Fig. 4. GATA4 blocks Dlx5 binding to Runx2 promoter region. (A) HEK 293T cells were transfected with Flag-GATA4 and HA-Dlx5. Lysates were immunoprecipitated (IP) with anti-Flag antibody. Immunoprecipitated samples (upper panel) or whole cell lysates (WCL; lower panel) were subjected to Western blotting for detection of GATA4 (Flag) and Dlx5 (HA). (B) ChIP assay of GATA4 binding to the Runx2 promoter region. C2C12 cells were transfected with pMX-IRES-EGFP (C) or pMX-IRES-GATA4-EGFP (G4) plasmid. After 48 hours of transfection, samples were immunoprecipitated with control IgG or with anti-Dlx5 antibodies and were subjected to PCR amplification with primers specific for the Dlx5 binding sites of Runx2 P1 promoter region.