(A and B) Effect of H222P Lamin A expression on levels of pERK1/2 in transfected Cos-7 (A) and C2C12 (B) cells. Immunoblotting with pERK1/2 Ab and total ERK1/2 Ab was performed. Data are shown as mean ± SD of 11 (A) and 7 (B) samples per group (*P < 0.05). Significance of the results was determined using paired Student’s t test (for parametric data) and a Wilcoxon rank-sum test (for nonparametric data). Immunoblotting with GFP Ab is shown to demonstrate expression of proteins encoded by transfected plasmids. Immunoblotting with β-actin Ab was used as a loading control. (C and D) Effect of H222P Lamin A on nuclear translocation of pERK1/2 in transfected Cos-7 (C) and C2C12 (D) cells. Representative photomicrographs are shown for nontransfected (NT) cells, transfected cells expressing a GFP fusion of wild-type Lamin A, and transfected cells expressing a GFP fusion of Lamin A with the H222P amino acid substitution. Arrowheads show enhanced nuclear localization of pERK1/2 in cells expressing GFP H222P Lamin A. Scale bars: 10 μm. (E and F) Percentage of Cos-7 (E) and C2C12 (F) cells with pERK1/2 primarily in the nucleus. Nontransfected cells, transfected cells expressing a GFP fusion of wild-type Lamin A, and transfected cells expressing a GFP fusion of Lamin A with the H222P aa substitution were randomly counted and scored for nuclear pERK1/2 (see arrowheads in C for example). Transfected cells were determined by presence of GFP signal. Values are mean ± SD for 200 cells per group (*P < 0.05). The person counting the cells was blinded as to which protein was expressed.