Project description:The Wnt/β-catenin signalling pathway is a key regulator of embryonic stem cell self-renewal and differentiation. Constitutive activation of this pathway has been shown to significantly increase mouse embryonic stem cell (mESC) self-renewal and pluripotency marker expression. In this study, we generated a novel β-catenin knock-out model in mESCs by using CRISPR/Cas9 technology to delete putatively functional N-terminally truncated isoforms observed in previous knock-out models. While we showed that aberrant N-terminally truncated isoforms are not functional in mESCS, we observed that canonical Wnt signalling is not active in mESCs, as β-catenin ablation does not alter mESC transcriptional profile in LIF-enriched culture conditions; on the other hand, Wnt signalling activation represses mESC spontaneous differentiation. We also showed that transcriptionally silent β-catenin (ΔC) isoforms can rescue β-catenin knock-out self-renewal defects in mESCs, cooperating with TCF1 and LEF1 in the inhibition of mESC spontaneous differentiation in a Gsk3 dependent manner.
Project description:Analysis of genes involved in the mESC maintainance between different conditions of growth. The hypothesis tested in the present study was that Myc can sustain mESC self renewal and pluripotency. Results provide important information on the mechanism by which Myc supports mESC self renewal, in comparison with the routinely used LIF+serum culture condition, such as a Myc-dependent specific transcriptional program, which involves induction of an alternative Core Regulatory Network, modulation of signallign pathways (Wnt/bcat) and repression of developmental genes. Total RNA obtained from mESC growth 3 days in the indicated condition compared to the routinely used LIF+serum culture condition. Total RNA obtained from EpiSC growth 3 days in standard condition were also used as control for primed stem cell state.
