title: Identification of Spen as a Crucial Factor for Xist Function through Forward Genetic Screening in Haploid Embryonic Stem Cells.
abstract: 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.
authors.affiliation: Institute of Molecular Health Sciences, Swiss Federal Institute of Technology, ETH Hönggerberg, Otto-Stern-Weg 7, 8093 Zurich, Switzerland.
authors.forename: Asun
authors.initials: A
authors.lastname: Monfort
authors.affiliation: Institute of Molecular Health Sciences, Swiss Federal Institute of Technology, ETH Hönggerberg, Otto-Stern-Weg 7, 8093 Zurich, Switzerland.
authors.forename: Giulio
authors.initials: G
authors.lastname: Di Minin
authors.affiliation: Institute of Molecular Health Sciences, Swiss Federal Institute of Technology, ETH Hönggerberg, Otto-Stern-Weg 7, 8093 Zurich, Switzerland.
authors.forename: Andreas
authors.initials: A
authors.lastname: Postlmayr
authors.affiliation: Institute of Molecular Health Sciences, Swiss Federal Institute of Technology, ETH Hönggerberg, Otto-Stern-Weg 7, 8093 Zurich, Switzerland.
authors.forename: Remo
authors.initials: R
authors.lastname: Freimann
authors.affiliation: CEITEC-Central European Institute of Technology, Masaryk University, Kamenice 753/5, Brno 62500, Czech Republic.
authors.forename: Fabiana
authors.initials: F
authors.lastname: Arieti
authors.affiliation: University of Bordeaux, European Institute for Chemistry and Biology (IECB), ARNA Laboratory, Bordeaux 33000, France; Institut National de la Sante et de la Recherche Medicale, INSERM, U869, ARNA Laboratory, Bordeaux 33000, France.
authors.forename: Stéphane
authors.initials: S
authors.lastname: Thore
authors.affiliation: Institute of Molecular Health Sciences, Swiss Federal Institute of Technology, ETH Hönggerberg, Otto-Stern-Weg 7, 8093 Zurich, Switzerland. Electronic address: awutz@ethz.ch.
authors.forename: Anton
authors.initials: A
authors.lastname: Wutz
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D004268:DNA-Binding Proteins; D053595:Embryonic Stem Cells / Q000378:metabolism*; D000071221:Enhancer of Zeste Homolog 2 Protein; D020868:Gene Silencing; D006238:Haploidy; D051379:Mice; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism*; D063151:Polycomb Repressive Complex 2 / Q000235:genetics / Q000378:metabolism; D062085:RNA, Long Noncoding / Q000235:genetics*; D016601:RNA-Binding Proteins
chemical_list: D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D062085:RNA, Long Noncoding; D016601:RNA-Binding Proteins; C121253:Spen protein, mouse; C077885:XIST non-coding RNA; D000071221:Enhancer of Zeste Homolog 2 Protein; C498410:Ezh2 protein, mouse; D063151:Polycomb Repressive Complex 2