Interplay Between the Effects of Dilated Cardiomyopathy Mutation (R206L) and the Protein Kinase C Phosphomimic (T204E) of Rat Cardiac Troponin T Are Differently Modulated by α- and β-Myosin Heavy Chain Isoforms.

BACKGROUND: We hypothesized that the functional effects of R206L-a rat analog of the dilated cardiomyopathy (DCM) mutation R205L in human cardiac troponin T (TnT)-were differently modulated by myosin heavy chain (MHC) isoforms and T204E, a protein kinase C (PKC) phosphomimic of TnT. Our hypothesis was based on two observations: (1) α- and β-MHC differentially influence the functional effects of TnT; and (2) PKC isoforms capable of phosphorylating TnT are upregulated in failing human hearts.

METHODS AND RESULTS: We generated 4 recombinant TnT variants: wild type; R206L; T204E; and R206L+T204E. Functional effects of the TnT variants were tested in cardiac muscle fibers (minimum 14 per group) from normal (α-MHC) and propylthiouracil-treated rats (β-MHC) using steady-state and dynamic contractile measurements. Notably, in α-MHC fibers, Ca(2+)-activated maximal tension was attenuated by R206L (≈32%), T204E (≈63%), and R206L+T204E (≈64%). In β-MHC fibers, maximal tension was unaffected by R206L, but was attenuated by T204E (≈33%) and R206L+T204E (≈40%). Thus, β-MHC differentially counteracted the attenuating effects of the TnT variants on tension. However, in β-MHC fibers, R206L+T204E attenuated tension to a greater extent when compared to T204E alone. In β-MHC fibers, R206L+T204E attenuated the magnitude of the length-mediated recruitment of new cross-bridges (≈28%), suggesting that the Frank-Starling mechanism was impaired.

CONCLUSIONS: Our findings are the first (to our knowledge) to demonstrate that the functional effects of a DCM-linked TnT mutation are not only modulated by MHC isoforms, but also by the pathology-associated post-translational modifications of TnT.