Abstract 1060: Regulation of Protease Activated Receptor-1 signaling by G-protein coupled receptor kinase 3
G protein-coupled receptors (GPCRs) play crucial roles in normal heart function and dysregulated GPCR signaling contributes to heart failure (HF). Protease-activated receptors (PARs) are one class of GPCR expressed in the heart. Emerging evidence demonstrates that excessive PAR-1 signaling induces cardiac dysfunction. An important component of PAR signaling is ERK1/2, which is phosphorylated in response to PAR stimulation and can influence myocyte hypertrophy and survival. Cytoplasmic pERK1/2 accumulation depends in part on scaffolding complexes that assemble with internalized GPCRs. Activation of an internalization-defective PAR mutant leads to enhanced nuclear pERK1/2 accumulation upon stimulation. The nuclear/cytoplasmic distribution of pERK1/2 may be a key factor in determining the cellular effects of PAR stimulation as elevated nuclear pERK in cardiomyocytes is suggested to promote survival and physiological hypertrophy. Phosphorylation of PARs by G-protein coupled receptor kinase 3 (GRK3) is thought to promote receptor internalization, potentially influencing overall pERK1/2 accumulation and subcellular distribution. We have used a dominant negative form of GRK3 lacking the kinase domain (GRK3ct) to test the hypothesis that GRK3 influences PAR1 internalization and ERK1/2 phosphorylation. By measuring cell surface receptor levels we demonstrate that GRK3ct interferes with PAR1 internalization. Immunofluorescent staining and cellular fractionation techniques further show that GRK3ct enhances nuclear accumulation of pERK1/2 in COS-7 cells and adult mouse cardiomyocytes. Furthermore we find that GRK3ct overexpression in neonatal rat cardiomyocytes increases PAR1-activation induced physiologic hypertrophy. In summary these results may explain recent unpublished reports that mice overexpressing GRK3ct in the heart are protected against ischemic injury, a heart failure model that involves pathologic PAR signaling. Thus we conclude that following ischemic injury, reducing PAR1 internalization via interfering with endogenous GRK3 activity or promoting nuclear pERK accumulation might improve cardiac recovery.