From mESCs to mEPSCs : a proteomics-based study

Abstract

The mouse expanded potential stem cells (mEPSCs) can contribute to both embryonic and extra-embryonic tissues. They can be derived directly from preimplantation embryos, or converted from mouse embryonic stem cells (mESCs) which normally only have the capacity to give rise to the embryonic cell lineages. This mESCs-to-mEPSCs transition is achieved by culturing mESCs in the mEPSC medium which contains several small molecule inhibitors targeting key signaling pathways in development. Among them, XAV939, a tankyrase inhibitor, has been validated to stabilize Axin1 which is a negative regulator of canonical Wnt signaling and promotes β-catenin degradation. Notably, the increased protein level of Axin1 is verified in mEPSCs. Genetic overexpression of Axin1 can also 3 efficiently convert mESCs to mEPSCs, as validated in the chimera assay. To uncover the underlying molecular mechanism of Axin1 in the mESCs-mEPSCs transition, I made use of an Axin1 overexpression systems engineered into mESC line to study the underlying mechanism in chimera assay and in proteomics. The Affinity Purification combined with Mass Spectrometry analysis shows that more than one novel candidate is found as the potential binding candidate of the Axin1 protein. Among these candidates, I identified that the histone acetyltransferase Kat2a (Gcn5) exhibits bona fide interaction and its activities were negatively correlated with Axin1 levels and acquisition of totipotency in mouse ESCs.