| Project/Area Number |
15380232
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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 |
Applied molecular and cellular biology
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| Research Institution | RIKEN |
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Principal Investigator |
MATSUOKA Ken RIKEN, Cell Function research Team, Team Leader, 細胞機能研究チーム, チームリーダー (40222294)
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| Co-Investigator(Kenkyū-buntansha) |
TOYOOKA Kiminori RIKEN, Cell Function Research Team, Researcher, 細胞機能研究チーム, 研究員 (10360596)
UOZUMI Noboyuki Nagoya University, Bioscience and Biotechnology Center, Professor, 生物機能開発利用センター, 教授 (40223515)
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| Project Period (FY) |
2003 – 2005
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| Project Status |
Completed (Fiscal Year 2005)
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| Budget Amount *help |
¥15,800,000 (Direct Cost: ¥15,800,000)
Fiscal Year 2005: ¥4,200,000 (Direct Cost: ¥4,200,000)
Fiscal Year 2004: ¥4,600,000 (Direct Cost: ¥4,600,000)
Fiscal Year 2003: ¥7,000,000 (Direct Cost: ¥7,000,000)
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| Keywords | Membrane protein / vacuole / localzation / Golgi apparatus / signal / transporter / tobacco / cultured cell / 植物細胞 / 分泌系 / プロリン / 水酸化酵素 / 輸送 / 膜蛋白質 / オルガネラ / カリウムイオン輸送体 / GFP / 融合遺伝子 |
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| Research Abstract |
In order to utilize higher ability of the glycan synthesis in plants and to produce valuable and bioactive glycans using plant cell culture or plant cultured roots, modified glycan syntheses should be localized to the Golgi apparatus, where most of the cell wall non-cellulose glycans synthesized. Plants also have metabolic pathways for the catabolism of complex compounds using membrane-anchored oxidoreductases. The industrial use of these enzymes requires high-level production of the proteins. Thus developing procedures to synthesize such membrane proteins with high quantity is necessary. Moreover, most of the stresses that prevent the growth of plants in non-optimum environment can overcome by the modulation of various transporters. Thus developing systems to engineer localizations and stabilities of integral membrane proteins including transporters are requested. At present, however, knowledge on the localization mechanism of membrane proteins are scarcer than the one of soluble proteins.
Therefore, in this work we aimed to clarify the signals and mechanisms of the localization and degradation of membrane proteins in plant cells using cytochrome b5 and prolyl hydroxylase as major tools. In addition to this work we cloned several membrane transporter cDNAs from tobacco and analyzed their localization. As a result, the following results were obtained. 1)Prolylhydroxylase is a typeII membrane protein localizing to the Golgi apparatus. The Golgi localization or efficient export from the ER requires basic residues in the cytoplasmic tail. 2)Fusion protein of cytochrome b5 and a red fluorescent protein form stable protein aggregate in the cell. Formation of the aggregate depends on the tetrameric nature of RFP. 3)From tobacco BY-2 cells several transporter and other membrane protein cDNAs were cloned and localization of some of them are characterized.
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