| Project/Area Number |
11680627
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Functional biochemistry
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| Research Institution | Nagoya University |
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Principal Investigator |
WATANABE Yasuo School of Medicine, Nagoya University, Assistant Professor, 医学部, 講師 (10273228)
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| Co-Investigator(Kenkyū-buntansha) |
NAITO Yasuhito School of Medicine, Nagoya University, Research Associate, 医学部, 助手 (80303618)
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| Project Period (FY) |
1999 – 2000
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| Project Status |
Completed (Fiscal Year 2000)
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| Budget Amount *help |
¥3,100,000 (Direct Cost: ¥3,100,000)
Fiscal Year 2000: ¥1,300,000 (Direct Cost: ¥1,300,000)
Fiscal Year 1999: ¥1,800,000 (Direct Cost: ¥1,800,000)
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| Keywords | calcium signal / neuronal NO synthase / calmodulin kinase / structure and function / phosphorylation / dephosphorylation / activation mechanism / neuronal cell / カルシウム情報ネットワーク / 一酸化窒素合成酵素 / リン酸化部位 / 活性制御 / リン酸化抗体 |
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| Research Abstract |
Purpose Ca2+ signals are delivered to appropriate intracellular target proteins via phosphorylation catalyzed by Ca2+/calmodulin (CaM)-dependent protein kinases (CaM-Ks). Nitric oxide (NO) plays a crucial role in synaptic plasticity in the brain. It is formed by the enzyme NO synthase (NOS), which generates NO and the by-product L-citrulline from its substrate L-arginine. While neuronal NOS (nNOS) phosphorylation has been investigated in several studies, the effects in vivo and the physiological consequences are not completely understood. To elucidate the activation mechanism of nNOS through its phosphorylation by CaM-Ks, the following projects were undertaken : 1. Determination of the amino acid residue of nNOS, being phosphorylated by CaM-Ks in vitro. 2. Regulation of nNOS by CaM-Ks in neuronal cells.
Results and discussion 1. We reported that nNOS is phosphorylated at Ser847 by CaM-Ks, including CaM-K Iα, CaM-K IIα, and CaM-K IV in vitro. We have also demonstrated that phosphorylation of nNOS at Ser847 by CaM-Ks attenuates the catalytic activity of the enzyme in vitro. 2. We have shown that direct phosphorylation of nNOS at Ser847 by CaM-K IIα results in decrease in NOS activity in NG108-15 cells. Furthermore, protein phosphatase 2A (PP2A) could be shown to dephosphorylate the Ser847 residue with recovery of enzyme activity. Our results suggest a potential involvement of phosphorylation by CaM-K IIα/ dephosphorylation by protein phosphatase 2A in affecting the balance of NOS activity in neuronal cells.
Future prospect Based on our present studies, further analyses are need to elucidate the in vivo regulation of nNOS in neural cells via its Ser847 phosphorylation by CaM kinases.
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