Induction of β-MHC transgene in overloaded skeletal muscle is not eliminated by mutation of conserved elements

Mechanical overload leads to hypertrophy, increased type I fiber composition, and β-myosin heavy chain (β-MHC) induction in the fast-twitch plantaris muscle. To better understand the mechanism(s) involved in β-MHC induction, we have examined inducible expression of transgenes carrying the simultaneous mutation of three DNA regulatory subregions [muscle CAT (MCAT), C-rich, and [βe3] in the context of either a 5,600-base pair (bp; β5.6mut3) or 600-bp (β0.6mut3) β-MHC promoter in overloaded plantaris muscles of transgenic mice. Protein extract from mechanically overloaded plantaris muscle of mice, harboring either mutant transgene β5.6mut3 or β0.6mut3, showed an unexpected 2.8- to 4.5-fold increase in chloramphenicol acetyltransferase (CAT) specific activity relative to their respective controls. Similar results were obtained with wild-type (wt) β-MHC transgenes (β5.6wt, β0.6wt). Histochemical staining for both myofibrillar ATPase and CAT activity and CAT immunohistochemistry revealed a striking increase in type I fibers and that CAT expression was restricted to these fibers in overloaded plantaris muscle of β5.6mut3 transgenic mice. Our transgenic data suggest that β-MHC transgenes, and perhaps the endogenous β-MHC gene, are induced by mechanical overload via a mechanism(s) that does not exclusively require the MCAT, C-rich, or βe3 subregions.