Intrinsic Modifier Effect of cTnT Isoform Switching in Sarcomeric Cardiomyopathies

Abstract

An oft-noted component of sarcomeric cardiomyopathies is that the same mutation can exhibit significant phenotypic variability suggesting independent modifiers exist differentially affecting susceptible individuals. Two highly penetrant tropomyosin (Tm) overlap mutations exhibiting divergent pathogenic remodeling are found in the cardiac thin filament (CTF): the DCM-causative D230N-Tm and the HCM-causative R92L-cardiac Troponin T (cTnT). Patients expressing D230N-Tm exhibit a “bimodal” distribution of severity, whereby children exhibit a severe, often fatal DCM, yet can recover systolic function into adulthood, whereas R92L-cTnT-carrying patients do not exhibit an early severe phenotype. We hypothesized that these age-dependent and mutation-specific changes are due to modulation by a closely linked Tm binding partner, cTnT, transitioning from its fetal (cTnT1) to adult (cTnT3) isoform. Persistence of cTnT1 to 4-months in D230N-Tm transgenic (Tg) mice additively reduced %-fractional shortening beyond that of D230N-Tm alone, and strikingly, persistence of cTnT1 in R92L-cTnT Tg mice decreased hyper-contractile function back to non-Tg levels. These data suggest that cTnT1 can act as a potent modifier of in vivo cardiac function. We investigated these changes at the molecular level via differential scanning calorimetry. cTnT1 alone had little effect on the CTF exhibiting decreased thermal stability and no change in cooperativity of unfolding (CU) of the Tm-overlap. However, D230N-cTnT1 filaments exhibited an additive increase in CU beyond D230N-cTnT3. Coupled with increased CU was a cTnT1-induced additive reduction in the calcium sensitivity of sliding velocity likely due to disrupted CU in the overlap. Conversely, R92L-cTnT3 filaments exhibited a decrease in CU with no additive change in the presence of cTnT1. cTnT1 reduced thermal stability regardless of the baseline effect of the mutation. These data suggest that changes in Tm-overlap CU could represent an early site of dysregulation contributing to the phenotypic variability in patient populations.