G alpha(q)-mediated activation of GRK2 by mechanical stretch in cardiac myocytes: the role of protein kinase C.
G protein-coupled receptor kinase-2 (GRK2) is a critical regulator of beta-adrenergic receptor (beta-AR) signaling and cardiac function. We studied the effects of mechanical stretch, a potent stimulus for cardiac myocyte hypertrophy, on GRK2 activity and beta-AR signaling. To eliminate neurohormonal influences, neonatal rat ventricular myocytes were subjected to cyclical equi-biaxial 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 beta-AR desensitization. The hypertrophic response to mechanical stretch is mediated primarily through the G alpha(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 alpha(q) coupling blunted stretch-induced hypertrophy and GRK2 activation. Short hairpin RNA-mediated knockdown of PKC alpha 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 alpha in transgenic mice led to impaired beta-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 beta-AR signaling after mechanical stretch. Cross-talk between hypertrophic signaling at the level of PKC and beta-AR signaling regulated by GRK2 may be an important mechanism in the transition from compensatory ventricular hypertrophy to heart failure.
Malhotra
Ricky
R
Department of Surgery, Section of Cardiac and Thoracic Surgery, University of Chicago Medical Center, Chicago, Illinois 60637, USA.
D'Souza
Karen M
KM
Staron
Michelle L
ML
Birukov
Konstantin G
KG
Bodi
Ilona
I
Akhter
Shahab A
SA
D000818:Animals; D002478:Cells, Cultured; D004789:Enzyme Activation / Q000235:genetics; D054769:G-Protein-Coupled Receptor Kinase 2 / Q000235:genetics / Q000378:metabolism*; D043802:GTP-Binding Protein alpha Subunits, Gq-G11 / Q000235:genetics / Q000378:metabolism*; D055785:Gene Knockdown Techniques; D006352:Heart Ventricles / Q000201:enzymology; D051379:Mice; D018345:Mice, Knockout; D032383:Myocytes, Cardiac / Q000201:enzymology*; D010766:Phosphorylation / Q000235:genetics; D051571:Protein Kinase C-alpha / Q000235:genetics / Q000378:metabolism; D051381:Rats; D044140:Receptor, Angiotensin, Type 1 / Q000235:genetics / Q000378:metabolism; D011943:Receptors, Adrenergic, beta / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction; D013312:Stress, Physiological; D013997:Time Factors
D044140:Receptor, Angiotensin, Type 1; D011943:Receptors, Adrenergic, beta; C496630:Prkca protein, mouse; D051571:Protein Kinase C-alpha; C496568:GRK2 protein, mouse; C496569:Grk2 protein, rat; D054769:G-Protein-Coupled Receptor Kinase 2; D043802:GTP-Binding Protein alpha Subunits, Gq-G11