title: Two orthogonal cleavages separate subunit RNAs in mouse ribosome biogenesis.
abstract: Ribosome biogenesis is a dynamic multistep process, many features of which are still incompletely documented. Here, we show that changes in this pathway can be captured and annotated by means of a graphic set of pre-rRNA ratios, a technique we call Ratio Analysis of Multiple Precursors (RAMP). We find that knocking down a ribosome synthesis factor produces a characteristic RAMP profile that exhibits consistency across a range of depletion levels. This facilitates the inference of affected steps and simplifies comparative analysis. We applied RAMP to examine how endonucleolytic cleavages of the mouse pre-rRNA transcript in the internal transcribed spacer 1 (ITS1) are affected by depletion of factors required for maturation of the small ribosomal subunit (Rcl1, Fcf1/Utp24, Utp23) and the large subunit (Pes1, Nog1). The data suggest that completion of early maturation in a subunit triggers its release from the common pre-rRNA transcript by stimulating cleavage at the proximal site in ITS1. We also find that splitting of pre-rRNA in the 3' region of ITS1 is prevalent in adult mouse tissues and quiescent cells, as it is in human cells. We propose a model for subunit separation during mammalian ribosome synthesis and discuss its implications for understanding pre-rRNA processing pathways.
authors.affiliation: Department of Cell Biology, Rowan University School of Osteopathic Medicine, Stratford, NJ 08084, USA Graduate School of Biomedical Sciences, Rowan University, Stratford, NJ 08084, USA.
authors.forename: Minshi
authors.initials: M
authors.lastname: Wang
authors.affiliation: Department of Cell Biology, Rowan University School of Osteopathic Medicine, Stratford, NJ 08084, USA Graduate School of Biomedical Sciences, Rowan University, Stratford, NJ 08084, USA.
authors.forename: Leonid
authors.initials: L
authors.lastname: Anikin
authors.affiliation: Department of Cell Biology, Rowan University School of Osteopathic Medicine, Stratford, NJ 08084, USA pestovdg@rowan.edu.
authors.forename: Dimitri G
authors.identifier: http://orcid.org/0000-0002-1425-9879
authors.initials: DG
authors.lastname: Pestov
mesh_terms: D000818:Animals; D041702:BALB 3T3 Cells; D002478:Cells, Cultured; D021903:DNA, Ribosomal Spacer; D051379:Mice; D008957:Models, Genetic; D041681:NIH 3T3 Cells; D059367:RNA Cleavage; D034622:RNA Interference; D012322:RNA Precursors / Q000737:chemistry / Q000378:metabolism; D012323:RNA Processing, Post-Transcriptional; D012335:RNA, Ribosomal / Q000737:chemistry / Q000378:metabolism*; D012337:RNA, Ribosomal, 18S / Q000378:metabolism; D012269:Ribosomal Proteins / Q000037:antagonists & inhibitors; D054683:Ribosome Subunits, Large, Eukaryotic / Q000378:metabolism*; D054682:Ribosome Subunits, Small, Eukaryotic / Q000378:metabolism*
chemical_list: D021903:DNA, Ribosomal Spacer; D012322:RNA Precursors; D012335:RNA, Ribosomal; D012337:RNA, Ribosomal, 18S; D012269:Ribosomal Proteins