Na+, K+ -ATPase is a ubiquitous plasma membrane-bound enzyme whose continuous function is of critical importance tot he normal function of virtually all animal cells. In the myocardium the enzyme represents the cellular receptor for digitalis glycosides and its activity is significantly stimulated by the action of thyroid hormone (T3). In the present application we explore the molecular basis for the physiological control of myocardial Na+, K+-ATPase expression by T3, and plan to accomplish the following three major goals: 1. Test the hypothesis that the induction of myocardial Na+, K+ -ATPase subunit mRNAs by T3 is mediated at the transcriptional level. a. Determine the role of enhanced transcription of cardiac Na+, K+-ATPase genes in the enhancement of alpha1, alpha2, and beta1 mRNA expression by T3. 1) If the response is transcriptional, then employing the recently described method of direct injection of expression plasmids into the contracting ventricular myocardium, identify DNA sequences in the 5'- flanking regions of the alpha1. alpha2. and beta1 genes that mediate the transcriptional response to T3 in myocardial tissue in vivo. 2) If the mechanism of the T3-induced increase of Na+, K+ -ATPase subunit mRNAs is post-transcriptional then test the hypothesis that sequences contained in the 3'-untranslated region of alpha1, alpha2, and beta1 RNA transcripts mediate the T3-induced stabilization of the mRNAs in the myocardium in vivo. 2. Test the hypothesis that Na+, K+ -ATPase gene expression is regulated by T3 in the heterotopic isografted heart and is hence independent of contractile work. a. Examine Na+, K+ -ATPase subunit mRNA protein and alpha1 and alpha2 enzyme activity in the heterotopic heart. b. Determine the effect of thyroid hormone deficiency and excess on Na+, K+ -ATPase expression in the heterotopic heart. 3. Test the hypothesis that various mRNAbeta1 species are differentially regulated by T3 in the myocardium and that they exhibit distinct translational efficiencies. a. Identify the different mRNAbeta1 species that are expressed in the myocardium of hypo-, eu-, and hyperthyroid rats, and determine the relative translational efficiency of each mRNAbeta1 species. Understanding of the regulation of Na+, K+ -ATPase expression and function is of relevance to the pathogenesis of several human diseases including congestive heart failure, hypertension, obesity, and altered thyroid state.
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