As female embryonic stem cells differentiate, one of their X chromosomes is inactivated. Indeed, blocking X-chromosome inactivation disrupts differentiation. And when differentiated female cells from any source are reprogrammed to behave like embryonic stem cells, inactive X chromosomes are reactivated. It would make sense, then, that the paths to and from pluripotency and X-chromosome inactivation are governed by the same proteins. Publishing in Nature, Jeannie Lee and colleagues at Harvard Medical School in Boston explain how this process works and reveal yet another role for the pluripotency regulator Oct4 (also called POUF5f)1.

Previous work had shown that genes essential for pluripotency (Nanog, Oct4, and Sox2) act together to temporarily repress gene expression on paternal X chromosomes2. Using bioinformatic scanning within a region known as the X inactivation centre (XIC), Lee and colleagues confirmed and identified binding sites for Oct4 within regions called Tsix and Xite, two regulatory noncoding RNA genes. They went on to work out a mechanism of action and showed that Oct4 bound to Tsix and Xite and enhanced their transcription. This, in turn, led to the repression of another, non–protein coding gene called Xist, which coats and inactivates the X chromosome. “Our analysis indicates that Oct4's interaction with Tsix/Xite may be the first direct connection between the pluripotency factor and the X-inactivation centre,” says Lee.

The protein seems to control how the X chromosomes pair with each other, and so is at the “top of the pathway” when X-chromosome inactivation initiates, says Lee. As Oct4 levels fall during differentiation, Lee says, there may only be enough of the protein to repress Xist on one of the chromosomes — the one fated to remain active. And Sox2 is also involved in these processes: it binds Xite, may co-bind Oct4 on Tsix and also binds genes for sorting chromosomes into daughter cells.

Working out the epigenetic reprogramming for X chromosomes could explain fundamental biology and uncover ways to attack X-chromosome–linked diseases, and it could also help with strategies to reprogram more cells. Anecdotally, Lee explains, reprogramming seems to happen more easily in male cells.

It's an interesting finding, says Kathrin Plath of the University of California, Los Angeles, an expert on both X-chromosome inactivation and reprogramming, but she'd like to see more experiments to understand just how Oct4 regulates X-chromosome inactivation.

A major question going forward is how Oct4 turns on the switch to reprogram the X chromosome, Lee says. In particular, why does cell differentiation stall if X-chromosome inactivation is incomplete? Such questions could provide significant insight into how embryonic stem cells differentiate and how reprogramming differentiated cells to pluripotency reverses this process.

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