Identification of Oct4 and Sox2 targets in mouse embryonic stem cells

Abstract

Oct4 and Sox2 are transcription factors that are important in maintaining embryonic stem cell (ESC) pluripotency and self-renewal. To study their mechanism of action, chromatin immunoprecipitation (ChIP) was used to identify DNA binding targets of Oct4 and Sox2 in mouse ESCs. The Oct4, Sox2 and Nanog binding network was established and more than 1000 novel Oct4 and Sox2 binding sites mapping to genes important in various cellular functions, were identified. Oct4 and Sox2 were found to co-occupy a substantial number of genes at a fifteen nucleotide Sox2-Oct4 joint DNA motif. Motif search on the Oct4 and Sox2 binding datasets identified Stat3 and Smad1 motifs proximal to Oct4 and Sox2 binding sites. ChIP-on-chip and conventional ChIP-qPCR confirmed that Stat3, Smad1, Oct4 and Sox2 co-localise at the same DNA binding sites. Co-immunoprecipitation further demonstrated that Stat3 and Smad1 are Oct4 partners. Interdependence of these factors was further inferred by differentiation assays and RNA interference-mediated depletion. Stat3 was also shown to bind with Oct4 to regulate Nanog, providing a link between the three transcriptional pathways in mouse ESCs. This study identifies Oct4 and Sox2 binding targets and suggests that Oct4 and Sox2 work in collaboration with other factors to maintain the ESC state.