NANOG is a direct target of TGFβ/Activin-mediated SMAD signaling in human ESCs

Self-renewal of human embryonic stem cells (ESCs) is promoted by FGF and TGF beta/Activin signaling, and differentiation is promoted by BMP signaling, but how these signals regulate genes critical to the maintenance of pluripotency has been unclear. Using a defined medium, we show here that both TGF beta and FGF signals synergize to inhibit BMP signaling; sustain expression of pluripotency-associated genes such as NANOG, OCT4, and SOX2; and promote long-term undifferentiated proliferation of human ESCs. We also show that both TGF beta- and BMP-responsive SMADs can bind with the NANOG proximal promoter. NANOG promoter activity is enhanced by TGF beta/Activin and FGF signaling and is decreased by BMP signaling. Mutation of putative SMAD binding elements reduces NANOG promoter activity to basal levels and makes NANOG unresponsive to BMP and TGF beta signaling. These results suggest that direct binding of TGF beta/Activin-responsive SMADs to the NANOG promoter plays an essential role in sustaining human ESC self-renewal.