Endothelin-1 (ET-1) stimulates carboxy terminal Smad2 phosphorylation in vascular endothelial cells by a mechanism dependent on ET receptors and de novo protein synthesis.

Objective G protein-coupled receptor ( GPCR) agonists through their receptors can transactivate protein tyrosine kinase receptors such as epidermal growth factor receptor and serine/threonine kinase receptors most notably transforming growth factor ( TGF)-β receptor (Tβ RI). This signalling mechanism represents a major expansion in the cellular outcomes attributable to GPCR signalling. This study addressed the role and mechanisms involved in GPCR agonist, endothelin-1 ( ET-1)-mediated transactivation of the Tβ RI in bovine aortic endothelial cells ( BAECs). Method The in-vitro model used BAECs. Signalling intermediate phospho-Smad2 in the carboxy terminal was detected and quantified by Western blotting. Key finding ET-1 treatment of BAECs resulted in a time and concentration-dependent increase in pSmad2C. Peak phosphorylation was evident with 100 n m treatment of ET-1 at 4-6 h. Tβ RI antagonist, SB431542 inhibited ET-1-mediated pSmad2C. In the presence of bosentan, a mixed ET_A and ET_B receptor antagonist ET-1-mediated pSmad2C levels were inhibited. The ET-mediated pSmad2C was blocked by the protein synthesis inhibitor, cycloheximide. Conclusion In BAECs, ET-1 via the ETB receptor is involved in transactivation of the Tβ RI. The transactivation-dependent response is dependent upon de novo protein synthesis. [ABSTRACT FROM AUTHOR]