Transcriptome-wide mapping reveals widespread dynamic-regulated pseudouridylation of ncRNA and mRNA

Schraga Schwartz; Douglas A Bernstein; Maxwell R Mumbach; Marko Jovanovic; Rebecca H Herbst; Brian X León-Ricardo; Jesse M Engreitz; Mitchell Guttman; Rahul Satija; Eric S Lander; Gerald Fink; Aviv Regev

doi:10.1016/j.cell.2014.08.028

Transcriptome-wide mapping reveals widespread dynamic-regulated pseudouridylation of ncRNA and mRNA

Cell. 2014 Sep 25;159(1):148-162. doi: 10.1016/j.cell.2014.08.028. Epub 2014 Sep 11.

Authors

Affiliations

¹ Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA.
² Whitehead Institute for Biomedical Research, Cambridge, MA 02142, USA.
³ Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA; Department of Systems Biology, Harvard Medical School, Boston, MA 02114, USA.
⁴ Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA; Department of Biology, University of Puerto Rico, Rio Piedras Campus, San Juan 00931, Puerto Rico.
⁵ Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA; Division of Health Sciences and Technology, MIT, Cambridge, MA 02139, USA.
⁶ Division of Biology and Biological Engineering, California Institute of Technology, Pasadena, CA 91125, USA.
⁷ Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA; Department of Systems Biology, Harvard Medical School, Boston, MA 02114, USA; Department of Biology, MIT, Cambridge, MA 02139, USA.
⁸ Whitehead Institute for Biomedical Research, Cambridge, MA 02142, USA; Department of Biology, MIT, Cambridge, MA 02139, USA. Electronic address: gfink@wi.mit.edu.
⁹ Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA; Department of Biology, MIT, Cambridge, MA 02139, USA; Howard Hughes Medical Institute, 4000 Jones Bridge Road, Chevy Chase, MD 20815, USA. Electronic address: aregev@broad.mit.edu.

Abstract

Pseudouridine is the most abundant RNA modification, yet except for a few well-studied cases, little is known about the modified positions and their function(s). Here, we develop Ψ-seq for transcriptome-wide quantitative mapping of pseudouridine. We validate Ψ-seq with spike-ins and de novo identification of previously reported positions and discover hundreds of unique sites in human and yeast mRNAs and snoRNAs. Perturbing pseudouridine synthases (PUS) uncovers which pseudouridine synthase modifies each site and their target sequence features. mRNA pseudouridinylation depends on both site-specific and snoRNA-guided pseudouridine synthases. Upon heat shock in yeast, Pus7p-mediated pseudouridylation is induced at >200 sites, and PUS7 deletion decreases the levels of otherwise pseudouridylated mRNA, suggesting a role in enhancing transcript stability. rRNA pseudouridine stoichiometries are conserved but reduced in cells from dyskeratosis congenita patients, where the PUS DKC1 is mutated. Our work identifies an enhanced, transcriptome-wide scope for pseudouridine and methods to dissect its underlying mechanisms and function.

Publication types

Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't

MeSH terms

Animals
Candida albicans / genetics
Candida albicans / metabolism
Cell Cycle Proteins / genetics
Cell Cycle Proteins / metabolism
Dyskeratosis Congenita / genetics
Dyskeratosis Congenita / metabolism
Gene Expression Profiling
Humans
Intramolecular Transferases / chemistry
Intramolecular Transferases / metabolism
Mice
Molecular Sequence Data
Nuclear Proteins / genetics
Nuclear Proteins / metabolism
Pseudouridine / analysis*
Pseudouridine / metabolism
RNA / chemistry
RNA / genetics
RNA, Messenger / chemistry*
RNA, Ribosomal / chemistry
RNA, Ribosomal / genetics
RNA, Untranslated / chemistry*
Saccharomyces cerevisiae / genetics
Saccharomyces cerevisiae / metabolism
Saccharomyces cerevisiae Proteins / chemistry
Saccharomyces cerevisiae Proteins / metabolism
Substrate Specificity
Telomerase / chemistry
Telomerase / genetics

Substances

Cell Cycle Proteins
DKC1 protein, human
Nuclear Proteins
Pus7 protein, S cerevisiae
RNA, Messenger
RNA, Ribosomal
RNA, Untranslated
Saccharomyces cerevisiae Proteins
telomerase RNA
Pseudouridine
RNA
Telomerase
Intramolecular Transferases
pseudouridine synthases

Associated data

GENBANK/GSE60047

Abstract

Publication types

MeSH terms

Substances

Associated data

Grants and funding