Transcriptional mechanisms governing the exit of mouse embryonic stem cells (mESCs) from the pluripotency ground state

Abstract

Embryonic stem cells (ESCs) are non-specialized cells able to self-renew and to differentiate in vitro giving rise to all cell types of an organism, mimicking the events that take place in vivo during the early stages of the development. For these characteristics, a deep knowledge of the mechanisms that govern ESC fate is fundamental for both basic research and cell replacement therapy. In the recent years, enormous progresses have been made in the understanding of ESC pluripotency, while the mechanisms governing the ESC differentiation and, in particular, the mechanisms governing the exit from the pluripotent state and the first steps of differentiation are still not definitively understood. In this context, our group has developed a systematic approach based on the screening of a shRNA library to identify factors governing ESC fate. Among the identified genes, we have found the high mobility group AT-hook 2 (HMGA2), a non-histone chromatin factor that is widely expressed during embryogenesis and is known to have important roles in development and cell adhesion/differentiation processes. We have found that the knockdown of HMGA2 promotes the maintenance of pluripotent state. Indeed, in conditions promoting differentiation, HMGA2 knock-down cells maintain the expression of stemness markers Oct3/4, Nanog. Interestingly, we have found that the expression of stemness markers is accompanied by a decrease of neuro-ectodermal marker Sox1. A more detailed analysis of the phenotype induced by HMGA2 suppression has shown that the block of differentiation occurs during first steps, i.e. the transition from ESCs to epiblast stem cells (EpiSCs). The same effects, accompanied by a more dramatic phenotype, were observed upon differentiation of induced pluripotent stem cells (iPS cells) knock-out for HMGA2. We have found that HMGA2 expression is regulated by a key transcription factor Otx2 that alone or in combination with Oct4 drives early enhancer activation during the exit from ground state of ESCs to EpiSCs. Indeed, we have shown that HMGA2 cooperates with Otx2 in enhanceosome formation. Finally, the results obtained during my doctoral thesis have demonstrated that HMGA2 acts taking part to the regulatory mechanisms that guide the exit of ESCs from ground state.