPR1 and HPR2) are expressed in pumpkin. cDNA and genomic DNA cloning for HPR strongly suggests that HPR1 and HPR2 are produced by alternative splicing. Two different hydroxypryuvate reductase, HPR1 and HPR2, are shown to be localized in microbodies and in the cytosol respectively, suggesting the possibility that the microbody transition may be regulated by alternative splicing.
3. Two proteins in glyoxysomal membranes with molecular masses of 31kDa and 28kDa were purified and characterized. The former was shown to be a novel ascorbate peroxidase. It was found that the amounts of these membrance proteins decreased during the microbody transition from glyoxysomes to leaf peroxisomes and that the large one was retained in leaf peroxisomes, whereas the small one could not be found in leaf peroxisomes after completion of the microbody transition. The results clearly showed that membrane proteins in glyoxysomes change dramatically during the microbody transition, as do the enzymes in the matrix. Less
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