Abrogation of E-Cadherin-Mediated Cell–Cell Contact in Mouse Embryonic Stem Cells Results in Reversible LIF-Independent Self-Renewal

Abstract

We have previously demonstrated that differentiation of embryonic stem (ES) cells is associated with downregulation of cell surface E-cadherin. In this study, we assessed the function of E-cadherin in mouse ES cell pluripotency and differentiation. We show that inhibition of E-cadherin-mediated cell-cell contact in ES cells using gene knockout (Ecad , EcadRNAi, and CHAVC-treated ES cells to the activin receptor-like kinase inhibitor SB431542 led to differentiation of the cells, which could be prevented by re-expression of E-cadherin. To confirm the role of transforming growth factor b family signaling in the self-renewal of Ecad 2/2 ES cells, we show that these cells maintain an undifferentiated phenotype when cultured in serum-free medium supplemented with Activin A and Nodal, with fibroblast growth factor 2 required for cellular proliferation. We conclude that transhomodimerization of E-cadherin protein is required for LIF-dependent ES cell selfrenewal and that multiple self-renewal signaling networks subsist in ES cells, with activity dependent upon the cellular context.

The Function of E-cadherin in ES Cell Pluripotency

“…Inhibition of Ecadherin expression using RNAi or an inhibitory peptide during iPS cell derivation lead to decreased iPS cell isolation. Interestingly, in contrast to signalling pathway alterations described by Soncin et al (Soncin et al, 2009), the -catenin binding domain of E-cadherin was not required for efficient iPS cell derivation and instead was dependent on the extracellular domain of the protein. We have previously demonstrated that the -catenin domain of E-cadherin is required to restore cell-cell contact in E-cadherin-/-ES cells (Soncin et al, 2009), suggesting that the function of E-cadherin in iPS cell derivation reflects a requirement for cell-extracellular matrix (ECM) interaction rather than cell-cell adhesion.…”

“…Interestingly, in contrast to signalling pathway alterations described by Soncin et al (Soncin et al, 2009), the -catenin binding domain of E-cadherin was not required for efficient iPS cell derivation and instead was dependent on the extracellular domain of the protein. We have previously demonstrated that the -catenin domain of E-cadherin is required to restore cell-cell contact in E-cadherin-/-ES cells (Soncin et al, 2009), suggesting that the function of E-cadherin in iPS cell derivation reflects a requirement for cell-extracellular matrix (ECM) interaction rather than cell-cell adhesion. Therefore, E-cadherin appears to function via two discreet mechanisms; firstly, as a regulator of pluripotent signalling pathways via the CCC and, secondly, as an enhancer of ES cell-ECM interactions to aid cell survival.…”

“…Although E-cadherin-/-ES cells exhibit an undifferentiated phenotype in medium supplemented with FBS/LIF, the cells maintain pluripotency and self-renewal by utilising Activin, Nodal and Fgf2 present within ES-screened FBS (Soncin et al, 2009) (Fig. 3.).…”

“…3.). E-cadherin-/-ES cells can be cultured in an undifferentiated state in serum-free medium supplemented with Activin A, Nodal and Fgf2 and exposure of the cells to the Activin-like kinase receptors (Alks)-4, -5 and -7 inhibitor (SB431542) under these conditions induces their differentiation (Soncin et al, 2009). Mutant E-cadherin protein analysis in our lab demonstrated that the -catenin binding domain of E-cadherin is critical for LIF/Bmp-mediated pluripotency in ES cells.…”

“…Mutant E-cadherin protein analysis in our lab demonstrated that the -catenin binding domain of E-cadherin is critical for LIF/Bmp-mediated pluripotency in ES cells. Although Ecadherin-/-ES cells utilise Activin/Nodal signalling as a default pluripotent pathway in serum-supplemented medium they are also able to maintain pluripotency via LIF/BMP signals in the absence of Activin/Nodal (Soncin et al, 2009). Therefore, E-cadherin-/-ES cells possess a functional "ground state" pluripotent signalling network (Ying et al, 2008) as well as the ability to circumvent this pathway by utilising Activin and Nodal.…”