Divergent effects of α and β myosin heavy chain isoforms on the N terminus of rat cardiac troponin T

Abstract

Divergent effects of α- and β-myosin heavy chain (MHC) isoforms on contractile behavior arise mainly because of their impact on thin filament cooperativity. The N terminus of cardiac troponin T (cTnT) also modulates thin filament cooperativity. Our hypothesis is that the impact of the N terminus of cTnT on thin filament activation is modulated by a shift from α- to β-MHC isoform. We engineered two recombinant proteins by deleting residues 1-43 and 44-73 in rat cTnT (RcTnT): RcTnT1-43δ and RcTnT44-73δ, respectively. Dynamic and steady-state contractile parameters were measured at sarcomere length of 2.3 μm after reconstituting proteins into detergent-skinned muscle fibers from normal (α-MHC) and propylthiouracil-treated (β-MHC) rat hearts. α-MHC attenuated Ca2+- activated maximal tension (̃46%) in RcTnT1-43δ fibers. In contrast, β-MHC decreased tension only by 19% in RcTnT1-43δ fibers. Both α- and β-MHC did not affect tension in RcTnT44-73δ fibers. The instantaneous muscle fiber stiffness measurements corroborated the divergent impact of α- and β-MHC on tension in RcTnT1-43δ fibers. pCa50 (-log of [Ca2+]free required for half-maximal activation) decreased significantly by 0.13 pCa units in β-MHC + RcTnT1-43δ fibers but remained unaltered in β-MHC + RcTnT1-43δ fibers, demonstrating that β-MHC counteracted the attenuating effect of RcTnT1-43δ on myofilament Ca2+ sensitivity. β-MHC did not alter the sudden stretch-mediated recruitment of new cross-bridges (ER) in RcTnT1-43δ fibers, but α-MHC attenuated ER by 36% in RcTnT1-43δ fibers. The divergent impact of α- and β-MHC on how the N terminus of cTnT modulates contractile dynamics has implications for heart disease; alterations in cTnT and MHC are known to occur via changes in isoform expression or mutations. © 2013 Mamidi and Chandra.