Self-renewal capacity and pluripotency, which are controlled by the 
Oct3/4-centered transcriptional regulatory network, are major characteristics of 
embryonic stem (ES) cells. Nuclear hormone receptor Dax1 is one of the crucial 
factors in the network. Here, we identified an orphan nuclear receptor, Esrrb 
(estrogen-related receptor beta), as a Dax1-interacting protein. Interaction of 
Dax1 and Esrrb was mediated through LXXLL motifs of Dax1 and the activation- and 
ligand-binding domains of Esrrb. Furthermore, Esrrb enhanced the promoter 
activity of the Dax1 gene via direct binding to Esrrb-binding site 1 (ERRE1, 
where "ERRE" represents "Esrrb-responsive element") of the promoter. Expression 
of Dax1 was suppressed followed by Oct3/4 repression; however, overexpression of 
Esrrb maintained expression of Dax1 even in the absence of Oct3/4, indicating 
that Dax1 is a direct downstream target of Esrrb and that Esrrb can regulate 
Dax1 expression in an Oct3/4-independent manner. We also found that the 
transcriptional activity of Esrrb was repressed by Dax1. Furthermore, we 
revealed that Oct3/4, Dax1, and Esrrb have a competitive inhibition capacity for 
each complex. These data, together with previous findings, suggest that Dax1 
functions as a negative regulator of Esrrb and Oct3/4, and these molecules form 
a regulatory loop for controlling the pluripotency and self-renewal capacity of 
ES cells.