MEF2D depletion results in increased PI3K/Akt activation in NRVMs.
A, Western blot and densitometry analysis reveals an increase in Akt phosphorylation at the activating residues threonine 308 and serine 473 in MEF2D-deficient cardiomyocytes. B, PI3K/Akt inhibition by treatment with GDC-0941 blunts up-regulation of positive cell cycle markers at the transcript level. C, FOXO3a protein is enriched in the nuclei of shlacZ-transduced NRVMs, whereas MEF2D-depleted NRVMs display significantly increased levels of cytoplasmic FOXO3a, suggesting FOXO3a inactivation in the absence of MEF2D. White arrows point to FOXO3a enrichment. Mcm3, minichromosome maintenance complex component 3; Mcm5, minichromosome maintenance complex component 5; Mcm5, minichromosome maintenance complex component 6; Pcna, proliferating cell nuclear antigen; Ccne1, cyclin E1; Ccne2, cyclin E2. The data are means ± S.E. (error bars). *, p < 0.05; **, p < 0.01; ***, p < 0.001.