Periodic Activation of Wnt/beta-Catenin Signaling Enhances Somatic Cell Reprogramming Mediated by Cell Fusion

Cosma M.P *, Lluis F , Pedone E , Pepe ,


Objective: Reprogramming of nuclei allows the dedifferentiation of differentiated cells. Somatic cells can undergo epigenetic modifications and reprogramming through their fusion with embryonic stem cells (ESCs) or after overexpression of a specific blend of ESC tranh1ion factor-encoding genes. Our goal was to demonstrate that the activation of Wnt/beta-catenin signaling triggered somatic cell reprogramming. Materials and Methods: We performed cell fusion experiments by fusing ES cells with Neural stem cells or ES cells with Thymocytes or ES cells with Mouse Embryonic Fibroblasts. ES cells or the hybrids were treated with Wnt3a or BIO (Wnt pathway activator) or Dkk1 (Wnt pathway inhibitor). Fusion between ES cells over-expressing different amounts of beta-catenin with NS cells were also carried out. Reprogrammed cells were analyzed in vivo and in vitro for their ability to differentiate in different cell types. Expression of stem cell markers and methylation profile of stem cell promoters was also analyzed in the reprogrammed cells. For further details see Lluis et al. Cell Stem Cell 2008. Results: We showed that cyclic activation of Wnt/beta-catenin signaling in ESCs with Wnt3a or the glycogen synthase kinase-3 (GSK-3) inhibitor 6-bromoindirubin-3’-oxime (BIO) strikingly enhanced the ability of ESCs to reprogram somatic cells after fusion. In addition we showed that reprogramming is triggered by a dose-dependent accumulation of active beta-catenin. Reprogrammed clones expressed ESC-specific genes silenced somatic differentiation markers became demethylated on Oct4 and Nanog CpG islands and were able to differentiate into cardiomyocytes in vitro and to generate teratomas in vivo. Conclusion: Our data thus demonstrate that in ESCs periodic beta-catenin accumulation via the Wnt/beta-catenin pathway provides a specific threshold that leads to the reprogramming of somatic cells after fusion.