Solving the "X" in embryos and stem cells

X-chromosome inactivation (XCI) is a complex epigenetic process that ensures that most X-linked genes are expressed equally for both sexes. Female eutherian mammals inactivate randomly the maternal or paternal inherited X-chromosome early in embryogenesis, whereas the extra-embryonic tissues experience an imprinting XCI that results in the inactivation of the paternal X-chromosome in mice. Although the phenomenon was initially described 40 years ago, many aspects remain obscure. In the last 2 years, some trademark publications have shed new light on the ongoing debate regarding the timing and mechanism of imprinted or random XCI. It has been observed that XCI is not accomplished at the blastocyst stage in bovines, rabbits, and humans, contrasting with the situation reported in mice, the standard model. All the species present 2 active X-chromosomes (Xa) in the early epiblast of the blastocyst, the cellular source for embryonic stem cells (ESCs). In this perspective, it would make sense to expect an absence of XCI in undifferentiated ESCs, but human ESCs are highly heterogeneous for this parameter and the presence of 2 Xa has been proposed as a true hallmark of ground-state pluripotency and a quality marker for female ESCs. Similarly, XCI reversal in female induced pluripotent stem cells is a key reprogramming event on the path to achieve the naïve pluripotency, and key pluripotency regulators can interact directly or indirectly with Xist. Finally, the presence of 2 Xa may lead to a sex-specific transcriptional regulation resulting in sexual dimorphism in reprogramming and differentiation.