Verbatim): Heart failure is a major cause of premature death and disability in the United States. The basis of abnormal myofilament function in the failing heart is not known. Myofilament function is regulated by troponin, a complex made of three subunits (cTnI, cTnC, cTnT): cTnI inhibits actin-myosin interaction, the binding of Ca2+ to cTnC disinhibits this interaction, and cTnT binds the complex to tropomyosin and is essential for Ca2+ -dependent force development and myofibrillar ATPase activity. This proposal aims to determine the role of the cTnT isoforms in the regulation of myocardial function, and how they affect the failing heart. The function of the cTnT isoforms is not known. We have identified four TnT isoforms in the human heart, whose expression is regulated by development and affected by heart failure. In contrast, the same single isoform of cTnI and cTnC are expressed in the normal and failing adult human heart. We focus on the cTnT isoforms because cTnT isoform expression is correlated with the fall in myofibrillar ATPase activity in the failing human heart. We will test the following: Hypothesis 1(a) cTnT isoforms modulate the binding characteristics of cTnC to Ca2+ in troponin in vitro and the characteristics are further modulated by the incorporation of troponin into the thin filament. Hypothesis 1(b). In myocardium, the cTnT isoforms alter myofilament function by changing the myofilaments' sensitivity to Ca2+ and the sarcomere length-dependence of this sensitivity (and consequently, the Frank-Starling relation). Hypothesis 2. cTnT isoform expression in vivo alters ventricular function in vivo and myofilament function in vitro. Hypothesis 3. cTnT isoform expression in vivo preserves left ventricular function in heart disease. We will use reagents we have recently developed, including recombinant cTnT proteins and transgenic mice overexpressing cTnT isoforms. These will be used to examine the functional role of cTnT isoforms in troponin, the thin filament, isolated myocytes, ventricular bundles and the in vivo heart and to test the effects of cTnI isoform expression of left ventricular structure and function in mouse models of heart disease. These models include constriction of the transverse aorta and over-expression of calsequestrin. Defining the role of cTnT isoforms in the normal and failing heart is expected to provide targets for potential new and effective pharmacological interventions in heart failure.
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