Premium
Cyclic GMP‐dependent protein kinase (PKG) regulates smooth muscle gene expression : Possible involvement of Elk‐1 sumoylation
Author(s) -
Choi ChungSik,
Sellak Hassan,
Brown Felricia M.,
Lincoln Thomas M.
Publication year - 2009
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.23.1_supplement.775.22
Subject(s) - sumo protein , myocardin , transfection , microbiology and biotechnology , immunoprecipitation , biology , gene expression , western blot , kinase , protein kinase a , regulation of gene expression , gene , ubiquitin , biochemistry , serum response factor
Although the regulation of smooth muscle cell (SMC) gene expression by cGMP‐dependent protein kinase (PKG) is now recognized, the mechanisms underling these effects are not fully understood. In this study, we report that PKG expression increased the level of a high molecular weight Elk‐1 as determined by western blot using a phospho‐Elk‐1 antibody. PKG expression also enhanced myocardin‐induced SM22 promoter activity in cultured SMC. With this background, we examined the hypothesis that PKG may modulate Elk‐1 activity through posttranslational modification by sumoylation, thus contributing to the expression of SMC specific genes. Using immunoprecipitation with sumo‐1 antibody and subsequent immunoblot with phospho‐Elk‐1 antibody, we identified the higher molecular weight species as sumoylated Elk‐1. PKG expression in SMC increased Elk‐1 sumoylation two‐fold compared to the control cells. The sumoylation of Elk‐1 was confirmed by the transfection of dominant negative form of the sumo‐conjugating enzyme, DN‐Ubc9. Furthermore, siRNA‐mediated suppression of PKG confirmed the role of the kinase in the up‐regulation of sumoylated phospho‐Elk‐1. Overexpression of Elk‐1, sumo‐1, and PKG in COS‐7 cells confirmed again that PKG increased the sumoylation of Elk‐1. Transfection of combined vectors demonstrated that PKG inhibited Elk‐1 binding and activity on the myocardin‐induced SMC specific gene promoter activity as measured by ChIP assay and reporter gene expression, respectively. Taken together, these data suggest that PKG increased sumoylation of Elk‐1, thereby increasing myocardin‐SRF activity on SMC‐specific gene expression.