Downregulation of C-terminal Src kinase (csk) and Csk-binding protein (cbp) is associated with increased aldosterone-induced C-src phosphorylation in Shr vascular smooth muscle cells

Callera, G. E., Montezano, A. C., Bruder-Nascimento, T. and Touyz, R. M. (2013) Downregulation of C-terminal Src kinase (csk) and Csk-binding protein (cbp) is associated with increased aldosterone-induced C-src phosphorylation in Shr vascular smooth muscle cells. Hypertension, 62(3 Sup), A350.

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Publisher's URL: http://hyper.ahajournals.org/cgi/content/meeting_abstract/62/3_MeetingAbstracts/A350

Abstract

c-Src phosphorylation is controlled by the recruitment of enzyme regulators, such as C-terminal Src kinase (CSK) which inhibits Src activity, and interactions with transmembrane adaptors. These complex regulatory mechanisms coordinate activity of c-Src at multiple levels. We previously showed that in aldosterone-stimulated SHR vascular smooth muscle cells (VSMCs), c-Src phosphorylation and its downstream signaling are upregulated. Here we hypothesized that mechanisms underlying vascular c-Src hyperactivation in SHR are related to dysregulated CSK and altered autophosphorylation at Tyr416 and Tyr527 in aldosterone-stimulated SHR cells. Studies were performed in cultured VSMCs from WKY and SHR. C-terminal Src kinase (Csk) cytosol/membrane translocation, Csk-binding protein (CBP), and c-Src phosphorylation were evaluated by western blot. Cholesterol-enriched fractions were obtained by sucrose-gradient centrifugation. Aldosterone (100 nM) induced Tyr527 c-Src phosphorylation (153.5 ± 13.6 %) which locks the kinase in an inactive conformation. This was blunted in SHR cells. Csk is a cytosolic kinase that catalyzes c-Src Tyr527 phosphorylation. ASN (10 uM), a Csk inhibitor, prevented the kinase translocation to the membrane and inhibited Tyr527 c-Src phosphorylation induced by aldosterone in WKY cells. Inhibition of Csk induced an increase in Tyr416 c-Src (180 ± 21%) under basal conditions. In SHR cells, Csk translocation to the membrane was reduced by aldosterone compared with WKY cells. Aldosterone induced an increase in expression (180.3 ± 39 %) and phosphorylation (169 ± 23 %) of the adaptor protein CBP in WKY, but not in SHR cells. Aldosterone stimulation increased Csk trafficking into lipid rafts/caveolae in WKY cells, without affecting the kinase content in the cholesterol-enriched fractions from SHR. Our findings demonstrate that 1) key regulators of c-Src activation by aldosterone, specifically Csk and CBP, are altered in SHR VSMCs; 2) c-Src regulation by aldosterone involves lipid rafts/caveolae. These novel findings suggest that modulation of Csk could be an important strategy to blunt c-Src-dependent aldosterone vascular effects.

Item Type:Articles
Additional Information:American-Heart-Association High Blood Pressure Research Scientific Sessions, New Orleans, LA, USA, 11-14 Sept 2013.
Status:Published
Refereed:Yes
Glasgow Author(s) Enlighten ID:Montezano, Dr Augusto and Touyz, Professor Rhian
Authors: Callera, G. E., Montezano, A. C., Bruder-Nascimento, T., and Touyz, R. M.
College/School:College of Medical Veterinary and Life Sciences > Institute of Cardiovascular and Medical Sciences
Journal Name:Hypertension
Publisher:American Heart Association
ISSN:0194-911X
ISSN (Online):1524-4563

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