Visualizing locus-specific sister chromatid exchange reveals differential patterns of replication stress-induced fragile site breakage

Irina Waisertreiger, Katherine Popovich, Maya Block, Krista R. Anderson, Jacqueline H. Barlow

Research output: Contribution to journalArticle

Abstract

Chromosomal fragile sites are genomic loci sensitive to replication stress which accumulate high levels of DNA damage, and are frequently mutated in cancers. Fragile site damage is thought to arise from the aberrant repair of spontaneous replication stress, however successful fragile site repair cannot be calculated using existing techniques. Here, we report a new assay measuring recombination-mediated repair at endogenous genomic loci by combining a sister chromatid exchange (SCE) assay with fluorescent in situ hybridization (SCE-FISH). Using SCE-FISH, we find that endogenous and exogenous replication stress generated unrepaired breaks and SCEs at fragile sites. We also find that distinct sources of replication stress induce distinct patterns of breakage: ATR inhibition induces more breaks at early replicating fragile sites (ERFS), while ERFS and late-replicating common fragile sites (CFS) are equally fragile in response to aphidicolin. Furthermore, SCEs were suppressed at fragile sites near centromeres in response to replication stress, suggesting that genomic location influences DNA repair pathway choice. SCE-FISH also measured successful recombination in human primary lymphocytes, and identificed the proto-oncogene BCL2 as a replication stress-induced fragile site. These findings demonstrate that SCE-FISH frequency at fragile sites is a sensitive indicator of replication stress, and that large-scale genome organization influences DNA repair pathway choice.

Original languageEnglish (US)
JournalOncogene
DOIs
StateAccepted/In press - Jan 1 2019

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Sister Chromatid Exchange
DNA Repair
Aphidicolin
Recombinational DNA Repair
Proto-Oncogenes
Centromere
Fluorescence In Situ Hybridization
Genetic Recombination
DNA Damage
Genome
Lymphocytes
Neoplasms

ASJC Scopus subject areas

  • Molecular Biology
  • Genetics
  • Cancer Research

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Visualizing locus-specific sister chromatid exchange reveals differential patterns of replication stress-induced fragile site breakage. / Waisertreiger, Irina; Popovich, Katherine; Block, Maya; Anderson, Krista R.; Barlow, Jacqueline H.

In: Oncogene, 01.01.2019.

Research output: Contribution to journalArticle

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abstract = "Chromosomal fragile sites are genomic loci sensitive to replication stress which accumulate high levels of DNA damage, and are frequently mutated in cancers. Fragile site damage is thought to arise from the aberrant repair of spontaneous replication stress, however successful fragile site repair cannot be calculated using existing techniques. Here, we report a new assay measuring recombination-mediated repair at endogenous genomic loci by combining a sister chromatid exchange (SCE) assay with fluorescent in situ hybridization (SCE-FISH). Using SCE-FISH, we find that endogenous and exogenous replication stress generated unrepaired breaks and SCEs at fragile sites. We also find that distinct sources of replication stress induce distinct patterns of breakage: ATR inhibition induces more breaks at early replicating fragile sites (ERFS), while ERFS and late-replicating common fragile sites (CFS) are equally fragile in response to aphidicolin. Furthermore, SCEs were suppressed at fragile sites near centromeres in response to replication stress, suggesting that genomic location influences DNA repair pathway choice. SCE-FISH also measured successful recombination in human primary lymphocytes, and identificed the proto-oncogene BCL2 as a replication stress-induced fragile site. These findings demonstrate that SCE-FISH frequency at fragile sites is a sensitive indicator of replication stress, and that large-scale genome organization influences DNA repair pathway choice.",
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