Ritchie, S. A., Kohlhaas, C. F., Boyd, A. R., Yalla, K. C., Walsh, K., Connell, J. M.C. and Salt, I. P. (2010) Insulin-stimulated phosphorylation of endothelial nitric oxide synthase at serine-615 contributes to nitric oxide synthesis. Biochemical Journal, 426(1), pp. 85-90. (doi: 10.1042/BJ20091580)
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Publisher's URL: http://dx.doi.org/10.1042/BJ20091580
Abstract
Insulin stimulates endothelial NO (nitric oxide) synthesis via PKB (protein kinase B)/Akt-mediated phosphorylation and activation of eNOS (endothelial NO synthase) at Ser-1177. In previous studies, we have demonstrated that stimulation of eNOS phosphorylation at Ser-1177 may be required, yet is not sufficient for insulin-stimulated NO synthesis. We therefore investigated the role of phosphorylation of eNOS at alternative sites to Ser-1177 as candidate parallel mechanisms contributing to insulin-stimulated NO synthesis. Stimulation of human aortic endothelial cells with insulin rapidly stimulated phosphorylation of both Ser-615 and Ser-1177 on eNOS, whereas phosphorylation of Ser-114, Thr-495 and Ser-633 was unaffected. Insulin-stimulated Ser-615 phosphorylation was abrogated by incubation with the PI3K (phosphoinositide 3-kinase) inhibitor wortmannin, infection with adenoviruses expressing a dominant-negative mutant PKB/Akt or pre-incubation with TNFα (tumour necrosis factor α), but was unaffected by high culture glucose concentrations. Mutation of Ser-615 to alanine reduced insulin-stimulated NO synthesis, whereas mutation of Ser-615 to aspartic acid increased NO production by NOS in which Ser-1177 had been mutated to an aspartic acid residue. We propose that the rapid PKB-mediated stimulation of phosphorylation of Ser-615 contributes to insulin-stimulated NO synthesis.
| Item Type: | Articles |
|---|---|
| Keywords: | Endothelial nitric oxide synthase (eNOS), endothelium, insulin, nitric oxide, phosphorylation, protein kinase B (PKB), signalling |
| Status: | Published |
| Refereed: | Yes |
| Glasgow Author(s) Enlighten ID: | Salt, Dr Ian and Yalla, Dr Krishna and Connell, Professor John and Boyd, Mr Alasdair and Akehurst, Dr Christine |
| Authors: | Ritchie, S. A., Kohlhaas, C. F., Boyd, A. R., Yalla, K. C., Walsh, K., Connell, J. M.C., and Salt, I. P. |
| Subjects: | Q Science > QH Natural history > QH345 Biochemistry |
| College/School: | College of Medical Veterinary and Life Sciences College of Medical Veterinary and Life Sciences > School of Cardiovascular & Metabolic Health College of Medical Veterinary and Life Sciences > School of Molecular Biosciences |
| Journal Name: | Biochemical Journal |
| Journal Abbr.: | Biochem. j. |
| Publisher: | Portland Press Ltd. |
| ISSN: | 0264-6021 |
| ISSN (Online): | 1470-8728 |
| Published Online: | 20 November 2009 |
| Copyright Holders: | Copyright © 2010 Biochemical Society |
| First Published: | First published in Biochemical Journal 426:85-90 |
| Publisher Policy: | Reproduced in accordance with the copyright policy of the publisher. |
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