Integrative Physiology Receptor-Independent Cardiac Protein Kinase C Activation by Calpain-Mediated Truncation of Regulatory Domains

Rationale: Protein kinase (PK)Cs and calpain cysteine proteases are highly expressed in myocardium. Ischemia produces calcium overload that activates calpains and conventional PKCs. However, calpains can proteolytically process PKCs, and the potential in vivo consequences of this interaction are unknown. Objective: To determine the biochemical and pathophysiological consequences of calpain-mediated cardiac PKC proteolysis. Methods and Results: Isolated mouse hearts subjected to global ischemia/reperfusion demonstrated cleavage of PKC . Calpain 1 overexpression was not sufficient to produce PKC cleavage in normal hearts, but ischemia-induced myocardial PKC cleavage and myocardial injury were greatly increased by cardiac-specific expression of calpain 1. In contrast, calpain 1 gene ablation or inhibition with calpastatin prevented ischemia/reperfusion induced PKC cleavage; infarct size was decreased and ventricular function enhanced in infarcted calpain 1 knockout hearts. To determine consequences of PKC fragmentation on myocardial protein phosphorylation, transgenic mice were created conditionally expressing full-length PKC or its N-terminal and C-terminal calpain 1 cleavage fragments. Two-dimensional mapping of ventricular protein extracts showed a distinct PKC phosphorylation profile that was exaggerated and distorted in hearts expressing the PKC C-terminal fragment. MALDI mass spectroscopy revealed hyperphosphorylation of myosin-binding protein C and phosphorylation of atypical substrates by the PKC C-terminal fragment. Expression of parent PKC produced a mild cardiomyopathy, whereas myocardial expression of the C-terminal PKC fragment induced a disproportionately severe, rapidly lethal cardiomyopathy. Conclusions: Proteolytic processing of PKC by calcium-activated calpain activates pathological cardiac signaling through generation of an unregulated and/or mistargeted kinase. Production of the PKC C-terminal fragment in ischemic hearts occurs via a receptor-independent mechanism. (Circ Res. 2010;107:903-912.)