Inhibition of GSK3beta enhances both adhesive and signalling activities of beta-catenin in mouse embryonic stem cells.

BACKGROUND: GSK3beta (glycogen synthase kinase 3beta) regulates the expression level and activity of various target proteins, including beta-catenin. beta-Catenin is a co-activator of Wnt-dependent genes as well as a partner for transmembrane cadherins to mediate cell-to-cell adhesion. In some cases, inhibition of GSK3beta activity was shown to promote self-renewal of ESCs (embryonic stem cells), but immediate effects of GSK3beta inhibitors in these cells still remain elusive.
RESULTS: Here, we address the effects of GSK3beta inhibitors BIO (6-bromoindirubin-3'-oxime) and CHIR99021 on mESCs (mouse ESCs), focusing on modulation of beta-catenin activities. We found that, upon GSK3beta inhibition, the colonies of undifferentiated mESCs acquire a more compact morphology. This change is paralleled by two somewhat polar effects: (i) the accumulation of the beta-catenin, which is co-localized with E-cadherin at the plasma membrane, and the cytoplasmic, tyrosine unphosphorylated beta-catenin, which is able to bind the GST (glutathione transferase)-fused cytoplasmic domain of E-cadherin; and (ii) the accumulation of the tyrosine phosphorylated beta-catenin and its nuclear translocation that is accompanied by activation of the Tcf (T-cell factor)/beta-catenin-dependent transcription of Top-Flash reporter. The Tcf-mediated activation, however, does not affect most of the analysed Wnt-responsive genes involved in EMT (epithelial-mesenchymal transition) or cell-cycle progression, suggesting that the adhesive function of beta-catenin is dominant over transcription in undifferentiated mESCs. Treatment with BIO decreases proliferation rates of mESCs. This is not due to apoptosis, but rather to accumulation of cells in G1 phase of the cell cycle and is accompanied by down-regulation of the c-myc mRNA content.
CONCLUSION: Our results suggest that inhibition of GSK3beta activity in mESCs enhances both the beta-catenin/E-cadherin-mediated adhesion and the Tcf/beta-catenin-dependent transcription, but does not activate transcription in most of the examined genes involved in EMT and cell cycle progression.