Src modulates contractile vascular smooth muscle function via regulation of focal adhesions

Abstract

Src is a known regulator of focal adhesion turnover in migrating cells; but, in contrast, Src is generally assumed to play little role in differentiated, contractile vascular smooth muscle (dVSM). The goal of the present study was to determine if Src-family kinases regulate focal adhesion proteins and how this might affect contractility of non-proliferative vascular smooth muscle. We demonstrate here, through the use of phosphotyrosine screening, deconvolution microscopy imaging, and differential centrifugation, that the activity of Src family kinases in aorta is regulated by the alpha agonist and vasoconstrictor phenylephrine, and leads to focal adhesion protein phosphorylation and remodeling in dVSM. Furthermore, Src inhibition via morpholino knockdown of Src or by the small molecule inhibitor PP2 prevents phenylephrine-induced adhesion protein phosphorylation, markedly slows the tissue’s ability to contract and decreases steady state contractile force amplitude. Significant vasoconstrictor-induced and Src-dependent phosphorylation of Cas pY-165, FAK pY-925, paxillin pY-118, and Erk1/2 were observed. However, increases in FAK 397 phosphorylation were not seen, demonstrating differences between cells in tissue versus migrating, proliferating cells. We show here that Src, in a cause-and effect manner, regulates focal adhesion protein function and consequently, modulates contractility during the action of a vasoconstrictor. These data point to the possibility that vascular focal adhesion proteins may be useful drug discovery targets for novel therapeutic approaches to cardiovascular disease.

Vasoconstrictor-induced endocytic recycling regulates focal adhesion protein localization and function in vascular smooth muscle

“…Not all FA-associated proteins exhibit the same redistribution pattern. It has previously been reported that CAS, a scaffolding protein and Src, a tyrosine kinase, are signaling molecules associated with dVSM FAs and that they redistribute to the membrane from the cytoskeletal fraction (37). We have confirmed those data and demonstrate in Fig.…”

“…In airway smooth muscle, agonist-induced vinculin (27), paxillin (53), FA kinase (FAK), talin (42), and ␣-actinin (64) redistribution occurs and is necessary for active tension development with cholinergic agonists. In dVSM, VASP shifts from the soluble fraction to the insoluble fraction in response to a phorbol ester (34), and Src and p130 Crk-associated substrate (CAS) redistribute between insoluble and soluble fractions in response to an ␣-agonist (37). Finally, norepinephrine-or endothelin-induced stimulation of small mesenteric arteries causes redistribution of paxillin from a soluble fraction to an insoluble fraction (41).…”

Differential MicroRNA Expression Profiles in Peripheral Arterial Disease

“…We combined the validated and predicted target list and performed a pathway enrichment analysis using the Kyoto Encyclopedia of Genes and Genome database. Several pathways with a clear involvement in vascular physiology were found to be significantly enriched, such as the mitogenactivated protein kinase (MAPK) signaling pathway, 34 focal adhesion, 35 insulin signaling pathway, 36 adherens junctions, 37 transforming growth factor (TGF)-β signaling pathway, 38 and the Wnt signaling pathway 39 ( Table V in …”

Exchange protein activated by cAMP (EPAC) controls migration of vascular smooth muscle cells in concentration- and timedependent manner

“…In doi: 10.7243/2055-0898-2-2 this light the GTPases Rho, Rac and cdc42 play critical roles: Rho mediates cell contractility, Rac regulates protrusions of lamellipodia and filopodia and cdc42 regulates directionality [3]. Other focal adhesion proteins such as focal adhesion kinase (FAK), Src and paxillin (PAX) also play key roles in this process [2,4]. Focal adhesion kinase is an intracellular protein tyrosine kinase (PTK) that is recruited to and activated at focal adhesion (FA) sites and that acts downstream of multiple ECM components [5].…”