Nucleotide-resolution DNA double-strand break mapping by next-generation sequencing

Nicola Crosetto; Abhishek Mitra; Maria Joao Silva; Magda Bienko; Norbert Dojer; Qi Wang; Elif Karaca; Roberto Chiarle; Magdalena Skrzypczak; Krzysztof Ginalski; Philippe Pasero; Maga Rowicka; Ivan Dikic

doi:10.1038/nmeth.2408

Nucleotide-resolution DNA double-strand break mapping by next-generation sequencing

Nicola Crosetto
, Abhishek Mitra
, Maria Joao Silva
, Magda Bienko
, Norbert Dojer
, Qi Wang
, Elif Karaca
, Roberto Chiarle
, Magdalena Skrzypczak
, Krzysztof Ginalski
, Philippe Pasero
, Maga Rowicka
, Ivan Dikic

Biochemistry & Molecular Biology

Research output: Contribution to journal › Article › peer-review

394 Scopus citations

Abstract

We present a genome-wide approach to map DNA double-strand breaks (DSBs) at nucleotide resolution by a method we termed BLESS (direct in situ breaks labeling, enrichment on streptavidin and next-generation sequencing). We validated and tested BLESS using human and mouse cells and different DSBs-inducing agents and sequencing platforms. BLESS was able to detect telomere ends, Sce endonuclease-induced DSBs and complex genome-wide DSB landscapes. As a proof of principle, we characterized the genomic landscape of sensitivity to replication stress in human cells, and we identified >2,000 nonuniformly distributed aphidicolin-sensitive regions (ASRs) overrepresented in genes and enriched in satellite repeats. ASRs were also enriched in regions rearranged in human cancers, with many cancer-associated genes exhibiting high sensitivity to replication stress. Our method is suitable for genome-wide mapping of DSBs in various cells and experimental conditions, with a specificity and resolution unachievable by current techniques.

Original language	English (US)
Pages (from-to)	361-365
Number of pages	5
Journal	Nature Methods
Volume	10
Issue number	4
DOIs	https://doi.org/10.1038/nmeth.2408
State	Published - Apr 2013

ASJC Scopus subject areas

Biotechnology
Biochemistry
Molecular Biology
Cell Biology

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10.1038/nmeth.2408

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@article{40776c2a00874453a827b82908407f64,

title = "Nucleotide-resolution DNA double-strand break mapping by next-generation sequencing",

abstract = "We present a genome-wide approach to map DNA double-strand breaks (DSBs) at nucleotide resolution by a method we termed BLESS (direct in situ breaks labeling, enrichment on streptavidin and next-generation sequencing). We validated and tested BLESS using human and mouse cells and different DSBs-inducing agents and sequencing platforms. BLESS was able to detect telomere ends, Sce endonuclease-induced DSBs and complex genome-wide DSB landscapes. As a proof of principle, we characterized the genomic landscape of sensitivity to replication stress in human cells, and we identified >2,000 nonuniformly distributed aphidicolin-sensitive regions (ASRs) overrepresented in genes and enriched in satellite repeats. ASRs were also enriched in regions rearranged in human cancers, with many cancer-associated genes exhibiting high sensitivity to replication stress. Our method is suitable for genome-wide mapping of DSBs in various cells and experimental conditions, with a specificity and resolution unachievable by current techniques.",

author = "Nicola Crosetto and Abhishek Mitra and Silva, \{Maria Joao\} and Magda Bienko and Norbert Dojer and Qi Wang and Elif Karaca and Roberto Chiarle and Magdalena Skrzypczak and Krzysztof Ginalski and Philippe Pasero and Maga Rowicka and Ivan Dikic",

note = "Funding Information: We acknowledge Y. Shiloh (Tel Aviv University) and A.J. Pierce (University of Kentucky) for kindly providing U2OS\_DRH-1 cells and I-Sce plasmids. We are grateful to T. W{\l}odarski, A.R. Lehmann, G. Fudenberg and A. Kudlicki for insightful discussions, critical reading of the manuscript and help with data analysis. This work was supported by grants from Deutsche Forschungsgemeinschaft, the Cluster of Excellence “Macromolecular Complexes” of the Goethe University Frankfurt (EXC115), the LOEWE-funded Oncogenic Signaling Frankfurt network, the LOEWE Gene and Cell Therapy Center and the European Research Council (ERC) under the European Union{\textquoteright}s Seventh Framework Programme (FP7/2007-2013) and ERC grant agreement 250241-LineUb to I.D.; from the Associazione Italiana per la Ricerca sul cancro (AIRC) and the International Association for Cancer Research (AICR) and grant FP7 ERC-2009-StG (proposal 242965–“Lunely”) to R.C.; from the Foundation for Polish Science (TEAM), Polish National Science Centre (2011/02/A/NZ2/00014) and European Regional Development Fund under the Innovative Economy Programme (POIG.02.02.00-14-024/08-00) to K.G.; from Ligue contre le Cancer ({\'e}quipe labellis{\'e}e), Agence Nationale de la Recherche (RepliCare) and Institut National du Cancer to P.P.; and by grant UL1TR000071 ITS “Novel Methods” from the National Center for Research Resources, US National Institutes of Health, to M.R. M.B. is a recipient of a Human Frontier Science Program Long-Term Fellowship.",

year = "2013",

month = apr,

doi = "10.1038/nmeth.2408",

language = "English (US)",

volume = "10",

pages = "361--365",

journal = "Nature Methods",

issn = "1548-7091",

publisher = "Public Library of Science",

number = "4",

}

TY - JOUR

T1 - Nucleotide-resolution DNA double-strand break mapping by next-generation sequencing

AU - Crosetto, Nicola

AU - Mitra, Abhishek

AU - Silva, Maria Joao

AU - Bienko, Magda

AU - Dojer, Norbert

AU - Wang, Qi

AU - Karaca, Elif

AU - Chiarle, Roberto

AU - Skrzypczak, Magdalena

AU - Ginalski, Krzysztof

AU - Pasero, Philippe

AU - Rowicka, Maga

AU - Dikic, Ivan

N1 - Funding Information: We acknowledge Y. Shiloh (Tel Aviv University) and A.J. Pierce (University of Kentucky) for kindly providing U2OS_DRH-1 cells and I-Sce plasmids. We are grateful to T. Włodarski, A.R. Lehmann, G. Fudenberg and A. Kudlicki for insightful discussions, critical reading of the manuscript and help with data analysis. This work was supported by grants from Deutsche Forschungsgemeinschaft, the Cluster of Excellence “Macromolecular Complexes” of the Goethe University Frankfurt (EXC115), the LOEWE-funded Oncogenic Signaling Frankfurt network, the LOEWE Gene and Cell Therapy Center and the European Research Council (ERC) under the European Union’s Seventh Framework Programme (FP7/2007-2013) and ERC grant agreement 250241-LineUb to I.D.; from the Associazione Italiana per la Ricerca sul cancro (AIRC) and the International Association for Cancer Research (AICR) and grant FP7 ERC-2009-StG (proposal 242965–“Lunely”) to R.C.; from the Foundation for Polish Science (TEAM), Polish National Science Centre (2011/02/A/NZ2/00014) and European Regional Development Fund under the Innovative Economy Programme (POIG.02.02.00-14-024/08-00) to K.G.; from Ligue contre le Cancer (équipe labellisée), Agence Nationale de la Recherche (RepliCare) and Institut National du Cancer to P.P.; and by grant UL1TR000071 ITS “Novel Methods” from the National Center for Research Resources, US National Institutes of Health, to M.R. M.B. is a recipient of a Human Frontier Science Program Long-Term Fellowship.

PY - 2013/4

Y1 - 2013/4

N2 - We present a genome-wide approach to map DNA double-strand breaks (DSBs) at nucleotide resolution by a method we termed BLESS (direct in situ breaks labeling, enrichment on streptavidin and next-generation sequencing). We validated and tested BLESS using human and mouse cells and different DSBs-inducing agents and sequencing platforms. BLESS was able to detect telomere ends, Sce endonuclease-induced DSBs and complex genome-wide DSB landscapes. As a proof of principle, we characterized the genomic landscape of sensitivity to replication stress in human cells, and we identified >2,000 nonuniformly distributed aphidicolin-sensitive regions (ASRs) overrepresented in genes and enriched in satellite repeats. ASRs were also enriched in regions rearranged in human cancers, with many cancer-associated genes exhibiting high sensitivity to replication stress. Our method is suitable for genome-wide mapping of DSBs in various cells and experimental conditions, with a specificity and resolution unachievable by current techniques.

AB - We present a genome-wide approach to map DNA double-strand breaks (DSBs) at nucleotide resolution by a method we termed BLESS (direct in situ breaks labeling, enrichment on streptavidin and next-generation sequencing). We validated and tested BLESS using human and mouse cells and different DSBs-inducing agents and sequencing platforms. BLESS was able to detect telomere ends, Sce endonuclease-induced DSBs and complex genome-wide DSB landscapes. As a proof of principle, we characterized the genomic landscape of sensitivity to replication stress in human cells, and we identified >2,000 nonuniformly distributed aphidicolin-sensitive regions (ASRs) overrepresented in genes and enriched in satellite repeats. ASRs were also enriched in regions rearranged in human cancers, with many cancer-associated genes exhibiting high sensitivity to replication stress. Our method is suitable for genome-wide mapping of DSBs in various cells and experimental conditions, with a specificity and resolution unachievable by current techniques.

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UR - https://www.scopus.com/pages/publications/84875754465#tab=citedBy

U2 - 10.1038/nmeth.2408

DO - 10.1038/nmeth.2408

M3 - Article

C2 - 23503052

AN - SCOPUS:84875754465

SN - 1548-7091

VL - 10

SP - 361

EP - 365

JO - Nature Methods

JF - Nature Methods

IS - 4

ER -

Nucleotide-resolution DNA double-strand break mapping by next-generation sequencing

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