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Cell cycle regulation of ES cell identity and reprogramming potential

Final Report Summary - REPLENICHE (Cell cycle regulation of ES cell identity and reprogramming potential)

Our team has been investigating how experimental reprogramming (turning one type of cell into another) works and asking whether cells that can divide are particularly prone to this sort of conversion. We have linked up with bio-engineers and used a variety of different tools to try and answer this question. We want to know how conversion occurs to try and make it more efficient and safe, so that in the longer-term we may be able to provide ‘replacement’ human tissues that are damaged by disease, infection or aging.

Our studies began by comparing how well we could reprogram cells that differed only in terms of where they were in the cell division cycle. To reprogram we fused, or joined, two different types of cell together and watched what happened. Previous studies had shown that stem cells from embryos (so-called embryonic stem cells) are very good at converting cells isolated from the body such as white blood cells, to become just like themselves; a property called ‘dominance’. We found out that one of the main reasons that they can do this is because they force blood cells to begin the process of DNA replication before they were ready (something we have termed precocious DNA replication). We can exploit this new understanding to improve reprogramming efficiencies around ten-fold.

We also investigated other sources of embryonic stem cells and found that embryonic germ cell lines could reprogram and also alter the expression of a group of genes called imprinted genes. The expression of these genes is normally resistant to alteration during reprogramming and we were able to show that an enzyme called Tet1 was required to reset the expression of imprinted genes.
These studies were technically challenging since there is a paucity of reporters that reliably tracked the expression of individual imprinted loci. Therefore, as part of work funded by the Medical Research Council UK, we generated a series of new mouse models where imprinted gene expression can be followed experimentally and in vivo using luciferase-based imaging. This work was published recently and illustrates how ERC funded research, together with national funding streams, can be used to generate important new tools for the research community.

Finally, we used our knowledge of reprogramming by cell-cell fusion to study how genes contained within a single human chromosome (the X chromosome) could be reactivated. In humans, genes from one of the two X chromosomes in female cells are expressed. The second X chromosome remains largely silent. Fusion of female human blood cells or fibroblasts with embryonic stem cells provokes their reprogramming and the partial reactivation of genes on the previously silent human X chromosome. We were able to demonstrate this and to carefully describe the molecular events that were required to reactivate genes on the human X chromosome. We showed some genes were reproducibly more susceptible to reactivation than others and that drugs that modified DNA (or the proteins bound to DNA) could be used to enhance the extent of reactivation. These exciting results suggest that in the future it may be possible to engineer reprogramming strategies that can compensate for a range of defects and disease states that are caused by mutations of genes on the human X chromosome.