Inhibition of glycogen synthase kinase-3 alleviates Tcf3 repression of the pluripotency network and increases embryonic stem cell resistance to differentiation

Self-renewal of rodent embryonic stem cells is enhanced by partial inhibition of glycogen synthase kinase-3 (Gsk3; refs 1,2). This effect has variously been attributed to stimulation of Wnt signalling by beta-catenin(1), stabilization of Myc protein(3) and global de-inhibition of anabolic processes(4). Here we demonstrate that beta-catenin is not necessary for embryonic stem cell identity or expansion, but its absence eliminates the self-renewal response to Gsk3 inhibition. Responsiveness is fully restored by truncated beta-catenin lacking the carboxy-terminal transactivation domain(5). However, requirement for Gsk3 inhibition is dictated by expression of T-cell factor3 (Tcf3) and mediated by direct interaction with beta-catenin. Tcf3 localizes to many pluripotency genes(6) in embryonic stem cells. Our findings confirm that Tcf3 acts as a transcriptional repressor and reveal that beta-catenin directly abrogates Tcf3 function. We conclude that Gsk3 inhibition stabilizes the embryonic stem cell state primarily by reducing repressive influence on the core pluripotency network.