Receptor-independent cardiac protein kinase Calpha activation by calpain-mediated truncation of regulatory domains.
BACKGROUND
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 PKCα proteolysis.


RESULTS
Isolated mouse hearts subjected to global ischemia/reperfusion demonstrated cleavage of PKCα. Calpain 1 overexpression was not sufficient to produce PKCα cleavage in normal hearts, but ischemia-induced myocardial PKCα 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 PKCα cleavage; infarct size was decreased and ventricular function enhanced in infarcted calpain 1 knockout hearts. To determine consequences of PKCα fragmentation on myocardial protein phosphorylation, transgenic mice were created conditionally expressing full-length PKCα or its N-terminal and C-terminal calpain 1 cleavage fragments. Two-dimensional mapping of ventricular protein extracts showed a distinct PKCα phosphorylation profile that was exaggerated and distorted in hearts expressing the PKCα C-terminal fragment. MALDI mass spectroscopy revealed hyperphosphorylation of myosin-binding protein C and phosphorylation of atypical substrates by the PKCα C-terminal fragment. Expression of parent PKCα produced a mild cardiomyopathy, whereas myocardial expression of the C-terminal PKCα fragment induced a disproportionately severe, rapidly lethal cardiomyopathy.


CONCLUSIONS
Proteolytic processing of PKCα by calcium-activated calpain activates pathological cardiac signaling through generation of an unregulated and/or mistargeted kinase. Production of the PKCα C-terminal fragment in ischemic hearts occurs via a receptor-independent mechanism.
Kang
Min-Young
MY
Washington University Center for Pharmacogenomics, St Louis, MO 63110, USA.
Zhang
Yan
Y
Matkovich
Scot J
SJ
Diwan
Abhinav
A
Chishti
Athar H
AH
Dorn
Gerald W
GW
D000818:Animals; D002118:Calcium / Q000378:metabolism; D002154:Calpain / Q000378:metabolism*; D009202:Cardiomyopathies / Q000378:metabolism* / Q000473:pathology; D002352:Carrier Proteins / Q000378:metabolism; D006801:Humans; D051379:Mice; D008822:Mice, Transgenic; D009203:Myocardial Infarction / Q000378:metabolism / Q000473:pathology; D015428:Myocardial Reperfusion Injury / Q000378:metabolism* / Q000473:pathology; D009206:Myocardium / Q000378:metabolism* / Q000473:pathology; D010766:Phosphorylation / Q000502:physiology; D051571:Protein Kinase C-alpha / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017434:Protein Structure, Tertiary; D015398:Signal Transduction / Q000502:physiology; D000099163:Myosin Binding Protein C
D002118:Calcium; D002154:Calpain; D002352:Carrier Proteins; D051571:Protein Kinase C-alpha; D017346:Protein Serine-Threonine Kinases; D000099163:Myosin Binding Protein C; C112682:Capn1 protein, mouse