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PMID: 12446767 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't

Rho GTPase/Rho kinase negatively regulates endothelial nitric oxide synthase phosphorylation through the inhibition of protein kinase B/Akt in human endothelial cells.

Molecular and cellular biology ·Vol. 22 ·No. 24 ·2002-12-00 ·Pages 8467-77

Ming XF, Viswambharan H, Barandier C, Ruffieux J, Kaibuchi K, Rusconi S, Yang Z

Abstract

Endothelial nitric oxide synthase (eNOS) is an important regulator of cardiovascular homeostasis by production of nitric oxide (NO) from vascular endothelial cells. It can be activated by protein kinase B (PKB)/Akt via phosphorylation at Ser-1177. We are interested in the role of Rho GTPase/Rho kinase (ROCK) pathway in regulation of eNOS expression and activation. Using adenovirus-mediated gene transfer in human umbilical vein endothelial cells (HUVECs), we show here that both active RhoA and ROCK not only downregulate eNOS gene expression as reported previously but also inhibit eNOS phosphorylation at Ser-1177 and cellular NO production with concomitant suppression of PKB activation. Moreover, coexpression of a constitutive active form of PKB restores the phosphorylation but not gene expression of eNOS in the presence of active RhoA. Furthermore, we show that thrombin inhibits eNOS phosphorylation, as well as expression via Rho/ROCK pathway. Expression of the active PKB reverses eNOS phosphorylation but has no effect on downregulation of eNOS expression induced by thrombin. Taken together, these data demonstrate that Rho/ROCK pathway negatively regulates eNOS phosphorylation through inhibition of PKB, whereas it downregulates eNOS expression independent of PKB.

MeSH Terms
Adenoviridae/genetics,metabolism Androstadienes/metabolism Cells, Cultured Down-Regulation/physiology Endothelium, Vascular/cytology,metabolism Enzyme Activation Gene Expression Regulation, Enzymologic Humans Intracellular Signaling Peptides and Proteins Nitric Oxide Synthase/genetics,metabolism Nitric Oxide Synthase Type III Phosphatidylinositol 3-Kinases/metabolism Phosphoinositide-3 Kinase Inhibitors Phosphorylation Protein Serine-Threonine Kinases/genetics,metabolism Proto-Oncogene Proteins/genetics,metabolism Proto-Oncogene Proteins c-akt Thrombin/metabolism Wortmannin rho GTP-Binding Proteins/genetics,metabolism rho-Associated Kinases
Chemicals
Androstadienes Intracellular Signaling Peptides and Proteins Phosphoinositide-3 Kinase Inhibitors Proto-Oncogene Proteins NOS3 protein, human Nitric Oxide Synthase Nitric Oxide Synthase Type III AKT1 protein, human Protein Serine-Threonine Kinases Proto-Oncogene Proteins c-akt rho-Associated Kinases Thrombin rho GTP-Binding Proteins Wortmannin
Authors & Affiliations
7 authors, click to expand affiliations / ORCID
Ming Xiu-Fen
Vascular Biology, Institute of Physiology, University of Fribourg, CH-1700 Fribourg, Switzerland.
Viswambharan Hema
Barandier Christine
Ruffieux Jean
Kaibuchi Kozo
Rusconi Sandro
Yang Zhihong
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Article Info
Journal
Molecular and cellular biology
Abbr.
Mol Cell Biol
ISSN
0270-7306
DOI
10.1128/MCB.22.24.8467-8477.2002
Published
2002-12-00
Pages
8467-77
Language
English
Region
United States
NLM ID
8109087
PMCID
PMC139860
Subset
IM
External Links
PubMed source DOI: 10.1128/MCB.22.24.8467-8477.2002 PMC Full Text View journal details
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