Systemic hypertension is a major public health problem in the United States. The mechanisms by which it affects an organs, such as the heart, remain poorly understood at the molecular level. Recent experiments in the sponsor's laboratory demonstrated that rat cardiac muscle can be transfected in vivo by the direct injection of plasmid DNA, and, further, that the expression of an injected reporter gene coupled to a cellular promoter is regulated in a tissue restricted and physiologically responsive manner. These findings suggest a straightforward means with which to study the regulation of cardiac genes in physiologic and pathologic states which cannot be modeled in vitro. Hypertension induces the expression of the beta-myosin heavy chain (MHC) gene in the ventricles of rats, in which the alpha-MHC isoform predominates in the normotensive state. The goal of this research project is to identify cis-acting elements responsive to hypertension in the human beta-MHC gene and the nuclear proteins with which these sequences interact.
The specific aims are 1) to determine the expression patterns of a reporter gene whose expression is driven by human beta=MHC 5' flanking sequence following direct gene transfer into rat heart in vivo; 2) to determine whether expression of this injected gene is induced by renovascular hypertension; 3) to map hypertension responsive elements in the 5' flanking sequence of the beta-MHC gene by direct injection of constructs which hav been altered by mutagenesis; and 4) to identify and to begin to purify and characterize nuclear proteins which bind specifically with cis-acting hypertension responsive elements. These studies are likely to provide an entry point from which to dissect the signal transduction by which hypertension influences the expression of specific cardiac genes.
