Fig. 6. Overexpression of p300 restores the expression of differentiation markers in Tax-expressing muscle cell. A, C2.7-Tax cells were transfected with either a pCMV-p300 expression plasmid or a GFP vector (pCMV-EGFP). After being cultured in differentiation medium for 72 h, the cells were subjected to immunofluorescence staining with a monoclonal anti-MHC antibody, as indicated in "Materials and Methods." Results are expressed as percentage of MHC-positive cells in cells transfected with pCMV-p300 relative to the number of fluorescent cells observed among cells transfected with pCMV-EGFP. B, C2.7-Tax cells transfected with the CMV-p300 vector or with the control plasmid, CMV-LacZ, were cultured in differentiation medium for 72 h. Cells were then harvested, and total cell lysates were prepared. Protein separation (100 µg of the lysate) and immunoblotting were carried out as indicated in "Materials and Methods." The same membrane was probed sequentially with a polyclonal anti-Tax, a monoclonal anti-MHC, and a monoclonal antiactin antibody, followed by the corresponding secondary HRP-coupled antibody. Antibody binding was visualized by enhanced chemiluminescence followed by autoradiography.