Wnt signaling promotes reprogramming of somatic cells to pluripotency.

Mice, cell lines, retroviral vectors, cell culture, flow cytometry, immunostainings, teratoma formation and blastocyst injections were previously described in detail (Wernig et al., 2007; Meissner et al., 2007; Brambrink et al., 2008). Wnt3a conditioned media (Willert et al., 2003) and control conditioned media from parental L cells were generated according to standard protocols (ATCC), and used in a 1:1 dilution with standard ES cell medium. It has been previously shown that the conditioned media from Wnt3aoverexpressing cells has Wnt signaling activity as determined by TOP flash assays (Ogawa et al., 2006) and that functional Wnt3a protein can be purified from this media (Willert et al., 2003). Furthermore, secretion of Wnt3a protein can be detected by immunoblot of the conditioned media from Wnt3a-overexpressing cells (Shibamoto et al., 1998). ICG-001 was dissolved in DMSO to a stock concentration of 0.1M and used at a final, working concentration of 4uM (or as indicated in Supplemental Figure S2).