In vivo regulation of beta-MHC gene in rodent heart: role of T-3 and evidence for an upstream enhancer

Cardiac beta-myosin heavy chain (beta-MHC) gene expression is mainly regula ted through transcriptional processes. Although these results are based pri marily on in vitro cell culture models, relatively little information is av ailable concerning the interaction of key regulatory factors thought to mod ulate MHC expression in the intact rodent heart. Using a direct gene transf er approach, we studied the in vivo transcriptional activity of different-l ength beta-MHC promoter fragments in normal control and in altered thyroid states. The test beta-MHC promoter was fused to a firefly luciferase report er gene, whereas the control alpha-MHC promoter was fused to the Renilla lu ciferase reporter gene and was used to account for variations in transfecti on efficiency. Absolute reporter gene activities showed that beta- and alph a-MHC genes were individually and reciprocally regulated by thyroid hormone . The beta-to-alpha ratios of reporter gene expression demonstrated an almo st threefold larger beta-MHC gene expression in the longest than in the sho rter promoter fragments in normal control animals, implying the existence o f an upstream enhancer. A mutation in the putative thyroid response element of the -408-bp beta-MHC promoter construct caused transcriptional activity to drop to null. When studied in the -3,500-bp beta-MHC promoter, construc t activity was reduced (similar to 100-fold) while thyroid hormone responsi veness was retained. These findings suggest that, even though the bulk of t he thyroid hormone responsiveness of the gene is contained within the first 215 bp of the beta-MHC promoter sequence, the exact mechanism of triiodoth yronine (T-3) action remains to be elucidated.