A Kinase-Independent Function of c-Src Mediates p130Cas Phosphorylation at the Serine-639 Site in Pressure Overloaded Myocardium.

Autor: Palanisamy AP; Division of Cardiology, Department of Medicine, Gazes Cardiac Research Institute, Medical University of South Carolina, Charleston, South Carolina, 29425-2221., Suryakumar G; Division of Cardiology, Department of Medicine, Gazes Cardiac Research Institute, Medical University of South Carolina, Charleston, South Carolina, 29425-2221., Panneerselvam K; Division of Cardiology, Department of Medicine, Gazes Cardiac Research Institute, Medical University of South Carolina, Charleston, South Carolina, 29425-2221., Willey CD; Division of Cardiology, Department of Medicine, Gazes Cardiac Research Institute, Medical University of South Carolina, Charleston, South Carolina, 29425-2221., Kuppuswamy D; Division of Cardiology, Department of Medicine, Gazes Cardiac Research Institute, Medical University of South Carolina, Charleston, South Carolina, 29425-2221.
Jazyk: angličtina
Zdroj: Journal of cellular biochemistry [J Cell Biochem] 2015 Dec; Vol. 116 (12), pp. 2793-803.
DOI: 10.1002/jcb.25224
Abstrakt: Early work in pressure overloaded (PO) myocardium shows that integrins mediate focal adhesion complex formation by recruiting the adaptor protein p130Cas (Cas) and nonreceptor tyrosine kinase c-Src. To explore c-Src role in Cas-associated changes during PO, we used a feline right ventricular in vivo PO model and a three-dimensional (3D) collagen-embedded adult cardiomyocyte in vitro model that utilizes a Gly-Arg-Gly-Asp-Ser (RGD) peptide for integrin stimulation. Cas showed slow electrophoretic mobility (band-shifting), recruitment to the cytoskeleton, and tyrosine phosphorylation at 165, 249, and 410 sites in both 48 h PO myocardium and 1 h RGD-stimulated cardiomyocytes. Adenoviral mediated expression of kinase inactive (negative) c-Src mutant with intact scaffold domains (KN-Src) in cardiomyocytes did not block the RGD stimulated changes in Cas. Furthermore, expression of KN-Src or kinase active c-Src mutant with intact scaffold function (A-Src) in two-dimensionally (2D) cultured cardiomyocytes was sufficient to cause Cas band-shifting, although tyrosine phosphorylation required A-Src. These data indicate that c-Src's adaptor function, but not its kinase function, is required for a serine/threonine specific phosphorylation(s) responsible for Cas band-shifting. To explore this possibility, Chinese hamster ovary cells that stably express Cas were infected with either β-gal or KN-Src adenoviruses and used for Cas immunoprecipitation combined with mass spectrometry analysis. In the KN-Src expressing cells, Cas showed phosphorylation at the serine-639 (human numbering) site. A polyclonal antibody raised against phospho-serine-639 detected Cas phosphorylation in 24-48 h PO myocardium. Our studies indicate that c-Src's adaptor function mediates serine-639 phosphorylation of Cas during integrin activation in PO myocardium.
(© 2015 Wiley Periodicals, Inc.)
Databáze: MEDLINE
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