Desensitizing Effect of N-terminal Truncated cTnI In RCM Myofibrils

Abstract

Cardiac TnI (cTnI) mutations have been associated with the development of restrictive cardiomyopathy (RCM) characterized by a Ca2+ hypersensitivity and diastolic dysfunction in cardiac myofibrils. Whereas cTnI N-terminal deletion (cTnI-ND) caused by restricted proteolysis in cardiac adaptation to stress manifests a lower left ventricular end diastolic pressure and an enhanced ventricular diastolic function. By crossing the RCM cTnI R193H transgenic mice (cTnI193His) with cTnI-ND transgenic mice (cTnI-ND) that contain 100% cTnI-ND in the heart, we have obtained double TG mice containing both the cTnI R193H mutant and cTnI-ND. In this study, by using these TG mouse lines, we have investigated the desensitizing effect of cTnI-ND on the RCM cTnI mutant mice and myofibrils. Our survival data for these mice indicated that cTnI-ND greatly reduced the mortality of the RCM cTnI193His mice. Ca2+ sensitivity measured in skinned myofibrils confirmed that increased myofibril Ca2+ sensitivity was the major mechanism that resulted in impaired relaxation and diastolic dysfunction in RCM cTnI193His mice and that cTnI-ND could reverse the cellular dysfunction by desensitizing the myofibrils to Ca2+. The PKA stimulation assays showed that cTnI193His myofibrils were able to respond to PKA activation, resulting in a right-shift of pCa curve after PKA treatment. However, since the myofibrils from cTnI-ND hearts lacked Ser residues 23 and 24, they had no response to PKA stimulation, showing a similar pCa curve before or after PKA. Our data have, for the first time, demonstrated a desensitizing effect by an endogenous myofibril protein proteolysis without the intervention of β-adrenergic stimulation mediated cTnI phosphorylation. The desensitizing function in cTnI-ND hearts indicates that the removal of cTnI N-terminal extension by restricted proteolysis represents a novel mechanism to improve myofibril relaxation and cardiac diastolic function in cardiac adaptation to hemodynamic and inotropic stresses.