Multisite phosphorylation of Bcl-2 via protein kinase Cδ facilitates apoptosis of hypertrophic cardiomyocytes.

Abstract

Activated protein kinase Cδ (PKCδ) associated with cardiac hypertrophy moves from the cytoplasm to the mitochondria and subsequently triggers the apoptotic signalling pathway. The underlying mechanisms remain unknown. The aim of the present study was to investigate whether mitochondrial translocation of PKCδ phosphorylates multiple sites of Bcl-2, resulting in an imbalance between Bcl-2 and Bak or Bax, thus enhancing the susceptibility of hypertrophic cardiomyocytes to angiotensin II (AngII)-induced apoptosis. Chronic pressure overload was induced by transverse aortic constriction (TAC) in rats. The apoptotic rate increased in hypertrophied cardiomyocytes. In AngII-treated hearts (10 nmol/L, 60 min), there was an increase in the number of TERMINAL deoxyribonucleotidyl transferase-mediated dUTP-digoxigenin nick end-labelling (TUNEL)-positive cells; PKCδ inhibition with 500 nmol/L δV1-1 for 60 min prevented the AngII-induced increase in apoptosis. In the hypertrophied myocardium, PKCδ expression increased, whereas that of Bcl-2 decreased compared with the synchronous control. Treatment of hearts with 10 nmol/L AngII for 60 min activated PKCδ and induced translocation of PKCδ to the mitochondria, where activated PKCδ facilitated the phosphorylation of Bcl-2 at serine-87 and serine-70 sites. The multisite phosphorylated Bcl-2 was released from the mitochondria, and exhibited reduced affinity for Bak and Bax. The imbalance between Bcl-2 and Bak/Bax induced the release of mitochondrial cytochrome c and then activated the caspase 3 apoptotic pathway during AngII stimulation (10 nmol/L, 60 min) of hypertrophied cardiomyocytes. Inhibition of PKCδ reduced these effects of AngII. The results suggest that PKCδ can counteract the anti-apoptotic effect of Bcl-2 and may promote cardiomyocyte apoptosis through multisite phosphorylation of Bcl-2 in hypertrophied cardiomyocytes.