RESUMO
Functional overload (OV) of the rat plantaris muscle results in a fast to slow change in muscle phenotype with induction of the slow contractile protein genes including myosin light chain 2 slow (MLC2s). To identify potential cis-acting DNA sites regulating MLC2s following ablation, plasmid constructs were transfected in vivo into regenerating overloaded plantaris muscles. Activity of the 270bp promoter (-270MLC2s) was increased in OV muscles at 28 days. Mutation of the MEF2 site (-270MEF2) knocked out the overload-induced activity of the promoter. Mutation of the Ebox (-270Ebox) resulted in an earlier induction with OV and mutation of the CACC site (-270CACC) resulted in increased activity in the CON PLN with OV induction detected by 21 days. These results demonstrate that the -270MLC2s promoter contains the elements necessary for expression of MLC2s in regenerating OV PLN. More importantly, mutation analysis of -270MLC2s promoter demonstrates that mechanical loading induced expression shares some common molecular mechanisms with slow nerve dependent model regulation. In these two models of physiological induction of MLC2s, the CACC site acts as a repressor region (on/off switch) and the MEF2 site acts to modulate quantitative expression.
