The Xist lncRNA interacts directly with SHARP to silence transcription through HDAC3
Copyright policy )The Xist lncRNA interacts directly with SHARP to silence transcription through HDAC3
Many long non-coding RNAs (lncRNAs) affect gene expression, but the mechanisms by which they act are still largely unknown. One of the best-studied lncRNAs is Xist, which is required for transcriptional silencing of one X chromosome during development in female mammals. Despite extensive efforts to define the mechanism of Xist-mediated transcriptional silencing, we still do not know any proteins required for this role. The main challenge is that there are currently no methods to comprehensively define the proteins that directly interact with a lncRNA in the cell. Here we develop a method to purify a lncRNA from cells and identify proteins interacting with it directly using quantitative mass spectrometry. We identify ten proteins that specifically associate with Xist, three of these proteins--SHARP, SAF-A and LBR--are required for Xist-mediated transcriptional silencing. We show that SHARP, which interacts with the SMRT co-repressor that activates HDAC3, is not only essential for silencing, but is also required for the exclusion of RNA polymerase II (Pol II) from the inactive X. Both SMRT and HDAC3 are also required for silencing and Pol II exclusion. In addition to silencing transcription, SHARP and HDAC3 are required for Xist-mediated recruitment of the polycomb repressive complex 2 (PRC2) across the X chromosome. Our results suggest that Xist silences transcription by directly interacting with SHARP, recruiting SMRT, activating HDAC3, and deacetylating histones to exclude Pol II across the X chromosome.
- California Institute of Technology United States
- University of California, San Francisco United States
- University of California Los Angeles United States
- Rowland Institute at Harvard United States
- University of California at Los Angeles United States
Male, DNA-Binding Proteins (mesh), Cytoplasmic and Nuclear, 3102 Bioinformatics and Computational Biology (for-2020), General Science & Technology (science-metrix), Mass Spectrometry (mesh), Heterogeneous-Nuclear Ribonucleoprotein U, Nuclear Receptor Co-Repressor 2 (mesh), Mass Spectrometry, Histones, Mice, X Chromosome Inactivation, Receptors, Protein Binding (mesh), X Chromosome (mesh), Animals (mesh), Male (mesh), Lamin B Receptor, Generic health relevance (hrcs-hc), Polycomb Repressive Complex 2, Mice (mesh), Nuclear Proteins, RNA-Binding Proteins, Acetylation, Biological Sciences, Histones (mesh), Gene Silencing (mesh), DNA-Binding Proteins, Long Noncoding, Female, RNA, Long Noncoding, RNA Polymerase II, Transcription, Lamin B Receptor (mesh), Protein Binding, X Chromosome, General Science & Technology, 1.1 Normal biological development and functioning, Bioinformatics and Computational Biology, Acetylation (mesh), Histone Deacetylases, Cell Line, Genetic, Nuclear Proteins (mesh), Underpinning research, Genetics, Long Noncoding (mesh), Animals, Nuclear Receptor Co-Repressor 2, Gene Silencing, Polycomb Repressive Complex 2 (mesh), Embryonic Stem Cells, 31 Biological Sciences (for-2020), Heterogeneous-Nuclear Ribonucleoprotein U (mesh), Genetics (rcdc), Histone Deacetylases (mesh), RNA Polymerase II (mesh), X Chromosome Inactivation (mesh), RNA-Binding Proteins (mesh), Embryonic Stem Cells (mesh), Female (mesh), Cytoplasmic and Nuclear (mesh), Cell Line (mesh), RNA, Genetic (mesh), Generic health relevance
Male, DNA-Binding Proteins (mesh), Cytoplasmic and Nuclear, 3102 Bioinformatics and Computational Biology (for-2020), General Science & Technology (science-metrix), Mass Spectrometry (mesh), Heterogeneous-Nuclear Ribonucleoprotein U, Nuclear Receptor Co-Repressor 2 (mesh), Mass Spectrometry, Histones, Mice, X Chromosome Inactivation, Receptors, Protein Binding (mesh), X Chromosome (mesh), Animals (mesh), Male (mesh), Lamin B Receptor, Generic health relevance (hrcs-hc), Polycomb Repressive Complex 2, Mice (mesh), Nuclear Proteins, RNA-Binding Proteins, Acetylation, Biological Sciences, Histones (mesh), Gene Silencing (mesh), DNA-Binding Proteins, Long Noncoding, Female, RNA, Long Noncoding, RNA Polymerase II, Transcription, Lamin B Receptor (mesh), Protein Binding, X Chromosome, General Science & Technology, 1.1 Normal biological development and functioning, Bioinformatics and Computational Biology, Acetylation (mesh), Histone Deacetylases, Cell Line, Genetic, Nuclear Proteins (mesh), Underpinning research, Genetics, Long Noncoding (mesh), Animals, Nuclear Receptor Co-Repressor 2, Gene Silencing, Polycomb Repressive Complex 2 (mesh), Embryonic Stem Cells, 31 Biological Sciences (for-2020), Heterogeneous-Nuclear Ribonucleoprotein U (mesh), Genetics (rcdc), Histone Deacetylases (mesh), RNA Polymerase II (mesh), X Chromosome Inactivation (mesh), RNA-Binding Proteins (mesh), Embryonic Stem Cells (mesh), Female (mesh), Cytoplasmic and Nuclear (mesh), Cell Line (mesh), RNA, Genetic (mesh), Generic health relevance
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