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  • Finally by sorting the Wnt off cells we obtained a

    2018-11-14

    Finally, by sorting the “Wnt-off” cells, we obtained a striking increase in the number of reprogrammed NANOG+ clones, thus clearly identifying the “Wnt-off” state as an early reprogramming marker. It is interesting to note how different the roles of TCF3 and TCF1 are in the control of somatic cell reprogramming. We showed previously that leukotriene receptor antagonist of Tcf3 induces increased AcH3 and a decreased number of H3K9me3 heterochromatin foci. These epigenome modifications ultimately enhance reprogramming efficiency (Lluis et al., 2011). Here, we showed that the role of TCF1 during the early reprogramming phase is correlated with the repression of senescence genes and with the activation of MET. Ho et al. (2013) provided evidence that TCF3 and TCF4 promote early reprogramming events by repressing Wnt pathway target genes, including TCF1 and LEF1. However, they also postulated that TCF3 and TCF4 could control other targets that are independent of TCF1 and LEF1 or of the Wnt pathway activation (Ho et al., 2013). It is tempting to speculate that the latter mentioned might be activated by the epigenetic modifications induced by Tcf3 derepression. Very little is known about TCF1 activity in ESCs, although it has been demonstrated that in medium lacking LIF, TCF1 contributes to the effect of Wnt3a stimulation to ESC self-renewal (Yi et al., 2011). Here, we have revealed that in medium containing LIF and serum, wild-type ESCs and ESCs silenced for Tcf1 do not show differences in self-renewal and differentiation, indicating that TCF1 does not control either process. TCF1 is instead a key regulator of the reprogramming process. This led to the interesting observation that some key transcription factors do not always control both somatic cell reprogramming processes and ESC self-renewal, as the effects on these two processes can be distinct. Both MET and the downregulation of senescence genes are essential processes to achieve somatic cell reprogramming (Esteban et al., 2012; Mahmoudi and Brunet, 2012). Our data show that TCF1 is an important regulator of these processes. Senescent cells are characterized by cell-cycle arrest due to p16INK4a induction. Cell proliferation is crucial in the reprogramming process, and thus cell-cycle arrest is a major barrier to the efficiency of this process. Indeed, mouse fibroblasts cannot be reprogrammed efficiently when two antiproliferative genes, p16 and p19, which are part of the lnk4/Arf locus, are highly expressed. Accordingly, silencing of the lnk4/Arf locus restores reprogramming efficiency in senescent cells. Furthermore, knockdown of p53 and p21 also accelerates reprogramming of human and mouse fibroblasts (Banito et al., 2009; Hong et al., 2009; Kawamura et al., 2009; Li et al., 2009; Marión et al., 2009; Utikal et al., 2009). Thus, senescence is a barrier to the reprogramming activity, and this can be overcome by TCF1 action, which maintains low expression of p21 and p19 during reprogramming. During MET, SOX2, OCT4, and C-MYC have been shown to supress Snai1 and TGF-β signaling, whereas KLF4 has been shown to upregulate E-cadherin (Li et al., 2010; Samavarchi-Tehrani et al., 2010). Here, we found that TCF1 activity is correlated not only with repression of senescence genes, but also with MET activation. Embryoid bodies derived from ESCs undergo epithelial-to-mesenchymal transition in a Wnt-dependent process, with the Wnt activity inducing upregulation of Snai1 and repression of E-cadherin (ten Berge et al., 2008). It is therefore intriguing that during reprogramming, TCF1 enhances MET by transcriptionally repressing Vim, leukotriene receptor antagonist Slug, and Snai1 and upregulating E-cadherin. Furthermore, we observed that TCF1 directly represses Snai1 by binding to its promoter. SNAI1 has been shown to be a transcriptional repressor of E-cadherin (Batlle et al., 2000). Thus, it might well be that E-cadherin is derepressed as a consequence of TCF1-mediated repression of Snai1.