Identification of Spen as a Crucial Factor for Xist Function through Forward Genetic Screening in Haploid Embryonic Stem Cells

By Asun Monfort, Giulio Di Minin, Andreas Postlmayr, Remo Freimann, Fabiana Arieti, S. Thore, Anton Wutz. 2015

230 citations recorded by the source corpus.

Summary In mammals, the noncoding Xist RNA triggers transcriptional silencing of one of the two X chromosomes in female cells. Here, we report a genetic screen for silencing factors in X chromosome inactivation using haploid mouse embryonic stem cells (ESCs) that carry an engineered selectable reporter system. This system was able to identify several candidate factors that are genetically required for chromosomal repression by Xist. Among the list of candidates, we identify the RNA-binding protein Spen, the homolog of split ends. Independent validation through gene deletion in ESCs confirms that Spen is required for gene repression by Xist. However, Spen is not required for Xist RNA localization and the recruitment of chromatin modifications, including Polycomb protein Ezh2. The identification of Spen opens avenues for further investigation into the gene-silencing pathway of Xist and shows the usefulness of haploid ESCs for genetic screening of epigenetic pathways.

Subfields: Genetics, Molecular Biology, Plant Science

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Citations in the corpus

How do lncRNAs regulate transcription? (2017, methodology): “Finally, two independent groups (63, 64) used elegant genetic screens to determine which genes are required for Xist-mediated XCI.”
Time-resolved structured illumination microscopy reveals key principles of Xist RNA spreading (2020, background): “The protein SPEN, which binds to the Arepeat region of Xist RNA through a triple RRM domain, plays a central role in Xistmediated gene silencing (8, 17, 22, 23).”
Xist nucleates local protein gradients to propagate silencing across the X chromosome (2021, background): “The protein SPEN is essential for initiating the silencing of virtually all X-linked genes (Chu et al., 2015; Dossin et al., 2020; McHugh et al., 2015; Moindrot et al., 2015; Monfort et al., 2015).”
Cross-talk between PRMT1-mediated methylation and ubiquitylation on RBM15 controls RNA splicing (2015, background): “Recently SPEN and RBM15 have been shown to be essential for Xist-mediated X 68 chromosome inactivation (Chu et al., 2015; McHugh et al., 2015; Minajigi et al., 2015; Moindrot et al., 69 2015; Monfort et al., 2015).”
RNA duplex map in living cells reveals higher order transcriptome structure (2016, background): “Previous studies of SPEN RRM domains suggest that they bind many RNA species, without preference 1276 Cell 165, 1267–1279, May 19, 2016 for single copies of the A-repeat motif (Monfort et al., 2015).”