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Protease‐activated receptor ‐3 signals independently to phosphorylate endothelial nitric oxide synthase in human endothelial cells
Author(s) -
Tillery Lakeisha C.,
Epperson Tenille,
Austin Anita,
Motley Evangeline
Publication year - 2011
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.25.1_supplement.lb491
Subject(s) - enos , phosphorylation , thrombin , signal transduction , microbiology and biotechnology , umbilical vein , nitric oxide , nitric oxide synthase , nitric oxide synthase type iii , endothelial dysfunction , receptor , endothelium , chemistry , biology , medicine , biochemistry , endocrinology , platelet , in vitro
Protease‐activated receptor‐3 (PAR‐3) is widely recognized as a non‐signaling elusive member of a G protein‐coupled receptor family known to regulate the endothelial nitric oxide synthase (eNOS)/nitric oxide (NO) signaling pathway via phosphorylation of its regulatory sites. Failure of the eNOS/NO pathway leads to endothelial dysfunction, a precursor to cellular induced hypertension and other cardiovascular diseases. Using human umbilical vein endothelial cells (HUVECs) as a model for human endothelium, we demonstrated that PAR‐3 is highly co‐expressed with PAR‐1 and ‐2 and can be activated using a synthetic peptide, SFNGGP‐NH 2 . Therefore, we hypothesize that PAR‐3 can signal independently of PAR‐1 and ‐2 to regulate eNOS. Immunoblot analysis using eNOS specific antibodies showed that 25uM of SFNGGP lead to the phosphorylation of the positive site of eNOS at ‐Ser1177 and the negative site ‐Thr495 in a time‐dependent manner similar to that observed with thrombin. After HUVECs were pretreated with 10uM of SCH79797, a selective PAR‐1 inhibitor, then stimulated with SFNGGP for 5 minutes, we observed an increase in phosphorylation at both eNOS regulatory sites. Thrombin‐induced phosphorylation of these sites was inhibited in the presence of SCH79797. Our data suggests that PAR‐3 is able to regulate eNOS/NO signaling via site specific phosphorylation that is independent of the activation of PAR‐1. Cardiovascular disease is the leading cause of death among Americans, where hypertension can be a silent aggressor. Studying the signaling pathways responsible for eNOS‐induced NO production will lead to a better understanding of endothelial dysfunction in cardiovascular diseases such as hypertension. NIH‐NIGMS‐SCORE and NIH‐NHLBI‐T32