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Learn MoreUnderstanding the mechanism of resistance of certain genes to reactivation will help improving the success of nuclear reprogramming. Here, in a Xenopus oocyte nuclear transfer assay we identify mouse genes that are resistant to reprogramming by oocyte factors. By using MEF nuclei with WT or reduced DNA methylation in combination with chromatin modifiers able to erase H3K9me3, H3K27me3, and H2Aub from transplanted nuclei, we reveal the basis for resistance to transcriptional reprogramming. We observe a complex relationship between loss of resistance and chromatin modifiers treatment as a majority of gene is affected by more than one type of treatment, suggesting that resistance requires simultaneous repression through multiple epigenetic mechanisms. Existence of synergistic as well as adverse mechanism of action of chromatin modifiers on removal of resistance is revealed by the classification of resistant genes according to their sensitivity to chromatin modifiers treatments. We further demonstrate that removal of H2Aub from transplanted chromatin explain loss of resistance upon USP21 expression. Finally, we provide evidence that H2A ubiquitylation is also contributing to resistance to transcriptional reprogramming in mouse nuclear transfer embryos. SOURCE: Charles Bradshaw University of Cambridge
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