Functional Properties of Slow Skeletal Troponin T Isoforms in Cardiac Muscle Regulation

Abstract

Since human slow skeletal troponin T (HSSTnT) mRNA expression has been shown to be upregulated in end-stage heart failure, it is important to understand its possible role in cardiac muscle regulation. At least three SSTnT isoforms have been found to exist in slow skeletal muscle: HSSTnT1 (+ Exons 5 and 12), HSSTnT2 (+5, −12), HSSTnT3 (−5, −12) and HSSTnT4 (−5, +12, only found at mRNA level). Porcine papillary skinned fibers displaced with HSSTnT1, 2 or 3 and reconstituted with HCTnI-TnC complex showed a decrease in the Ca2+ sensitivity of force development compared to adult human cardiac troponin T (HCTnT3). The extent of TnT displacement was analyzed by measuring the unregulated tension at pCa 8.0 after TnT treatment. The maximal recovered force was increased in fibers displaced with HSSTnT1 and 3. In contrast, HSSTnT4, showed an increase in the Ca2+ sensitivity of force development and no change in maximal force recovered compared to HCTnT3. Using a reconstituted system, actomyosin ATPase activity containing different HSSTnT isoforms was determined. None of the HSSTnTs were found to alter the ATPase activation in the presence of Ca2+. In contrast, HSSTnT2 and 3 showed an increase in the ATPase inhibition at 0.8 μM [Tn] in the absence of Ca2+. All of the HSSTnTs were less soluble than HCTnT3 at three different pHs, with the HSSTnT4 having the lowest solubility. Circular dichroism experiments performed in the presence of 1M KCl, revealed structural changes between HSSTnTs and HCTnT3. Proteolytic digestion assays are being carried out to determine the susceptibility of HSSTnTs to proteolysis. These results suggest that HSSTnT isoforms may play distinct functional roles in muscle regulation and the molecular mechanism may lie in their physical-chemical properties. Supported by NIH HL-042325 and AR-050199 and AHA 0825368E.