Abstract 15540: p63RhoGEF is a Key Signaling Mediator in Vascular Smooth Muscle Ca2+ Sensitization Initiated by Activation of G alpha(q/11)
In normal and diseased vascular smooth muscle, the RhoA pathway activated by multiple agonists through G protein coupled receptors (GPCRs) plays a central role in the regulation of smooth muscle contraction controlling basal tone and peripheral resistance. A down stream substrate of active RhoA is Rho kinase which phosphorylates and inhibits the targeting subunit (MYPT1) of myosin phosphatase. This results in an increase in myosin regulatory light chain phosphorylation and force development without a further increase in intracellular Ca2+ concentration; a physiologically important process known as Ca2+ sensitization. While it is thought that specific agonists and GPCRs may couple to specific RhoA guanine nucleotide exchange factors (RhoGEFs), it is largely unexplored for smooth muscle contraction. Consequently, selective targeting of specific RhoGEFs may be of therapeutic importance. Therefore, we generated an expression profile of various RhoGEFs in smooth muscle, identified p63RhoGEF as one of RhoGEFs specific to smooth muscle and dissected its physiological role in smooth muscle contraction. p63RhoGEF had been shown to couple to G alpha(q/11) and RhoA in vitro and its selectivity to G alpha(q/11) coupled agonists as a mediator of RhoA activation had been shown in vivo. In this study, we further demonstrate that silencing of the endogenous p63RhoGEF in blood vessels inhibits G alpha(q/11) mediated endothelin-1-induced force to a greater extent than the predominantly G alpha(12/13) mediated thromboxane analogue, U46619. Introduction of the exogenous isolated Pleckstrin-homology (PH) domain of p63RhoGEF (residues 331-580) into permeabilized portal vein, or its expression in cultured cells, acted as a dominant negative and inhibited Gαq/11-coupled, phenylephrine-induced Ca2+ sensitized force and activation of RhoA. Thus, we demonstrate that p63RhoGEF selectively couples G alpha(q/11), but not G alpha(12/13), to RhoA activation in blood vessels and cultured cells and mediates the physiologically important Ca2+ sensitization of force induced with G alpha(q/11) coupled agonists. Our results suggest that signaling through p63RhoGEF provides a novel mechanism for selective regulation of blood pressure.
- © 2011 by American Heart Association, Inc.