The role of BRCA2 in replication-coupled DNA interstrand cross-link repair in vitro

Lubos Cipak; Norifumi Watanabe; Tadayoshi Bessho

doi:10.1038/nsmb1120

The role of BRCA2 in replication-coupled DNA interstrand cross-link repair in vitro

Lubos Cipak, Norifumi Watanabe, Tadayoshi Bessho

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45 Scopus citations

Abstract

Using a defined substrate DNA with a single psoralen interstrand cross-link (ICL), we studied the molecular mechanism of human ICL repair. In vitro ICL repair by human extracts is dependent on replication and is a largely error-free process. Extracts from a human BRCA2-defective mutant cell line, CAPAN-1, are severely compromised in ICL repair. Specifically, 'unhooked' but not fully repaired products accumulate in the reaction with CAPAN-1, and transient expression of BRCA2 in CAPAN-1 restores repair activity. Together, these results reveal that BRCA2 participates in repair of replication-mediated double-strand breaks generated when replication forks encounter ICLs. We also show that nucleotide excision repair is essential for the removal of the lesion left behind on one strand after unhooking. This study provides new mechanistic insights into the repair of ICLs in human cells.

Original language	English (US)
Pages (from-to)	729-733
Number of pages	5
Journal	Nature Structural and Molecular Biology
Volume	13
Issue number	8
DOIs	https://doi.org/10.1038/nsmb1120
State	Published - Aug 2006

ASJC Scopus subject areas

Structural Biology
Molecular Biology

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title = "The role of BRCA2 in replication-coupled DNA interstrand cross-link repair in vitro",

abstract = "Using a defined substrate DNA with a single psoralen interstrand cross-link (ICL), we studied the molecular mechanism of human ICL repair. In vitro ICL repair by human extracts is dependent on replication and is a largely error-free process. Extracts from a human BRCA2-defective mutant cell line, CAPAN-1, are severely compromised in ICL repair. Specifically, 'unhooked' but not fully repaired products accumulate in the reaction with CAPAN-1, and transient expression of BRCA2 in CAPAN-1 restores repair activity. Together, these results reveal that BRCA2 participates in repair of replication-mediated double-strand breaks generated when replication forks encounter ICLs. We also show that nucleotide excision repair is essential for the removal of the lesion left behind on one strand after unhooking. This study provides new mechanistic insights into the repair of ICLs in human cells.",

author = "Lubos Cipak and Norifumi Watanabe and Tadayoshi Bessho",

note = "Funding Information: We thank J. Hearst for the psoralen-modified oligonucleotides, S. Powell (Washington University) for the HA-BRCA2 plasmid and J. Reardon, R. Lahue, R. Lewis and A. Tomkinson for critical reading of the manuscript. This work was supported by grant CA095291 from the US National Institutes of Health and grant DAMD17-03-1-0225 from the US Army Medical Research and Materiel Command.",

year = "2006",

month = aug,

doi = "10.1038/nsmb1120",

language = "English (US)",

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AU - Watanabe, Norifumi

AU - Bessho, Tadayoshi

N1 - Funding Information: We thank J. Hearst for the psoralen-modified oligonucleotides, S. Powell (Washington University) for the HA-BRCA2 plasmid and J. Reardon, R. Lahue, R. Lewis and A. Tomkinson for critical reading of the manuscript. This work was supported by grant CA095291 from the US National Institutes of Health and grant DAMD17-03-1-0225 from the US Army Medical Research and Materiel Command.

PY - 2006/8

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N2 - Using a defined substrate DNA with a single psoralen interstrand cross-link (ICL), we studied the molecular mechanism of human ICL repair. In vitro ICL repair by human extracts is dependent on replication and is a largely error-free process. Extracts from a human BRCA2-defective mutant cell line, CAPAN-1, are severely compromised in ICL repair. Specifically, 'unhooked' but not fully repaired products accumulate in the reaction with CAPAN-1, and transient expression of BRCA2 in CAPAN-1 restores repair activity. Together, these results reveal that BRCA2 participates in repair of replication-mediated double-strand breaks generated when replication forks encounter ICLs. We also show that nucleotide excision repair is essential for the removal of the lesion left behind on one strand after unhooking. This study provides new mechanistic insights into the repair of ICLs in human cells.

AB - Using a defined substrate DNA with a single psoralen interstrand cross-link (ICL), we studied the molecular mechanism of human ICL repair. In vitro ICL repair by human extracts is dependent on replication and is a largely error-free process. Extracts from a human BRCA2-defective mutant cell line, CAPAN-1, are severely compromised in ICL repair. Specifically, 'unhooked' but not fully repaired products accumulate in the reaction with CAPAN-1, and transient expression of BRCA2 in CAPAN-1 restores repair activity. Together, these results reveal that BRCA2 participates in repair of replication-mediated double-strand breaks generated when replication forks encounter ICLs. We also show that nucleotide excision repair is essential for the removal of the lesion left behind on one strand after unhooking. This study provides new mechanistic insights into the repair of ICLs in human cells.

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The role of BRCA2 in replication-coupled DNA interstrand cross-link repair in vitro

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