Tri-lineage Differentiation of Embryonic Stem Cells: Role of Wnt Signaling.

Abstract

Since embryonic stem cells (ESC) and the epiblast share a gene expression profile and an attenuated cell cycle, ESC are an attractive model system to study lineage choice at gastrulation. We have employed mouse ESC as a model system to examine the role of Wnt signaling in the transition from pluripotency to neural precursors and have identified a new function for the Geminin gene in mesendodermal differentiation. Wnt signaling is required for differentiation of both epiblast and ESC to the mesendodermal lineage. To test the hypothesis that blocking Wnt signaling in the absence of BMP signaling would promote differentiation of mESC to neural precursors, we developed a mESC line that inducibly expresses a dominant negative Tcf4 (dnTcf4) protein to block canonical Wnt signaling. Cells expressing the dnTcf4 protein differentiated largely to Sox3 positive neural precursors but were unable to progress to beta-III tubulin positive neurons unless Wnt signaling was de-repressed. This cell line will provide an important resource for other investigators studying the role of Wnt signaling in embryonic development. Geminin has been described as a bi-functional protein that inhibits endoreduplication during cell division and promotes neural differentiation by inhibiting BMP signaling. Geminin also binds transcription factors and chromatin remodeling proteins in the nucleus to inhibit their function. In mESC over-expression of Geminin induced mesendodermal differentiation and an epithelial to mesenchymal transition (EMT) via increasing Wnt signaling. We hypothesize that Geminin binds to Groucho/TLE proteins in the nucleus thereby dis-inhibiting Wnt target gene transcritption by Tcf/Lef proteins. Germinin is frequently over-expressed in cancer cells and EMT is an integral part of cancer metastasis. These observations make Geminin an attractive candidate for cancer therapy; reduction of Geminin protein could reduce metastasis as well as induce cell death due to endoreduplication. Overall, this work demonstrates that Wnt signaling is required for mesendodermal differentiation, EMT, and the transition from proliferative neural precursor to primitive neurons during lineage allocation of mESC.