title: Dissecting the nascent human transcriptome by analysing the RNA content of transcription factories.
abstract: While mapping total and poly-adenylated human transcriptomes has now become routine, characterizing nascent transcripts remains challenging, largely because nascent RNAs have such short half-lives. Here, we describe a simple, fast and cost-effective method to isolate RNA associated with transcription factories, the sites responsible for the majority of nuclear transcription. Following stimulation of human endothelial cells with the pro-inflammatory cytokine TNFα, we isolate and analyse the RNA content of factories by sequencing. Comparison with total, poly(A)(+) and chromatin RNA fractions reveals that sequencing of purified factory RNA maps the complete nascent transcriptome; it is rich in intronic unprocessed transcript, as well as long intergenic non-coding (lincRNAs) and enhancer-associated RNAs (eRNAs), micro-RNA precursors and repeat-derived RNAs. Hence, we verify that transcription factories produce most nascent RNA and confer a regulatory role via their association with a set of specifically-retained non-coding transcripts.
authors.affiliation: Deutsches Krebsforschungszentrum (DKFZ) & BioQuant, D-69120 Heidelberg, Germany.
authors.forename: Maïwen
authors.initials: M
authors.lastname: Caudron-Herger
authors.affiliation: Sir William Dunn School of Pathology, University of Oxford, OX1 3RE Oxford, UK.
authors.forename: Peter R
authors.initials: PR
authors.lastname: Cook
authors.affiliation: Deutsches Krebsforschungszentrum (DKFZ) & BioQuant, D-69120 Heidelberg, Germany.
authors.forename: Karsten
authors.initials: K
authors.lastname: Rippe
authors.affiliation: Sir William Dunn School of Pathology, University of Oxford, OX1 3RE Oxford, UK Center for Molecular Medicine, University of Cologne, D-50931 Cologne, Germany argyris.papantonis@uni-koeln.de.
authors.forename: Argyris
authors.initials: A
authors.lastname: Papantonis
mesh_terms: D002478:Cells, Cultured; D002843:Chromatin / Q000378:metabolism; D020869:Gene Expression Profiling / Q000379:methods*; D006801:Humans; D007438:Introns; D012313:RNA / Q000302:isolation & purification; D012323:RNA Processing, Post-Transcriptional; D062085:RNA, Long Noncoding / Q000096:biosynthesis; D058727:RNA, Small Untranslated / Q000096:biosynthesis; D022661:RNA, Untranslated / Q000096:biosynthesis*; D017423:Sequence Analysis, RNA / Q000379:methods*; D059467:Transcriptome
chemical_list: D002843:Chromatin; D062085:RNA, Long Noncoding; D058727:RNA, Small Untranslated; D022661:RNA, Untranslated; D012313:RNA