Evidence is presented that RSK1 (ribosomal S6 kinase 1), a downstream target of MAPK (mitogen-activated protein kinase), directly phosphorylates nNOS (neuronal nitric oxide synthase) on Ser847 in response to mitogens. The phosphorylation thus increases greatly following EGF (epidermal growth factor) treatment of rat pituitary tumour GH3 cells and is reduced by exposure to the MEK (MAPK/extracellular-signal-regulated kinase kinase) inhibitor PD98059. Furthermore, it is significantly enhanced by expression of wild-type RSK1 and antagonized by kinase-inactive RSK1 or specific reduction of endogenous RSK1. EGF treatment of HEK-293 (human embryonic kidney) cells, expressing RSK1 and nNOS, led to inhibition of NOS enzyme activity, associated with an increase in phosphorylation of nNOS at Ser847, as is also the case in an in vitro assay. In addition, these phenomena were significantly blocked by treatment with the RSK inhibitor Ro31-8220. Cells expressing mutant nNOS (S847A) proved resistant to phosphorylation and decrease of NOS activity. Within minutes of adding EGF to transfected cells, RSK1 associated with nNOS and subsequently dissociated following more prolonged agonist stimulation. EGF-induced formation of the nNOS–RSK1 complex was significantly decreased by PD98059 treatment. Treatment with EGF further revealed phosphorylation of nNOS on Ser847 in rat hippocampal neurons and cerebellar granule cells. This EGF-induced phosphorylation was partially blocked by PD98059 and Ro31-8220. Together, these data provide substantial evidence that RSK1 associates with and phosphorylates nNOS on Ser847 following mitogen stimulation and suggest a novel role for RSK1 in the regulation of nitric oxide function in brain.
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Research Article|
December 21 2006
p90 RSK-1 associates with and inhibits neuronal nitric oxide synthase
Tao Song;
Tao Song
*Department of Cell Physiology, Kagawa University, Faculty of Medicine, Kagawa 761-0793, Japan
†Department of Anesthesiology, The First Affiliated Hospital, China Medical University, Shenyang 110001, China
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Katsuyoshi Sugimoto;
Katsuyoshi Sugimoto
*Department of Cell Physiology, Kagawa University, Faculty of Medicine, Kagawa 761-0793, Japan
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Hideshi Ihara;
Hideshi Ihara
‡Department of Biological Science, Graduate School of Science, Osaka Prefecture University, Sakai, Osaka 599-8531, Japan
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Akihiro Mizutani;
Akihiro Mizutani
§Division of Molecular Neurobiology, Department of Basic Medical Sciences, Institute of Medical Science, University of Tokyo, Tokyo 108-8639, Japan
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Naoya Hatano;
Naoya Hatano
*Department of Cell Physiology, Kagawa University, Faculty of Medicine, Kagawa 761-0793, Japan
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Kodai Kume;
Kodai Kume
*Department of Cell Physiology, Kagawa University, Faculty of Medicine, Kagawa 761-0793, Japan
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Toshie Kambe;
Toshie Kambe
¶Department of Pharmacology, Showa Pharmaceutical University, Machida, Tokyo 194-8543, Japan
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Fuminori Yamaguchi;
Fuminori Yamaguchi
*Department of Cell Physiology, Kagawa University, Faculty of Medicine, Kagawa 761-0793, Japan
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Masaaki Tokuda;
Masaaki Tokuda
*Department of Cell Physiology, Kagawa University, Faculty of Medicine, Kagawa 761-0793, Japan
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Yasuo Watanabe
Yasuo Watanabe
1
*Department of Cell Physiology, Kagawa University, Faculty of Medicine, Kagawa 761-0793, Japan
¶Department of Pharmacology, Showa Pharmaceutical University, Machida, Tokyo 194-8543, Japan
1To whom correspondence should be sent at the present address: Department of Pharmacology, Showa Pharmaceutical University, Machida, Tokyo 194-8543, Japan (email yasuwata@ac.shoyaku.ac.jp).
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Publisher: Portland Press Ltd
Received:
April 19 2006
Revision Received:
September 15 2006
Accepted:
September 19 2006
Accepted Manuscript online:
September 19 2006
Online ISSN: 1470-8728
Print ISSN: 0264-6021
The Biochemical Society, London
2007
Biochem J (2007) 401 (2): 391–398.
Article history
Received:
April 19 2006
Revision Received:
September 15 2006
Accepted:
September 19 2006
Accepted Manuscript online:
September 19 2006
Citation
Tao Song, Katsuyoshi Sugimoto, Hideshi Ihara, Akihiro Mizutani, Naoya Hatano, Kodai Kume, Toshie Kambe, Fuminori Yamaguchi, Masaaki Tokuda, Yasuo Watanabe; p90 RSK-1 associates with and inhibits neuronal nitric oxide synthase. Biochem J 15 January 2007; 401 (2): 391–398. doi: https://doi.org/10.1042/BJ20060580
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