Delineation of sequences essential for specific promoter activation during pressure overloaded hypertrophy or...

Earlier studies from our laboratory have demonstrated the appearance of a high Mr (182-kDa) phosphoprotein during early stages of development of cardiac hypertrophy in the sera of animals subjected to aortic constriction. Furthermore, it has been reported that the injection of purified 182-kDa protein into normal animals led to the development of hypertrophy, and the injection of polyclonal antibodies into the aorta constricted animals completely, abolished the development of hypertrophy, and downregulated the expression of the beta-Myosin heavy chain (MHC) gene. To identify the cis-acting regulatory element(s), which controls induction of the beta-MHC gene in acute pressure-overloaded cardiac hypertrophy induced by the 182-kDa protein, the beta-MHC promoter fragments of various lengths linked to the chloramphenicol acetyl transferase (CAT) reporter were injected into the left ventricular apex of adult rats, which underwent aortic constriction/182-kDa protein injection or were sham-operated. Activation of the beta-MHC gene by the 182-kDa protein was studied by a chimeric gene constructed by fusion of the 5' regulatory regions of the beta-MHC gene to bacterial CAT, demonstrating that at least 431 bp of the beta-MHC promoter (+103 to -328) with one E-box motif, along with upstream regulatory sequences such as the TATA box, N-Fe, C-rich, and M-CAT elements are required for beta-MHC gene expression in vivo during cardiac hypertrophy induced by the 182-kDa protein.