The Nkx2-5 homeodomain protein plays a key role in cardiomyogenesis. Ectopic 
expression in frog and zebrafish embryos results in an enlarged myocardium; 
however, expression of Nkx2-5 in fibroblasts was not able to trigger the 
development of beating cardiac muscle. In order to examine the ability of Nkx2-5 
to modulate endogenous cardiac specific gene expression in cells undergoing 
early stages of differentiation, P19 cell lines overexpressing Nkx2-5 were 
differentiated in the absence of Me2SO. Nkx2-5 expression induced 
cardiomyogenesis in these cultures aggregated without Me2SO. During 
differentiation into cardiac muscle, Nkx2-5 expression resulted in the 
activation of myocyte enhancer factor 2C (MEF2C), but not MEF2A, -B, or -D. In 
order to compare the abilities of Nkx2-5 and MEF2C to induce cellular 
differentiation, P19 cells overexpressing MEF2C were aggregated in the absence 
of Me2SO. Similar to Nkx2-5, MEF2C expression initiated cardiomyogenesis, 
resulting in the up-regulation of Brachyury T, bone morphogenetic protein-4, 
Nkx2-5, GATA-4, cardiac alpha-actin, and myosin heavy chain expression. These 
findings indicate the presence of a positive regulatory network between Nkx2-5 
and MEF2C and show that both factors can direct early stages of cell 
differentiation into a cardiomyogenic pathway.