BAF45D Cooperates with SMAD Signaling and Promotes Early Neural Differentiation in P19 Cells

We sought to explore whether BAF45D regulates PAX6 expression through cooperating with TGF-beta/SMAD signaling in RA treated P19 cells. Here we identified BAF45D is required for expression of phosphorylated SMAD3 and PAX6 induced by RA. Genome-wide analysis revealed that during RA-induced early neural differentiation, BAF45D knockdown failed to activate TGF-beta/SMAD signaling and induce expression of Stat3 and Smad7, two negative regulators of TGF-beta/SMAD signaling. Moreover, BAF45D was immunoprecipited with BRG1and phosphorylated SMAD3. In addition, Smad3 siRNA abolished RA-indueced expression of phosphorylated SMAD3, PAX6, STAT3 and SMAD7. Finally, overexpression of BAF45D directly induced expression of PAX6 and phosphorylated SMAD3.These results suggest that a novel effect of BAF45D cooperating with TGF-beta/SMAD signaling pathway on PAX6 level control, which may shed new light on neural differentiation of P19 cells. P19 cells from the NC siRNA and BAF45D siRNA groups (three biological replicates per group) were subjected to RA-induced differentiation for 5 days and treated with TRIzol (Invitrogen).