Gαq-mediated Activation of GRK2 by Mechanical Stretch in Cardiac Myocytes

Abstract

G protein-coupled receptor kinase-2 (GRK2) is a critical regulator of ␤-adrenergic receptor (␤-AR) signaling and cardiac function. We studied the effects of mechanical stretch, a potent stimulus for cardiac myocyte hypertrophy, on GRK2 activity and ␤-AR signaling. To eliminate neurohormonal influences, neonatal rat ventricular myocytes were subjected to cyclical equibiaxial stretch. A hypertrophic response was confirmed by "fetal" gene up-regulation. GRK2 activity in cardiac myocytes was increased 4.2-fold at 48 h of stretch versus unstretched controls. Adenylyl cyclase activity was blunted in sarcolemmal membranes after stretch, demonstrating ␤-AR desensitization. The hypertrophic response to mechanical stretch is mediated primarily through the G␣ q -coupled angiotensin II AT 1 receptor leading to activation of protein kinase C (PKC). PKC is known to phosphorylate GRK2 at the N-terminal serine 29 residue, leading to kinase activation. Overexpression of a mini-gene that inhibits receptor-G␣ q coupling blunted stretch-induced hypertrophy and GRK2 activation. Short hairpin RNA-mediated knockdown of PKC␣ also significantly attenuated stretch-induced GRK2 activation. Overexpression of a GRK2 mutant (S29A) in cardiac myocytes inhibited phosphorylation of GRK2 by PKC, abolished stretch-induced GRK2 activation, and restored adenylyl cyclase activity. Cardiac-specific activation of PKC␣ in transgenic mice led to impaired ␤-agonist-stimulated ventricular function, blunted cyclase activity, and increased GRK2 phosphorylation and activity. Phosphorylation of GRK2 by PKC appears to be the primary mechanism of increased GRK2 activity and impaired ␤-AR signaling after mechanical stretch. Cross-talk between hypertrophic signaling at the level of PKC and ␤-AR signaling regulated by GRK2 may be an important mechanism in the transition from compensatory ventricular hypertrophy to heart failure.

β-Adrenergic Receptor–Mediated Cardiac Contractility Is Inhibited via Vasopressin Type 1A-Receptor–Dependent Signaling

“…Although desensitization of one GPCR upon activation of another distinct GPCR has been shown, for instance opioid receptor-like 1 (ORL1)-mediated desensitization of the μ opioid receptor and M3 muscarinic receptor-mediated desensitization of β2AR, these effects have also been shown to be PKC-dependent, even when involving GRK signaling 38, 39 . Similarly, while mechanical stretch activates the AT1R with subsequent up-regulation of GRK2 and reduction in βAR signaling in neonatal rat ventricular myocytes, inhibition of either AT1R-Gq protein coupling or PKC-dependent phosphorylation of GRK2 restored normal βAR signaling 40 .…”

Uterine Stretch and Progesterone Action

“…Lei et al Uterine Stretch and Progesterone Action J Clin Endocrinol Metab, June 2011, 96(6):E1013-E1024 lipase C (PLC) have been shown to mediate stretch-induced changes in gene expression (36,37), but PKC and PLC activators had no effect on basal PR mRNA expression, and both PKC and PLC inhibitors failed to block the stretch-induced reduction in PR mRNA expression (Fig. 4, H and I).…”

Protein kinase C mechanisms that contribute to cardiac remodelling

“…Although studies in genetic models of PKC α gene deletion or PKC α overexpression in mice have focused on inhibitor-1 as the PKC α substrate that mediates changes in cardiac contractility, it is important to note that PKC α phosphorylates a large number of other cellular proteins that are predicted to also either directly or indirectly regulate cardiac contraction, including the α 1C subunit of the L-type calcium channel [66], various sarcomeric proteins that influence cardiomyocyte contraction and myocardial stiffness (including cardiac troponin I, cardiac troponin T, myosin binding protein C and titin [67–69]) and G protein-coupled receptor kinase 2 (GRK2); PKC α -dependent phosphorylation of GRK2 leads to enhanced enzyme activity and β -adrenergic receptor desensitization [70]. Studies to date have not formally excluded possible alternate roles for these PKC α substrates in the cardiac phenotypes that develop in these models.…”