title: Revealing long noncoding RNA architecture and functions using domain-specific chromatin isolation by RNA purification.
abstract: Little is known about the functional domain architecture of long noncoding RNAs (lncRNAs) because of a relative paucity of suitable methods to analyze RNA function at a domain level. Here we describe domain-specific chromatin isolation by RNA purification (dChIRP), a scalable technique to dissect pairwise RNA-RNA, RNA-protein and RNA-chromatin interactions at the level of individual RNA domains in living cells. dChIRP of roX1, a lncRNA essential for Drosophila melanogaster X-chromosome dosage compensation, reveals a 'three-fingered hand' ribonucleoprotein topology. Each RNA finger binds chromatin and the male-specific lethal (MSL) protein complex and can individually rescue male lethality in roX-null flies, thus defining a minimal RNA domain for chromosome-wide dosage compensation. dChIRP improves the RNA genomic localization signal by >20-fold relative to previous techniques, and these binding sites are correlated with chromosome conformation data, indicating that most roX-bound loci cluster in a nuclear territory. These results suggest dChIRP can reveal lncRNA architecture and function with high precision and sensitivity.
authors.affiliation: Howard Hughes Medical Institute and Program in Epithelial Biology, Stanford University School of Medicine, Stanford CA 94305, USA.|Department of Bioengineering, Stanford University Schools of Medicine and Engineering, Stanford CA 94305, USA.
authors.forename: Jeffrey J
authors.initials: JJ
authors.lastname: Quinn
authors.affiliation: Max Planck Institute of Immunobiology and Epigenetics, Stübeweg 51, 79108 Freiburg im Breisgau, Germany.
authors.forename: Ibrahim A
authors.initials: IA
authors.lastname: Ilik
authors.affiliation: Howard Hughes Medical Institute and Program in Epithelial Biology, Stanford University School of Medicine, Stanford CA 94305, USA.
authors.forename: Kun
authors.initials: K
authors.lastname: Qu
authors.affiliation: Max Planck Institute of Immunobiology and Epigenetics, Stübeweg 51, 79108 Freiburg im Breisgau, Germany.
authors.forename: Plamen
authors.initials: P
authors.lastname: Georgiev
authors.affiliation: Howard Hughes Medical Institute and Program in Epithelial Biology, Stanford University School of Medicine, Stanford CA 94305, USA.
authors.forename: Ci
authors.initials: C
authors.lastname: Chu
authors.affiliation: Max Planck Institute of Immunobiology and Epigenetics, Stübeweg 51, 79108 Freiburg im Breisgau, Germany.
authors.forename: Asifa
authors.initials: A
authors.lastname: Akhtar
authors.affiliation: Howard Hughes Medical Institute and Program in Epithelial Biology, Stanford University School of Medicine, Stanford CA 94305, USA.
authors.forename: Howard Y
authors.initials: HY
authors.lastname: Chang
mesh_terms: D000818:Animals; D001665:Binding Sites; D002843:Chromatin / Q000235:genetics* / Q000302:isolation & purification; D004303:Dosage Compensation, Genetic; D005260:Female; D008297:Male; D012313:RNA / Q000302:isolation & purification*; D062085:RNA, Long Noncoding / Q000235:genetics*
chemical_list: D002843:Chromatin; D062085:RNA, Long Noncoding; D012313:RNA