We have introduced 1 to 2 copies of a deletion mutant (betaDeltaC) of the human retinoic acid receptor beta into mouse embryonic stem (ES) cells. The betaDeltaC-expressing cells were 10 to 100 times less sensitive to RA-induced differentiation in comparison to their parental cells. In the presence of 10-7 M RA in monolayer culture, they showed no growth arrest or differentiation, but remained pluripotent. Embryoid Bodies (EBs) derived from betaDeltaC-expressing cells differentiated into cardiomyocytes rather than neurons after treatment with 10-6 M RA, and became neurons upon exposure to 10-5 or 10-4 M RA. Remarkably, after 10 passages of continuous culture in the presence of 10-7 M RA, they still were able to form chimeras after injection into blastocysts. These data suggest that appropriate levels of normal retinoid receptors are crucial for lineage-specific differentiation of mouse ES cells in vitro. The betaDeltaC mutant protein may prove to be useful in promoting "stemness" of ES cells in culture.