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Title: Transcriptome-Wide Cleavage Site Mapping on Cellular mRNAs Reveals Features Underlying Sequence-Specific Cleavage by the Viral Ribonuclease SOX

Abstract

Many viruses express factors that reduce host gene expression through widespread degradation of cellular mRNA. An example of this class of proteins is the mRNA-targeting endoribonuclease SOX from the gamma-herpesvirus Kaposi’s sarcoma-associated herpesvirus (KSHV). Previous studies indicated that cleavage of messenger RNAs (mRNA) by SOX occurs at specific locations defined by the sequence of the target RNA, which is at odds with the down-regulation of a large portion of cellular transcripts. In this study, we address this paradox by using high-throughput sequencing of cleavage intermediates combined with a custom bioinformatics-based analysis pipeline to identify SOX cleavage sites across the mRNA transcriptome. These data, coupled with targeted mutagenesis, reveal that while cleavage sites are specific and reproducible, they are defined by a degenerate sequence motif containing a small number of conserved residues rather than a strong consensus sequence. This degenerate element is well represented in both human and KSHV mRNA, and its presence correlates with RNA destabilization by SOX. This represents a new endonuclease targeting strategy, in which use of a degenerate targeting element enables RNA cleavage at specific locations without restricting the range of targets. Furthermore, it shows that strong target selectivity can be achieved without a high degree ofmore » sequence specificity.« less

Authors:
 [1];  [2];  [3]
+ Show Author Affiliations
  1. Tufts Univ., Medford, MA (United States); Tufts Univ. School of Medicine, Boston, MA (United States)
  2. Harvard Univ., Cambridge, MA (United States); Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States)
  3. Univ. of California, Berkeley, CA (United States). Howard Hughes Medical Inst.; Univ. of California, Berkeley, CA (United States). Dept. of Plant and Microbial Biology
Publication Date:
Research Org.:
Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States)
Sponsoring Org.:
USDOE Office of Science (SC); National Institutes of Health (NIH)
OSTI Identifier:
1524004
Grant/Contract Number:  
AC02-05CH11231; NIH CA136367; NIH CA160556
Resource Type:
Accepted Manuscript
Journal Name:
PLoS Pathogens
Additional Journal Information:
Journal Volume: 11; Journal Issue: 12; Journal ID: ISSN 1553-7374
Publisher:
Public Library of Science
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES; 60 APPLIED LIFE SCIENCES

Citation Formats

Gaglia, Marta Maria, Rycroft, Chris H., and Glaunsinger, Britt A. Transcriptome-Wide Cleavage Site Mapping on Cellular mRNAs Reveals Features Underlying Sequence-Specific Cleavage by the Viral Ribonuclease SOX. United States: N. p., 2015. Web. doi:10.1371/journal.ppat.1005305.
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Gaglia, Marta Maria, Rycroft, Chris H., & Glaunsinger, Britt A. Transcriptome-Wide Cleavage Site Mapping on Cellular mRNAs Reveals Features Underlying Sequence-Specific Cleavage by the Viral Ribonuclease SOX. United States. https://doi.org/10.1371/journal.ppat.1005305
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Gaglia, Marta Maria, Rycroft, Chris H., and Glaunsinger, Britt A. Tue . "Transcriptome-Wide Cleavage Site Mapping on Cellular mRNAs Reveals Features Underlying Sequence-Specific Cleavage by the Viral Ribonuclease SOX". United States. https://doi.org/10.1371/journal.ppat.1005305. https://www.osti.gov/servlets/purl/1524004.
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@article{osti_1524004,
title = {Transcriptome-Wide Cleavage Site Mapping on Cellular mRNAs Reveals Features Underlying Sequence-Specific Cleavage by the Viral Ribonuclease SOX},
author = {Gaglia, Marta Maria and Rycroft, Chris H. and Glaunsinger, Britt A.},
abstractNote = {Many viruses express factors that reduce host gene expression through widespread degradation of cellular mRNA. An example of this class of proteins is the mRNA-targeting endoribonuclease SOX from the gamma-herpesvirus Kaposi’s sarcoma-associated herpesvirus (KSHV). Previous studies indicated that cleavage of messenger RNAs (mRNA) by SOX occurs at specific locations defined by the sequence of the target RNA, which is at odds with the down-regulation of a large portion of cellular transcripts. In this study, we address this paradox by using high-throughput sequencing of cleavage intermediates combined with a custom bioinformatics-based analysis pipeline to identify SOX cleavage sites across the mRNA transcriptome. These data, coupled with targeted mutagenesis, reveal that while cleavage sites are specific and reproducible, they are defined by a degenerate sequence motif containing a small number of conserved residues rather than a strong consensus sequence. This degenerate element is well represented in both human and KSHV mRNA, and its presence correlates with RNA destabilization by SOX. This represents a new endonuclease targeting strategy, in which use of a degenerate targeting element enables RNA cleavage at specific locations without restricting the range of targets. Furthermore, it shows that strong target selectivity can be achieved without a high degree of sequence specificity.},
doi = {10.1371/journal.ppat.1005305},
journal = {PLoS Pathogens},
number = 12,
volume = 11,
place = {United States},
year = {2015},
month = {12}
}
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Journal Article:
Free Publicly Available Full Text
Publisher's Version of Record
https://doi.org/10.1371/journal.ppat.1005305
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Cited by: 20 works
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Figures / Tables:

Fig 1 Fig 1: PARE experimental procedure and peak finding analysis pipeline. A) Diagram of the PARE procedure. B) Schematic of PyDegradome analysis approach, which uses read counts in a control sample to generate a table of thresholds to compare test sample counts to. The table lists thresholds for a particular user-definedmore » confidence level and for a range of ratios between control and test samples. The applicable ratio for each position is computed by multiplying a user-defined multiplicative factor by the ratio of total read counts for the exon in test vs. control samples, thus accounting for variation in RNA levels and total mapped reads. Read counts in test sample at each position must exceed the threshold to be identified as part of a peak. C) Example of plot of read counts (5’ end only) for 200 nt surrounding a SOX cut site identified by PyDegradome within the 3’–most exon of the LIMD1 NM_014240 transcript in the four samples. Note that y-axis has a logarithmic scale. This example shows the expected distribution for a cut site followed by exonucleolytic degradation due to residual Xrn1 activity or to the action of another nuclease, with the highest read count at cut site and decaying counts at following positions. Positions referred to as “peak” and “cut site” in the text are marked on the graph. D) Flow chart of steps used to defined SOX cut sites used for further analyses.« less
All figures and tables (6 total)
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Works referenced in this record:

Methods for analyzing deep sequencing expression data: constructing the human and mouse promoterome with deepCAGE data
journal, January 2009

An Overlapping Protein-Coding Region in Influenza A Virus Segment 3 Modulates the Host Response
journal, June 2012

Coordinated Destruction of Cellular Messages in Translation Complexes by the Gammaherpesvirus Host Shutoff Factor and the Mammalian Exonuclease Xrn1
journal, October 2011

A Ribonucleoprotein Complex Protects the Interleukin-6 mRNA from Degradation by Distinct Herpesviral Endonucleases
journal, May 2015

RNase L Targets Distinct Sites in Influenza A Virus RNAs
journal, December 2014

  • Cooper, Daphne A.; Banerjee, Shuvojit; Chakrabarti, Arindam
  • Journal of Virology, Vol. 89, Issue 5
  • DOI: 10.1128/JVI.02953-14

Cutadapt removes adapter sequences from high-throughput sequencing reads
journal, May 2011

SARS Coronavirus nsp1 Protein Induces Template-Dependent Endonucleolytic Cleavage of mRNAs: Viral mRNAs Are Resistant to nsp1-Induced RNA Cleavage
journal, December 2011

Host shutoff during productive Epstein-Barr virus infection is mediated by BGLF5 and may contribute to immune evasion
journal, February 2007

  • Rowe, M.; Glaunsinger, B.; van Leeuwen, D.
  • Proceedings of the National Academy of Sciences, Vol. 104, Issue 9
  • DOI: 10.1073/pnas.0611128104

Gammaherpesviral Gene Expression and Virion Composition Are Broadly Controlled by Accelerated mRNA Degradation
journal, January 2014

SMG6 promotes endonucleolytic cleavage of nonsense mRNA in human cells
journal, December 2008

  • Eberle, Andrea B.; Lykke-Andersen, Søren; Mühlemann, Oliver
  • Nature Structural & Molecular Biology, Vol. 16, Issue 1
  • DOI: 10.1038/nsmb.1530

Ribonuclease L and metal-ion–independent endoribonuclease cleavage sites in host and viral RNAs
journal, February 2014

  • Cooper, Daphne A.; Jha, Babal K.; Silverman, Robert H.
  • Nucleic Acids Research, Vol. 42, Issue 8
  • DOI: 10.1093/nar/gku118

Crystal structure of a KSHV–SOX–DNA complex: insights into the molecular mechanisms underlying DNase activity and host shutoff
journal, March 2011

  • Bagnéris, Claire; Briggs, Louise C.; Savva, Renos
  • Nucleic Acids Research, Vol. 39, Issue 13
  • DOI: 10.1093/nar/gkr111

TopHat2: accurate alignment of transcriptomes in the presence of insertions, deletions and gene fusions
journal, January 2013

Global or local? Predicting secondary structure and accessibility in mRNAs
journal, February 2012

  • Lange, Sita J.; Maticzka, Daniel; Möhl, Mathias
  • Nucleic Acids Research, Vol. 40, Issue 12
  • DOI: 10.1093/nar/gks181

Viral Nucleases Induce an mRNA Degradation-Transcription Feedback Loop in Mammalian Cells
journal, August 2015

ViennaRNA Package 2.0
journal, November 2011

  • Lorenz, Ronny; Bernhart, Stephan H.; Höner zu Siederdissen, Christian
  • Algorithms for Molecular Biology, Vol. 6, Issue 1
  • DOI: 10.1186/1748-7188-6-26

KSHV and the pathogenesis of Kaposi sarcoma: listening to human biology and medicine
journal, April 2010

  • Ganem, Don
  • Journal of Clinical Investigation, Vol. 120, Issue 4
  • DOI: 10.1172/JCI40567

Bacterial RNA motif in the 5' UTR of rpsF interacts with an S6:S18 complex
journal, December 2013

Construction of Parallel Analysis of RNA Ends (PARE) libraries for the study of cleaved miRNA targets and the RNA degradome
journal, February 2009

  • German, Marcelo A.; Luo, Shujun; Schroth, Gary
  • Nature Protocols, Vol. 4, Issue 3
  • DOI: 10.1038/nprot.2009.8

Human nonsense-mediated RNA decay initiates widely by endonucleolysis and targets snoRNA host genes
journal, November 2014

  • Lykke-Andersen, Søren; Chen, Yun; Ardal, Britt R.
  • Genes & Development, Vol. 28, Issue 22
  • DOI: 10.1101/gad.246538.114

The UL41 protein of herpes simplex virus mediates selective stabilization or degradation of cellular mRNAs
journal, December 2004

  • Esclatine, A.; Taddeo, B.; Roizman, B.
  • Proceedings of the National Academy of Sciences, Vol. 101, Issue 52
  • DOI: 10.1073/pnas.0408272102

Decay of Endoplasmic Reticulum-Localized mRNAs During the Unfolded Protein Response
journal, July 2006

A two-pronged strategy to suppress host protein synthesis by SARS coronavirus Nsp1 protein
journal, October 2009

  • Kamitani, Wataru; Huang, Cheng; Narayanan, Krishna
  • Nature Structural & Molecular Biology, Vol. 16, Issue 11
  • DOI: 10.1038/nsmb.1680

Characterization of PA-N terminal domain of Influenza A polymerase reveals sequence specific RNA cleavage
journal, July 2013

  • Datta, Kausiki; Wolkerstorfer, Andrea; Szolar, Oliver H. J.
  • Nucleic Acids Research, Vol. 41, Issue 17
  • DOI: 10.1093/nar/gkt603

The herpes simplex virus 1 UL41 gene-dependent destabilization of cellular RNAs is selective and may be sequence-specific
journal, March 2004

  • Esclatine, A.; Taddeo, B.; Evans, L.
  • Proceedings of the National Academy of Sciences, Vol. 101, Issue 10
  • DOI: 10.1073/pnas.0400354101

Lytic KSHV Infection Inhibits Host Gene Expression by Accelerating Global mRNA Turnover
journal, March 2004

Host Shutoff Is a Conserved Phenotype of Gammaherpesvirus Infection and Is Orchestrated Exclusively from the Cytoplasm
journal, July 2009

  • Covarrubias, S.; Richner, J. M.; Clyde, K.
  • Journal of Virology, Vol. 83, Issue 18
  • DOI: 10.1128/JVI.01051-09

Highly Selective Escape from KSHV-mediated Host mRNA Shutoff and Its Implications for Viral Pathogenesis
journal, August 2004

  • Glaunsinger, Britt; Ganem, Don
  • The Journal of Experimental Medicine, Vol. 200, Issue 3
  • DOI: 10.1084/jem.20031881

Host Transcript Accumulation during Lytic KSHV Infection Reveals Several Classes of Host Responses
journal, August 2007

Zipf’s Law in Gene Expression
journal, February 2003

An RNA Element in Human Interleukin 6 Confers Escape from Degradation by the Gammaherpesvirus SOX Protein
journal, February 2013

  • Hutin, S.; Lee, Y.; Glaunsinger, B. A.
  • Journal of Virology, Vol. 87, Issue 8
  • DOI: 10.1128/JVI.00159-13

Rapid construction of parallel analysis of RNA end (PARE) libraries for Illumina sequencing
journal, May 2014

A Common Strategy for Host RNA Degradation by Divergent Viruses
journal, June 2012

  • Gaglia, M. M.; Covarrubias, S.; Wong, W.
  • Journal of Virology, Vol. 86, Issue 17
  • DOI: 10.1128/JVI.01230-12

NIH Image to ImageJ: 25 years of image analysis
journal, June 2012

  • Schneider, Caroline A.; Rasband, Wayne S.; Eliceiri, Kevin W.
  • Nature Methods, Vol. 9, Issue 7
  • DOI: 10.1038/nmeth.2089

Diverse Endonucleolytic Cleavage Sites in the Mammalian Transcriptome Depend upon MicroRNAs, Drosha, and Additional Nucleases
journal, June 2010

WebLogo: A Sequence Logo Generator
journal, May 2004

  • Crooks, Gavin E.; Hon, Gary; Chandonia, John-Marc
  • Genome Research, Vol. 14, Issue 6, p. 1188-1190
  • DOI: 10.1101/gr.849004

Identification of SMG6 cleavage sites and a preferred RNA cleavage motif by global analysis of endogenous NMD targets in human cells
journal, November 2014

  • Schmidt, Skye A.; Foley, Patricia L.; Jeong, Dong-Hoon
  • Nucleic Acids Research, Vol. 43, Issue 1
  • DOI: 10.1093/nar/gku1258

The Herpes Simplex Virus vhs Protein Induces Endoribonucleolytic Cleavage of Target RNAs in Cell Extracts
journal, September 1999

Integration of PCR Fragments at Any Specific Site within Cloning Vectors without the Use of Restriction Enzymes and DNA Ligase
journal, July 2001

  • Geiser, Martin; Cébe, Régis; Drewello, Delia
  • BioTechniques, Vol. 31, Issue 1
  • DOI: 10.2144/01311st05

AGO-bound mature miRNAs are oligouridylated by TUTs and subsequently degraded by DIS3L2
journal, June 2020

TopHat2: accurate alignment of transcriptomes in the presence of insertions, deletions and gene fusions
text, January 2013

Zipf's Law in Gene Expression
text, January 2002

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    Works referencing / citing this record:

    Site specific target binding controls RNA cleavage efficiency by the Kaposi's sarcoma-associated herpesvirus endonuclease SOX
    journal, October 2018

    • Mendez, Aaron S.; Vogt, Carolin; Bohne, Jens
    • Nucleic Acids Research, Vol. 46, Issue 22
    • DOI: 10.1093/nar/gky932

    Translational Control in Virus-Infected Cells
    journal, June 2018

    • Stern-Ginossar, Noam; Thompson, Sunnie R.; Mathews, Michael B.
    • Cold Spring Harbor Perspectives in Biology, Vol. 11, Issue 3
    • DOI: 10.1101/cshperspect.a033001

    C19ORF66 Broadly Escapes Virus-Induced Endonuclease Cleavage and Restricts Kaposi’s Sarcoma-Associated Herpesvirus
    journal, April 2019

    • Rodriguez, William; Srivastav, Kumaraman; Muller, Mandy
    • Journal of Virology, Vol. 93, Issue 12
    • DOI: 10.1128/jvi.00373-19

    Selective Degradation of Host RNA Polymerase II Transcripts by Influenza A Virus PA-X Host Shutoff Protein
    journal, February 2016

    Nuclease escape elements protect messenger RNA against cleavage by multiple viral endonucleases
    journal, August 2017

    Shutoff of Host Gene Expression in Influenza A Virus and Herpesviruses: Similar Mechanisms and Common Themes
    journal, April 2016

    • Rivas, Hembly; Schmaling, Summer; Gaglia, Marta
    • Viruses, Vol. 8, Issue 4
    • DOI: 10.3390/v8040102
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