| Project Area | Molecular interaction and modal shift of cellular sensors |
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| Project/Area Number |
18077009
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| Research Category |
Grant-in-Aid for Scientific Research on Priority Areas
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| Allocation Type | Single-year Grants |
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| Review Section |
Biological Sciences
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| Research Institution | National Institute for Physiological Sciences |
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Principal Investigator |
NABEKURA Junichi National Institute for Physiological Sciences, 発達生理学研究系, 教授 (50237583)
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| Co-Investigator(Kenkyū-buntansha) |
MAEJIMA Takashi 生理学研究所, 発達生理学研究系, 助教 (70399319)
WATANABE Miho 生理学研究所, 発達生理学研究系, 特任助教 (10399321)
TAKATSURU Yusuke 生理学研究所, 発達生理学研究系, 特別協力研究員 (30446265)
石橋 仁 生理学研究所, 発達生理学研究系, 准教授 (50311874)
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| Co-Investigator(Renkei-kenkyūsha) |
WATANABE Miho 生理学研究所, 発達生理学研究系, 特任助教 (10399321)
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| Project Period (FY) |
2006 – 2010
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| Project Status |
Completed (Fiscal Year 2010)
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| Budget Amount *help |
¥58,500,000 (Direct Cost: ¥58,500,000)
Fiscal Year 2010: ¥16,500,000 (Direct Cost: ¥16,500,000)
Fiscal Year 2009: ¥16,000,000 (Direct Cost: ¥16,000,000)
Fiscal Year 2008: ¥8,700,000 (Direct Cost: ¥8,700,000)
Fiscal Year 2007: ¥8,700,000 (Direct Cost: ¥8,700,000)
Fiscal Year 2006: ¥8,600,000 (Direct Cost: ¥8,600,000)
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| Keywords | KCC2 / GABA / モーダルシフト / Cl / 細胞移動 / 2光子顕微鏡 / 未熟期 / スイッチ / リン酸化 / リピッドラフト / 遺伝子改変動物 / 神経回路 / クロール / lipid raft / オリゴマー / クロールトランスポーター / 障害 / 細胞死 / 脱燐酸化 |
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| Research Abstract |
The neuronal K(+)-Cl(-) cotransporter (KCC2) is a membrane transport protein that extrudes Cl(-) from neurons and helps maintain low intracellular [Cl(-)] and hyperpolarizing GABAergic synaptic potentials. Depolarizing gamma-aminobutyric acid (GABA) responses in neonatal neurons and following various forms of neuronal injury are associated with reduced levels of KCC2 expression. Mutation to the putative tyrosine phosphorylation site within the long intracellular carboxyl terminus of KCC2(Y1087D) or application of the tyrosine kinase inhibitor genistein shifted the GABA reversal potential (E(GABA)) to more depolarized values, indicating reduced KCC2 function. This was associated with a change in the expression pattern of KCC2 from a punctate distribution to a more uniform distribution, suggesting that functional tyrosine-phosphorylated KCC2 forms clusters in restricted membrane domains. A tyrosine phosphatase inhibitor increased the proportion of KCC2 associated with lipid rafts membrane domains. These results indicate that direct tyrosine phosphorylation of KCC2 results in membrane clusters and functional transport activity. Oxidative stress resulted in a rapid dephosphorylation of KCC2 that preceded the decreases in KCC2 protein or mRNA expression. Dephosphorylation of KCC2 is correlated with a reduction of transport activity and a decrease in [Cl-]i, as well as a reduction in KCC2 surface expression. Manipulation of KCC2 tyrosine phosphorylation resulted in altered neuronal viability in response to in vitro oxidative stress. We propose that neuronal stress induces a rapid loss of tyrosine phosphorylation of KCC2 that results in translocation of the protein and functional loss of transport activity.
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