Fig. 2. NFATp interacts with MEF2D and enhances MEF2D-dependent reporter gene activation. (A) NFATp and MEF2D form a complex. Jurkat T cells (4 × 10⁷/lane) were transfected with pSG-flag-NFATp or with pSG-c-myc-MEF2D, respectively. NFAT-overexpressing cell lysates were prepared and immunoprecipitated with anti-Flag antibodies and the bound endogenous MEF2D was detected by western blotting using anti-MEF2D antibodies. A similar procedure was used to detect the presence of endogenous NFAT in c-Myc-MEF2D immuno precipitate. (B) Association of endogenous MEF2D, NFAT and p300 with the Nur77 promoter. Various endogenous proteins were immuno precipitated from nuclear lysates prepared from Jurkat T cells transfected with pGLNur77(–307 to –242)-luc with specific antibodies as indicated, and the co-precipitated Nur77 promoter DNA was amplified using primers specific to the promoter region of Nur77 encoded in pGLNur77(–307 to –242)-luc. (C) NFATp enhances the activation of the MEF2D-dependent Gal4 reporter gene.