The cytokine leukemia inhibitory factor (LIF) drives self-renewal of mouse embryonic stem (ES) cells by activating the transcription factor STAT3. In serum-free cultures, however, LIF is insufficient to block neural differentiation and maintain pluripotency. Here, we report that bone morphogenetic proteins (BMPs) act in combination with LIF to sustain self-renewal and preserve multilineage differentiation, chimera colonization, and germline transmission properties. ES cells can be propagated from single cells and derived de novo without serum or feeders using LIF plus BMP. The critical contribution of BMP is to induce expression of Id genes via the Smad pathway. Forced expression of Id liberates ES cells from BMP or serum dependence and allows self-renewal in LIF alone. Upon LIF withdrawal, Id-expressing ES cells differentiate but do not give rise to neural lineages. We conclude that blockade of lineage-specific transcription factors by Id proteins enables the self-renewal response to LIF/STAT3.

Embryonic stem (ES) cells have been available from inbred mice since 1981 but have not been validated for other rodents. Failure to establish ES cells from a range of mammals challenges the identity of cultivated stem cells and our understanding of the pluripotent state. Here we investigated derivation of ES cells from the rat. We applied molecularly defined conditions designed to shield the ground state of authentic pluripotency from inductive differentiation stimuli. Undifferentiated cell lines developed that exhibited diagnostic features of ES cells including colonization of multiple tissues in viable chimeras. Definitive ES cell status was established by transmission of the cell line genome to offspring. Derivation of germline-competent ES cells from the rat paves the way to targeted genetic manipulation in this valuable biomedical model species. Rat ES cells will also provide a refined test-bed for functional evaluation of pluripotent stem cell-derived tissue repair and regeneration.

Wnt/β-catenin signalling has a variety of roles in regulating stem cell fates. Its specific role in mouse epiblast stem cell self-renewal, however, remains poorly understood. Here we show that Wnt/β-catenin functions in both self-renewal and differentiation in mouse epiblast stem cells. Stabilization and nuclear translocation of β-catenin and its subsequent binding to T-cell factors induces differentiation. Conversely, retention of stabilized β-catenin in the cytoplasm maintains self-renewal. Cytoplasmic retention of β-catenin is effected by stabilization of Axin2, a downstream target of β-catenin, or by genetic modifications to β-catenin that prevent its nuclear translocation. We also find that human embryonic stem cell and mouse epiblast stem cell fates are regulated by β-catenin through similar mechanisms. Our results elucidate a new role for β-catenin in stem cell self-renewal that is independent of its transcriptional activity and will have broad implications in understanding the molecular regulation of stem cell fate.

combinations of known signal molecules to promote the development of OPCs from selected, ES-cell-derived, neuroepithelial cells. We show that the earliest stages of oligodendrocyte development follow an ordered sequence that is remarkably similar to that observed in vivo, suggesting that the ES-cell-derived neuroepithelial cells follow a normal developmental pathway to produce oligodendrocytes. These engineered ES cells thus provide a powerful system to study both the mechanisms that direct CNS stem cells down the oligodendrocyte pathway and those that influence subsequent oligodendrocyte differentiation. This strategy may also be useful for producing human cells for therapy and drug screening.