S-Nitrosoglutathione and Endothelial Nitric Oxide Synthase-Derived Nitric Oxide Regulate Compartmentalized Ras S-Nitrosylation and Stimulate Cell Proliferation

S-Nitrosoglutathione and Endothelial Nitric Oxide Synthase-Derived Nitric Oxide Regulate Compartmentalized Ras S-Nitrosylation and Stimulate Cell Proliferation

Author Batista, Wagner L. Autor UNIFESP Google Scholar
Ogata, Fernando T. Autor UNIFESP Google Scholar
Curcio, Marli F. Autor UNIFESP Google Scholar
Miguel, Rodrigo B. Autor UNIFESP Google Scholar
Arai, Roberto J. Google Scholar
Matsuo, Alisson L. Autor UNIFESP Google Scholar
Moraes, Miriam S. Autor UNIFESP Google Scholar
Stern, Arnold Google Scholar
Monteiro, Hugo P. Autor UNIFESP Google Scholar
Institution Universidade Federal de São Paulo (UNIFESP)
Universidade de São Paulo (USP)
NYU
Abstract Aims: S-nitrosylation of Cys118 is a redox-based mechanism for Ras activation mediated by nitric oxide (NO) at the plasma membrane. Results: Ras signaling pathway stimulation by 50 and/or 100 mu M of S-nitrosoglutathione (GSNO) causes proliferation of HeLa cells. Proliferation was not observed in HeLa cells overexpressing non-nitrosatable H-Ras(C118S). HeLa cells overexpressing H-Ras(wt) containing the spatiotemporal probe green fluorescent protein (GFP) fused to the Ras-binding domain of Raf-1 (GFP-RBD) incubated with 100 mu M GSNO stimulated a rapid and transient redistribution of GFP-RBD to the plasma membrane, followed by a delayed and sustained recruitment to the Golgi. No activation of H-Ras at the plasma membrane occurred in cells overexpressing H-Ras(C118S), contrasting with a robust and sustained activation of the GTPase at the Golgi. Inhibition of Src kinase prevented cell proliferation and activation of H-Ras by GSNO at the Golgi. Human umbilical vein endothelial cells (HUVECs) stimulated with bradykinin to generate NO were used to differentiate cell proliferation and Ras activation at the plasma membrane versus Golgi. in this model, Src kinase was not involved in cell proliferation, whereas Ras activation proceeded only at the plasma membrane, indicating that HUVEC proliferation induced by NO resulted only from stimulation of Ras. Innovation: the present work is the first to demonstrate that NO-mediated activation of Ras in different subcellular compartments regulates different downstream signaling pathways. Conclusion: S-nitrosylation of H-Ras at Cys(118) and the activation of Src kinase are spatiotemporally linked events of the S-nitrosothiol-mediated signaling pathway that occurs at the plasma membrane and at the Golgi. the nonparticipation of Src kinase and the localized production of NO by endothelial NO synthase at the plasma membrane limited NO-mediated Ras activation to the plasma membrane. Antioxid. Redox Signal. 18, 221-238.
Language English
Sponsor Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)
Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)
Grant number FAPESP: 07/59617-6
FAPESP: 09/52730-7
FAPESP: 11/14392-2
Date 2013-01-01
Published in Antioxidants & Redox Signaling. New Rochelle: Mary Ann Liebert Inc, v. 18, n. 3, p. 221-238, 2013.
ISSN 1523-0864 (Sherpa/Romeo, impact factor)
Publisher Mary Ann Liebert Inc
Extent 221-238
Origin http://dx.doi.org/10.1089/ars.2011.4455
Access rights Closed access
Type Article
Web of Science ID WOS:000312007900001
URI http://repositorio.unifesp.br/handle/11600/35828

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