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.
authors.affiliation: Institute of Biochemistry II, Goethe University Medical School, Frankfurt, Germany. crosetto@mit.edu
authors.forename: Nicola
authors.initials: N
authors.lastname: Crosetto
authors.forename: Abhishek
authors.initials: A
authors.lastname: Mitra
authors.forename: Maria Joao
authors.initials: MJ
authors.lastname: Silva
authors.forename: Magda
authors.initials: M
authors.lastname: Bienko
authors.forename: Norbert
authors.initials: N
authors.lastname: Dojer
authors.forename: Qi
authors.initials: Q
authors.lastname: Wang
authors.forename: Elif
authors.initials: E
authors.lastname: Karaca
authors.forename: Roberto
authors.initials: R
authors.lastname: Chiarle
authors.forename: Magdalena
authors.initials: M
authors.lastname: Skrzypczak
authors.forename: Krzysztof
authors.initials: K
authors.lastname: Ginalski
authors.forename: Philippe
authors.initials: P
authors.lastname: Pasero
authors.forename: Maga
authors.initials: M
authors.lastname: Rowicka
authors.forename: Ivan
authors.initials: I
authors.lastname: Dikic
mesh_terms: D000818:Animals; D016590:Aphidicolin; D001483:Base Sequence; D045744:Cell Line, Tumor; D003001:Cloning, Molecular; D053903:DNA Breaks, Double-Stranded; D004261:DNA Replication; D005347:Fibroblasts / Q000378:metabolism; D023281:Genomics / Q000379:methods*; D006801:Humans; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D018895:Microsatellite Repeats; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*; D020161:Physical Chromosome Mapping / Q000379:methods; D017422:Sequence Analysis, DNA; D013154:Spleen; D013737:Testis; D014779:Virus Replication
chemical_list: D016590:Aphidicolin