Abstract 12575: Nucleoside Diphosphate Kinase C is a Novel Regulator of cAMP Levels and Gβγ Signaling in Cardiomyocytes
Introduction: Nucleoside diphosphate kinases (NDPKs) regulate Gβγ signaling and cAMP formation under basal conditions and during β-adrenoceptor stimulation and are enhanced in human heart failure. Treatment with β-blockers reverse the up-regulation of NDPK, implicating NDPKs in heart failure pathophysiology. This study addressed the specific role of the NDPK C isoform in cardiac Gs protein signaling.
Methods: NDPK C function was assessed in neonatal (NRCM) and adult (ARCM) rat cardiomyocytes and zebrafish hearts with real-time PCR, immunoblotting, biochemistry and adenoviral transfection. Heart failure was induced with chronic treatment with isoproterenol (ISO, 2.4 mg/kg/d 4 days) via minipumps.
Results: Chronic ISO increased mRNA levels of NDPK C by 9.4±1.9-fold and its protein levels by 2.1±0.11-fold (Fig A, p<0.05*). Immunoprecipitation of the G protein β subunit resulted in co-immunoprecipitation of NDPK C and the stimulatory Gαs subunit, and ISO enhanced this interaction. Upon ISO stimulation, NDPK C translocated from the cytosol to the plasma membrane within 3 hours in both NRCMs and ARCMs (Fig B). Adenoviral overexpression of NDPK C in NRCMs caused a 1.5-fold increase in basal and ISO induced cAMP synthesis (Fig C), whereas siRNA mediated knockdown of endogenous NDPK C decreased cAMP levels by ∼50% (Fig D). In vivo knockdown of the NDPK C ortholog in zebrafish resulted in reduced fractional shortening (-75%*) and a severe contractile dysfunction of zebrafish ventricle, but not of the atrium (Fig E), implicating NDPK C in cAMP formation and contractility regulation at whole heart level.
Conclusions: Our findings identify NDPK C as a novel and critical regulator of cAMP synthesis and Gs signaling in the heart. The up-regulation of NDPK C and the increased responsiveness to ISO in failing hearts as well as the severe loss of contractility in NDPK C depleted zebrafish implicate NDPK C as novel counterregulatory factor in the onset of heart failure.
- Heart failure
- Beta-adrenergic receptor agonists
- Cardiac hypertrophy
- Cell signaling
- Receptor-mediated signaling
- © 2011 by American Heart Association, Inc.