title: High-Resolution Mapping of RNA-Binding Regions in the Nuclear Proteome of Embryonic Stem Cells.
abstract: Interactions between noncoding RNAs and chromatin proteins play important roles in gene regulation, but the molecular details of most of these interactions are unknown. Using protein-RNA photocrosslinking and mass spectrometry on embryonic stem cell nuclei, we identified and mapped, at peptide resolution, the RNA-binding regions in ∼800 known and previously unknown RNA-binding proteins, many of which are transcriptional regulators and chromatin modifiers. In addition to known RNA-binding motifs, we detected several protein domains previously unknown to function in RNA recognition, as well as non-annotated and/or disordered regions, suggesting that many functional protein-RNA contacts remain unexplored. We identified RNA-binding regions in several chromatin regulators, including TET2, and validated their ability to bind RNA. Thus, proteomic identification of RNA-binding regions (RBR-ID) is a powerful tool to map protein-RNA interactions and will allow rational design of mutants to dissect their function at a mechanistic level.
authors: Epigenetics Program, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA; Department of Cell and Developmental Biology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA.
authors: Chongsheng
authors: C
authors: He
authors: Epigenetics Program, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA; Department of Biochemistry and Biophysics, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA.
authors: Simone
authors: S
authors: Sidoli
authors: Epigenetics Program, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA; Graduate Group in Biochemistry and Biophysics, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA.
authors: Robert
authors: R
authors: Warneford-Thomson
authors: Department of Biochemistry and Biophysics, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA.
authors: Deirdre C
authors: DC
authors: Tatomer
authors: Department of Biochemistry and Biophysics, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA.
authors: Jeremy E
authors: JE
authors: Wilusz
authors: Epigenetics Program, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA; Department of Biochemistry and Biophysics, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA.
authors: Benjamin A
authors: BA
authors: Garcia
authors: Epigenetics Program, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA; Department of Cell and Developmental Biology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA. Electronic address: rbon@mail.med.upenn.edu.
authors: Roberto
authors: R
authors: Bonasio
mesh_terms: D000818:Animals; D001665:Binding Sites; D002843:Chromatin / Q000737:chemistry* / Q000378:metabolism / Q000528:radiation effects; D015870:Gene Expression; D057809:HEK293 Cells; D006801:Humans; D051379:Mice; D000066450:Mouse Embryonic Stem Cells / Q000166:cytology / Q000378:metabolism* / Q000528:radiation effects; D009687:Nuclear Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D009690:Nucleic Acid Conformation; D010449:Peptide Mapping / Q000379:methods; D055668:Photochemical Processes; D011485:Protein Binding; D054730:Protein Interaction Domains and Motifs; D017433:Protein Structure, Secondary; D020543:Proteome / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D040901:Proteomics / Q000379:methods; D022661:RNA, Untranslated / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D014466:Ultraviolet Rays
chemical_list: D002843:Chromatin; D009687:Nuclear Proteins; D020543:Proteome; D022661:RNA, Untranslated; D016601:RNA-Binding Proteins