title: Simultaneous processing and degradation of mitochondrial RNAs revealed by circularized RNA sequencing.
abstract: Mammalian mitochondrial RNAs are unique as they are derived from primary transcripts that encompass almost the entire mitochondrial genome. This necessitates extensive processing to release the individual mRNAs, rRNAs and tRNAs required for gene expression. Recent studies have revealed many of the proteins required for mitochondrial RNA processing, however the rapid turnover of precursor RNAs has made it impossible to analyze their composition and the hierarchy of processing. Here, we find that circularization of RNA prior to deep sequencing enables the discovery and characterization of unprocessed RNAs. Using this approach, we identify the most stable processing intermediates and the presence of intermediate processing products that are partially degraded and polyadenylated. Analysis of libraries constructed using RNA from mice lacking the nuclease subunit of the mitochondrial RNase P reveals the identities of stalled processing intermediates, their order of cleavage, and confirms the importance of RNase P in generating mature mitochondrial RNAs. Using RNA circularization prior to library preparation should provide a generally useful approach to studying RNA processing in many different biological systems.
authors.affiliation: Harry Perkins Institute of Medical Research and Centre for Medical Research, The University of Western Australia, Nedlands 6009, Australia.
authors.forename: Irina
authors.initials: I
authors.lastname: Kuznetsova
authors.affiliation: Harry Perkins Institute of Medical Research and Centre for Medical Research, The University of Western Australia, Nedlands 6009, Australia.
authors.forename: Stefan J
authors.initials: SJ
authors.lastname: Siira
authors.affiliation: Harry Perkins Institute of Medical Research and Centre for Medical Research, The University of Western Australia, Nedlands 6009, Australia.
authors.forename: Anne-Marie J
authors.initials: AJ
authors.lastname: Shearwood
authors.affiliation: Harry Perkins Institute of Medical Research and Centre for Medical Research, The University of Western Australia, Nedlands 6009, Australia.
authors.forename: Judith A
authors.initials: JA
authors.lastname: Ermer
authors.affiliation: Harry Perkins Institute of Medical Research and Centre for Medical Research, The University of Western Australia, Nedlands 6009, Australia.|School of Molecular Sciences, The University of Western Australia, Crawley 6009, Australia.
authors.forename: Aleksandra
authors.initials: A
authors.lastname: Filipovska
authors.affiliation: Harry Perkins Institute of Medical Research and Centre for Medical Research, The University of Western Australia, Nedlands 6009, Australia.|School of Molecular Sciences, The University of Western Australia, Crawley 6009, Australia.
authors.forename: Oliver
authors.initials: O
authors.lastname: Rackham
mesh_terms: D000818:Animals; D019295:Computational Biology; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D026723:Polyadenylation; D012313:RNA / Q000235:genetics / Q000378:metabolism*; D012323:RNA Processing, Post-Transcriptional; D020871:RNA Stability; D000079962:RNA, Circular; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D000077278:RNA, Mitochondrial; D043262:Ribonuclease P / Q000378:metabolism; D017423:Sequence Analysis, RNA / Q000379:methods*
chemical_list: D000079962:RNA, Circular; D012333:RNA, Messenger; D000077278:RNA, Mitochondrial; D012313:RNA; D043262:Ribonuclease P