Abstract
Dilated cardiomyopathy (DCM), characterized by ventricular dilation and diminished contractile function is one of the leading causes of heart failure. Recently, mutations in cardiac Troponin T (TnT) were shown to cause DCM. The major goal of this study is to investigate the functional properties of TnT mutations causing DCM in the fetal and adult troponin (Tn).At least four different TnT isoforms are expressed in a developmentally regulated manner in the human heart. TnT3 is the dominant TnT isoform in the adult heart. In TnT3 isoform, DeltaK210 mutation caused a significant decrease in Ca2+ sensitivity of force development while the R141W mutation did not alter the Ca2+ sensitivity compared to TnT3-WT at both pH 7.0 and 6.5. In addition, both TnT3-DCM mutations decreased maximal ATPase activity in the presence of Ca2+.The dominant cardiac TnT isoform in the embryonic heart is TnT1 and the DCM mutations were created in the TnT1 isoform. In skinned fiber studies, TnT1-WT treated fibers reconstituted with cardiac TnI·TnC, significantly increased Ca2+ sensitivity of force development when compared to TnT3-WT. Fibers treated with both TnT1-DCM mutations significantly decreased Ca2+ sensitivity of force development compared to TnT1-WT. Also, both the TnT1-DCM mutations decreased maximal actomyosin ATPase activity relative to TnT1-WT. In addition to a different TnT isoform, a different TnI isoform (slow skeletal TnI) is present in the embryonic heart. In order to understand the behavior of TnT DCM mutants in fetal troponin, we used slow skeletal TnI instead of cardiac TnI in skinned fiber studies as well as in reconstituted actomyosin ATPase assays. Fiber studies showed that TnT1-WT treated fibers reconstituted with ssTnI·TnC increased Ca2+ sensitivity of force development compared to TnT3-WT treated fibers. However, both TnT1-DCM mutations decreased Ca2+ sensitivity of force development compared to TnT1-WT treated fibers in the presence of ssTnI·TnC. The maximal ATPase activity also decreased in TnT1-DCM mutations complexed with ssTnI·TnC when compared to TnT1-WT.The decrease in maximal actomyosin ATPase activity (associated with the velocity of a shortening muscle) consistently observed in the fetal and adult Tn may provide the stimulus for the sarcomere to remodel leading to the phenotype of DCM.