Gene conversion and deletion frequencies during double-strand break repair in human cells are controlled by the distance between direct repeats

Schildkraut, Ezra; Miller, Cheryl A.; Nickoloff, Jac A.

doi:10.1093/nar/gki295

Cited by 40 publications

(41 citation statements)

References 46 publications

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“…Many of these higher order parallelisms involve sites in tandem duplicate clusters, a structural confirmation associated with frequent, recurrent non-allelic homologous recombination [26, 27, 52–54]. In particular, 33.4 % of higher order parallelisms involve ≥3 SNPs within 5 Mb of each other (Additional file 7: Table S6).…”

Section: Resultsmentioning

confidence: 99%

Interlocus gene conversion explains at least 2.7 % of single nucleotide variants in human segmental duplications

Dumont

2015

BMC Genomics

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BackgroundInterlocus gene conversion (IGC) is a recombination-based mechanism that results in the unidirectional transfer of short stretches of sequence between paralogous loci. Although IGC is a well-established mechanism of human disease, the extent to which this mutagenic process has shaped overall patterns of segregating variation in multi-copy regions of the human genome remains unknown. One expected manifestation of IGC in population genomic data is the presence of one-to-one paralogous SNPs that segregate identical alleles.ResultsHere, I use SNP genotype calls from the low-coverage phase 3 release of the 1000 Genomes Project to identify 15,790 parallel, shared SNPs in duplicated regions of the human genome. My approach for identifying these sites accounts for the potential redundancy of short read mapping in multi-copy genomic regions, thereby effectively eliminating false positive SNP calls arising from paralogous sequence variation. I demonstrate that independent mutation events to identical nucleotides at paralogous sites are not a significant source of shared polymorphisms in the human genome, consistent with the interpretation that these sites are the outcome of historical IGC events. These putative signals of IGC are enriched in genomic contexts previously associated with non-allelic homologous recombination, including clear signals in gene families that form tandem intra-chromosomal clusters.ConclusionsTaken together, my analyses implicate IGC, not point mutation, as the mechanism generating at least 2.7 % of single nucleotide variants in duplicated regions of the human genome.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-015-1681-3) contains supplementary material, which is available to authorized users.

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Section: Resultsmentioning

confidence: 99%

Interlocus gene conversion explains at least 2.7 % of single nucleotide variants in human segmental duplications

Dumont

2015

BMC Genomics

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“…A bias toward proximal donors is more likely when donors are separated by larger distances; such a bias would serve to stabilize the genome by reducing the probability of interactions over large distances and the associated risk of large-scale deletions and rearrangements. In both yeast and mammalian cells, SSA efficiency depends on the timing with which complementary single strands are exposed in direct repeats (17,35,36,59). Given that crossovers are restricted, most distal-proximal deletions probably arise by SSA, and the lack of bias in favor of proximal deletions suggests that end processing is rapid and extensive, exposing the relatively 32 P-labeled neo probe; the autoradiograph is shown above; equal loading is shown by ethidium bromide staining of 28S rRNA (B) Percentage of gene conversion events using 5Ј donor as a repair template for 5Ј3Ј, the net value for 5Ј3Ј and 5Ј3ЈSwitch, and MMTV5Ј3Ј Ϯ dex.…”

Section: Discussionmentioning

confidence: 99%

“…The complex patterns might result from multiple HR events or coupled HR/NHEJ events (54). This high deletion rate is not intrinsic to the triple repeat structure but is typical of closely spaced direct repeats (59). Among DP19 gene conversion products, there was a modest (2:1) bias in favor of the proximal donor (Table 3), but this is not significantly different from a 1:1 distribution (P ϭ 0.49).…”

Section: Experimental Designmentioning

confidence: 92%

Transcription of a Donor Enhances Its Use during Double-Strand Break-Induced Gene Conversion in Human Cells

Schildkraut

Miller

Nickoloff

2006

Molecular and Cellular Biology

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Homologous recombination (HR) mediates accurate repair of double-strand breaks (DSBs) but carries the risk of large-scale genetic change, including loss of heterozygosity, deletions, inversions, and translocations. Nearly one-third of the human genome consists of repetitive sequences, and DSB repair by HR often requires choices among several homologous repair templates, including homologous chromosomes, sister chromatids, and linked or unlinked repeats. Donor preference during DSB-induced gene conversion was analyzed by using several HR substrates with three copies of neo targeted to a human chromosome. Repair of I-SceI nucleaseinduced DSBs in one neo (the recipient) required a choice between two donor neo genes. When both donors were downstream, there was no significant bias for proximal or distal donors. When donors flanked the recipient, we observed a marked (85%) preference for the downstream donor. Reversing the HR substrate in the chromosome eliminated this preference, indicating that donor choice is influenced by factors extrinsic to the HR substrate. Prior indirect evidence suggested that transcription might increase donor use. We tested this question directly and found that increased transcription of a donor enhances its use during gene conversion. A preference for transcribed donors would minimize the use of nontranscribed (i.e., pseudogene) templates during repair and thus help maintain genome stability.DNA double-strand breaks (DSBs) are potentially lethal events that can be repaired by homologous recombination (HR) or nonhomologous end-joining (NHEJ). If left unrepaired, DSBs can lead to chromosome loss or cell death. DSBs are caused by ionizing radiation, X-rays, free radicals, chemicals, and nucleases and may occur at stalled replication forks (30). Although DSB repair can be accurate, misrepair can have serious consequences. Genomic rearrangements arising during DSB repair can lead to loss of heterozygosity and, ultimately, carcinogenesis through the activation of proto-oncogenes or inactivation of tumor suppressor genes (4, 45). HR appears to be essential for maintaining genome stability. Cells with defects in HR proteins such as BRCA1/2, XRCC2/3, and other RAD51 paralogs exhibit high levels of genomic instability (6,18,28,42,43,52,68). DSBs are a critical type of DNA damage; unlike single-strand breaks, which have a readily available template for repair, DSB repair by HR requires a search for a homologous template.Repetitive elements make up one-third of the mammalian genome and consist of coding DNA, and noncoding DNA such as satellites that can exist in thousands of copies. Repetitive sequences are scattered throughout the genome, including SINE and LINE elements, and ribosomal RNA gene repeats. HR between linked or unlinked repetitive elements can result in a variety of rearrangements including translocations, duplications, and deletions (45).When DNA is broken, there are many potential homologous sequences that could be used as a repair template, including linked or unlinked repeats, sister...

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“…Next, to investigate whether YY1 is involved in homology-directed repair, we used a reporterbased functional assay in which the Neo gene can be expressed only upon repair of DSBs produced by the endonuclease I-SceI 33 (Fig. 5a).…”

Section: Yy1 and Ino80 Promote Homology-directed Dna Repairmentioning

confidence: 99%

“…The human 293T cell line with a homologous recombination substrate (HR-293T) has been described 33 . I-SceI-induced homologous recombination was assayed as follows.…”

Section: Recombination Assaysmentioning

confidence: 99%

A YY1–INO80 complex regulates genomic stability through homologous recombination–based repair

Shi

Mulligan

et al. 2007

Nat Struct Mol Biol

187

220

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DNA damage repair is crucial for the maintenance of genome integrity and cancer suppression. We found that loss of the mouse transcription factor YY1 resulted in polyploidy and chromatid aberrations, which are signatures of defects in homologous recombination. Further biochemical analyses identified a YY1 complex comprising components of the evolutionarily conserved INO80 chromatin-remodeling complex. Notably, RNA interference-mediated knockdown of YY1 and INO80 increased cellular sensitivity toward DNA-damaging agents. Functional assays revealed that both YY1 and INO80 are essential in homologous recombination-based DNA repair (HRR), which was further supported by the finding that YY1 preferentially bound a recombination-intermediate structure in vitro. Collectively, these observations reveal a link between YY1 and INO80 and roles for both in HRR, providing new insight into mechanisms that control the cellular response to genotoxic stress.Genomic instability is a hallmark of cancer development and progression. Defects in DNA double-strand break (DSB) repair are believed to be responsible for chromosomal rearrangements and can be rectified by homologous recombination, which is particularly active in the S and G2 phases of the cell cycle, or by nonhomologous end joining (NHEJ), active in G1 phase 1,2 . Homologous recombination is an important mechanism for the repair of damaged chromosomes and damaged replication forks, and for several other aspects of chromosome maintenance 3 . Furthermore, impairment of homologous recombination is probably one of the underlying causes of breast, ovarian and other cancers 4,5 . INO80, a member of the Snf2p family of DNA-dependent ATPases, has been shown to positively regulate homologous recombination in a number of organisms [6][7][8] . The Ino80 complex is evolutionarily conserved, and components of the complex have been identified in mammalian cells 9 induces phosphorylation of histone H2A, and the yeast INO80 complex is known to be recruited directly to DSBs through an interaction with the phosphorylated histone H2A 7,8,10,11 ; however, the precise function of INO80 in DSB repair and its recruitment to the damage site are not completely understood 12 .Yin Yang-1 (YY1) is a zinc finger-containing Polycomb group (PcG) transcription factor that is essential in development [13][14][15][16] . Recently, a number of studies have shown that loss of YY1 increases the p53 protein level 17,18 , raising the possibility that YY1 may contribute to the regulation of genomic integrity. To investigate this possibility, we used genetic, biochemical and proteomic approaches. We discovered essential roles for YY1 in the cellular response to genotoxic stress and in the maintenance of chromosomal stability. Furthermore, we found multiple lines of evidence that link YY1 and the ATP-dependent chromatin-remodeling complex INO80 in DNA repair. We identified a YY1 complex containing components of the INO80 complex and confirmed this interaction of YY1 with components of the INO80 complex by ad...

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Gene conversion and deletion frequencies during double-strand break repair in human cells are controlled by the distance between direct repeats

Cited by 40 publications

References 46 publications

Interlocus gene conversion explains at least 2.7 % of single nucleotide variants in human segmental duplications

Interlocus gene conversion explains at least 2.7 % of single nucleotide variants in human segmental duplications

Transcription of a Donor Enhances Its Use during Double-Strand Break-Induced Gene Conversion in Human Cells

A YY1–INO80 complex regulates genomic stability through homologous recombination–based repair

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