Assessment of anti-recombination and double-strand break-induced gene conversion in human cells by a chromosomal reporter

Keqian Xu; Xiling Wu; Joshua D Tompkins; Chengtao Her

doi:10.1074/jbc.M112.352302

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Assessment of anti-recombination and double-strand break-induced gene conversion in human cells by a chromosomal reporter

Journal article

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Assessment of anti-recombination and double-strand break-induced gene conversion in human cells by a chromosomal reporter

Keqian Xu, Xiling Wu, Joshua D Tompkins and Chengtao Her

The Journal of biological chemistry, Vol.287(35), pp.29543-29553

08/24/2012

DOI: https://doi.org/10.1074/jbc.M112.352302

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https://hdl.handle.net/2376/105338

PMCID: PMC3436158

PMID: 22773873

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https://doi.org/10.1074/jbc.M112.352302View

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Abstract

Cell Line

Gene Conversion - genetics

MutL Protein Homolog 1

Chromosomes, Human - genetics

Humans

Nuclear Proteins - metabolism

Biological Assay - methods

MutS Homolog 2 Protein - genetics

Chromosomes, Human - metabolism

DNA-Binding Proteins - genetics

MutS Homolog 2 Protein - metabolism

DNA Breaks, Double-Stranded

DNA-Binding Proteins - metabolism

MRE11 Homologue Protein

Adaptor Proteins, Signal Transducing - genetics

Adaptor Proteins, Signal Transducing - metabolism

Nuclear Proteins - genetics

Genes, Reporter

Gene conversion is one of the frequent end results of homologous recombination, and it often underlies the inactivation of tumor suppressor genes in cancer cells. Here, we have developed an integrated assay system that allows simultaneous examination of double-strand break (DSB)-induced gene conversion events at the site of a DSB (proximal region) and at a surrounding region ~1 kb away from the break (distal region). Utilizing this assay system, we find that gene conversion events at the proximal and distal regions are relatively independent of one another. The results also indicate that synthesis-dependent strand annealing (SDSA) plays a major role in DSB-induced gene conversion. In addition, our current study has demonstrated that hMLH1 plays an essential role in anti-recombination and gene conversion. Specifically, the anti-recombination activity of hMLH1 is partially dependent on its interaction with hMRE11. Our data suggests that the role of hMLH1 and hMRE11 in the process of gene conversion is complex, and these proteins play different roles in DSB-induced proximal and distal gene conversions. In particular, the involvement of hMLH1 and hMRE11 in the distal gene conversion requires both hMSH2 and heteroduplex formation.

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Details

Title: Assessment of anti-recombination and double-strand break-induced gene conversion in human cells by a chromosomal reporter
Creators: Keqian Xu - From the School of Molecular Biosciences, Washington State University, Pullman, Washington 99164-7520
Xiling Wu
Joshua D Tompkins
Chengtao Her
Publication Details: The Journal of biological chemistry, Vol.287(35), pp.29543-29553
Academic Unit: Molecular Biosciences, School of
Publisher: United States
Grant note: R01 GM084353 / NIGMS NIH HHS GM084353 / NIGMS NIH HHS
Identifiers: 99900546759601842
Language: English
Resource Type: Journal article