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Cell. 2015 Apr 9;161(2):404-16. doi: 10.1016/j.cell.2015.03.025. Epub 2015 Apr 2.

Systematic discovery of Xist RNA binding proteins.

Author information

1
Howard Hughes Medical Institute and Program in Epithelial Biology, Stanford University School of Medicine, Stanford, CA 94305, USA; Genome Institute of Singapore, 60 Biopolis Street, Singapore 138672, Singapore.
2
Howard Hughes Medical Institute and Program in Epithelial Biology, Stanford University School of Medicine, Stanford, CA 94305, USA.
3
Institut Curie, CNRS UMR3215, INSERM U934, 26 rue d'Ulm, Paris 75248, France.
4
Department of Pharmacology and Lineberger Comprehensive Cancer Center, University of North Carolina, Chapel Hill, NC 27599, USA.
5
Department of Genetics and Lineberger Comprehensive Cancer Center, University of North Carolina, Chapel Hill, NC 27599, USA.
6
Howard Hughes Medical Institute and Program in Epithelial Biology, Stanford University School of Medicine, Stanford, CA 94305, USA. Electronic address: howchang@stanford.edu.

Abstract

Noncoding RNAs (ncRNAs) function with associated proteins to effect complex structural and regulatory outcomes. To reveal the composition and dynamics of specific noncoding RNA-protein complexes (RNPs) in vivo, we developed comprehensive identification of RNA binding proteins by mass spectrometry (ChIRP-MS). ChIRP-MS analysis of four ncRNAs captures key protein interactors, including a U1-specific link to the 3' RNA processing machinery. Xist, an essential lncRNA for X chromosome inactivation (XCI), interacts with 81 proteins from chromatin modification, nuclear matrix, and RNA remodeling pathways. The Xist RNA-protein particle assembles in two steps coupled with the transition from pluripotency to differentiation. Specific interactors include HnrnpK, which participates in Xist-mediated gene silencing and histone modifications but not Xist localization, and Drosophila Split ends homolog Spen, which interacts via the A-repeat domain of Xist and is required for gene silencing. Thus, Xist lncRNA engages with proteins in a modular and developmentally controlled manner to coordinate chromatin spreading and silencing.

PMID:
25843628
PMCID:
PMC4425988
DOI:
10.1016/j.cell.2015.03.025
[Indexed for MEDLINE]
Free PMC Article
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