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. Overall design: 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).