Single strand and double strand DNA damage-induced reciprocal recombination in yeast. Dependence on nucleotide excision repair and<i>RAD1</i>recombination

Saffran, Wilma A.; Greenberg, Ross B.; Thaler-Scheer, Mindy S.; Jones, Monica M.

doi:10.1093/nar/22.14.2823

Cited by 29 publications

(27 citation statements)

References 47 publications

(42 reference statements)

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“…Psoralen-stimulated recombination has been reported for bacterial (5,22), yeast (26,(29)(30)(31), and mammalian (8,34,39) cells. With two exceptions (8,34), the adducts in these studies were random in location and variable in number.…”

Section: Discussionmentioning

confidence: 99%

“…Cross-links present a particular challenge to the repair machinery, since both strands are damaged at a single site. Like some other types of lesion, psoralen cross-links induce recombination in bacterial (5, 22, 36), yeast (26,(29)(30)(31), and mammalian (8, 34, 39) cells. Double-strand breaks (DSBs) have been either inferred or directly demonstrated to be involved in this process.…”

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confidence: 99%

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Processing of Targeted Psoralen Cross-Links in Xenopus Oocytes

Segal

Faruqi

Glazer

et al. 1997

Molecular and Cellular Biology

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Psoralen cross-links have been shown to be both mutagenic and recombinagenic in bacterial, yeast, and mammalian cells. Double-strand breaks (DSBs) have been implicated as intermediates in the removal of psoralen cross-links. Recent work has suggested that site-specific mutagenesis and recombination might be achieved through the use of targeted psoralen adducts. The fate of plasmids containing psoralen adducts was evaluated in Xenopus oocytes, an experimental system that has well-characterized recombination capabilities and advantages in the analysis of intermediates in DNA metabolism. Psoralen adducts were delivered to a specific site by a triplex-forming oligonucleotide. These lesions are clearly recognized and processed in oocytes, since mutagenesis was observed at the target site. The spectrum of induced mutations was compared with that found in similar studies in mammalian cells. Plasmids carrying multiple random adducts were preferentially degraded, perhaps due to the introduction of DSBs. However, when DNAs carrying site-specific adducts were examined, no plasmid loss was observed and removal of cross-links was found to be very slow. Sensitive assays for DSB-dependent homologous recombination were performed with substrates with one or two cross-link sites. No adduct-stimulated recombination was observed with a single lesion, and only very low levels were observed with paired lesions, even when a large proportion of the cross-links was removed by the oocytes. We conclude that DSBs or other recombinagenic structures are not efficiently formed at psoralen adducts in Xenopus oocytes. While psoralen is not a promising reagent for stimulating site-specific recombination, it is effective in inducing targeted mutations.Photoinduced interstrand psoralen cross-links in cellular DNA are detected as genomic damage, and cells of all types attempt to repair them. Cross-links present a particular challenge to the repair machinery, since both strands are damaged at a single site. Like some other types of lesion, psoralen cross-links induce recombination in bacterial (5, 22, 36), yeast (26,(29)(30)(31), and mammalian (8, 34, 39) cells. Double-strand breaks (DSBs) have been either inferred or directly demonstrated to be involved in this process. In yeast cells (7,18,23,31), DSBs have been shown to be intermediates in the cellular repair of psoralen cross-links, and their production requires nucleotide excision repair (NER) machinery (RAD3 epistasis group). Recombinational repair (RAD52 epistasis group) is required to rejoin the breaks made during cross-link removal and may participate in generating the DSBs (7).NER is the major mechanism by which all living systems repair DNA that has been damaged by helix-distorting modifications, such as pyrimidine dimers and psoralen adducts (10,32,42). Briefly, NER proteins recognize and bind to the damaged site. In eukaryotes, incisions are made on the damaged strand 22 to 24 nucleotides 5Ј and 5 nucleotides 3Ј to the damage. The 27-to 29-base oligonucleotide containing the damage is e...

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Section: Discussionmentioning

confidence: 99%

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confidence: 99%

Processing of Targeted Psoralen Cross-Links in Xenopus Oocytes

Segal

Faruqi

Glazer

et al. 1997

Molecular and Cellular Biology

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“…One oligonucleotide contains a central poly(C) 30 region, whereas the other contains poly(T) 30 to produce a centrally unpaired poly(C)⅐poly(T) region of 30 nucleotides flanked by duplex DNA. The DNA sequences and method of preparation are described elsewhere (19).…”

Section: Methodsmentioning

confidence: 99%

“…The DNA sequences and method of preparation are described elsewhere (19). The poly(T) 30 -containing strand (oligo 2) was labeled with 32 P at the 5Ј-end prior to annealing. Synthetic Holliday junction X12 was prepared as described previously (20).…”

Section: Methodsmentioning

confidence: 99%

Role of the Rad1 and Rad10 Proteins in Nucleotide Excision Repair and Recombination

Davies¹,

Friedberg

Tomkinson

et al. 1995

Journal of Biological Chemistry

121

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In Saccharomyces cerevisiae, the RAD1 and RAD10 genes are involved in DNA nucleotide excision repair (NER) and in a pathway of mitotic recombination that occurs between direct repeat DNA sequences. In this paper, we show that purified Rad1 and Rad10 interact with a synthetic bubble structure and incise the DNA at the 5-side of the centrally unpaired region. When Rad1-Rad10 and purified XPG protein (the human homolog of yeast Rad2 protein) were co-incubated with the DNA substrate, we observed incisions at both ends of the bubble. This reaction mimics the dual incision step in nucleotide excision repair in vivo. In addition, the recent suggestion that Rad1 can act to resolve Holliday junctions (Habraken, Y., Sung, P., Prakash, L., and Prakash, S. (1994) Nature 371, 531-534), explaining the recombination defect observed in rad1 mutants, has been further investigated. However, using proteins purified in two different laboratories we were unable to show any interaction between Rad1 and synthetic Holliday junctions. The role that Rad1-Rad10 plays in recombination is likely to resemble its activity in NER by acting upon partially unpaired DNA intermediates such as those formed by recombination mechanisms involving single-strand DNA annealing.The process of nucleotide excision repair plays a major role in the removal of DNA lesions following DNA damage. In bacteria, excision repair is initiated by three proteins, UvrA, UvrB, and UvrC, which promote a dual incision reaction by cleaving the 3rd to 5th phosphodiester bond 3Ј to the lesion and the 8th bond at the 5Ј-side of the lesion. The 12-13-nucleotide-long oligomer containing the lesion is then removed from the DNA, and the single-stranded gap is filled and sealed by DNA polymerases and DNA ligase (1, 2). Dual incision is also observed in eukaryotes, and higher organisms excise a fragment of 27-29 nucleotides in length (3).The isolation of UV-sensitive mutants has helped to define the genetic complexity of nucleotide excision repair in Saccharomyces cerevisiae, and it is now known that many genes are involved in the process. These include RAD1, RAD2, RAD3, RAD4, RAD7, RAD10, RAD14, RAD16, RAD23, TFB1, SSL1, and SSL2 (1,4). Remarkably, the sequences of excision repair genes in yeast and mammalian cells are highly homologous, and the enzymology of the two systems is very similar. The RAD1 and RAD10 genes of S. cerevisiae encode polypeptides that interact to form a stable complex (5-7) that exhibits a structure-specific endonuclease activity. The endonuclease cleaves single-stranded DNA tails with 3Ј-ends at the junction with duplex DNA (8), indicating that Rad1-Rad10 promotes the 5Ј-incision event in NER.1 Genetic studies show that rad1 and rad10 mutants also exhibit defects in mitotic recombination (9 -14). In particular, RAD1 and RAD10 are required for intrachromosomal recombination between direct repeats and affect the integration of linear DNA molecules and circular plasmids into genomic sequences. RAD1 is also required for doublestrand break repair between two dir...

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“…Mutations in RAD1 and RAD10 reduce intrachromosomal recombination between directly repeated sequences and decrease the efficiency of homologous integration of linear DNA fragments and circular plasmids (30,45,47,48). The RAD1 function is also required for elevated recombination in cdc9 and top3 mutants (2,47) as well as for mitotic recombination stimulated by RNA polymerase I-or II-dependent transcription (59,67).…”

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confidence: 99%

RAD1 and RAD10, but Not Other Excision Repair Genes, Are Required for Double-Strand Break-Induced Recombination in Saccharomyces cerevisiae

Ivanov

Haber

1995

Molecular and Cellular Biology

209

158

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HO endonuclease-induced double-strand breaks (DSBs) in the yeast Saccharomyces cerevisiae can be repaired by the process of gap repair or, alternatively, by single-strand annealing if the site of the break is flanked by directly repeated homologous sequences. We have shown previously (J. Fishman-Lobell and J. E. Haber, Science 258:480-484, 1992) that during the repair of an HO-induced DSB, the excision repair gene RAD1 is needed to remove regions of nonhomology from the DSB ends. In this report, we present evidence that among nine genes involved in nucleotide excision repair, only RAD1 and RAD10 are required for removal of nonhomologous sequences from the DSB ends. rad1⌬ and rad10⌬ mutants displayed a 20-fold reduction in the ability to execute both gap repair and single-strand annealing pathways of HO-induced recombination. Mutations in RAD2, RAD3, and RAD14 reduced HO-induced recombination by about twofold. We also show that RAD7 and RAD16, which are required to remove UV photodamage from the silent HML locus, are not required for MAT switching with HML or HMR as a donor. Our results provide a molecular basis for understanding the role of yeast nucleotide excision repair genes and their human homologs in DSB-induced recombination and repair.Genes belonging to the epistasis group RAD3 are required for nucleotide excision repair (NER) of UV-damaged DNA in the yeast Saccharomyces cerevisiae (for a recent review, see reference 40). Some of these genes also function in other pathways of DNA metabolism. Genomic deletions of RAD3 (34) and SSL2 (RAD25) (17, 38) are lethal in haploid cells, indicating their essential role in maintaining cell viability. Experiments using conditional lethal mutations of RAD3 and SSL2 have demonstrated that their essential function is most likely their role in RNA polymerase II transcription (18,19,41). Substantiating the idea of an intimate linkage between NER and transcription were recent findings that protein products of at least RAD2, RAD3, RAD4, and SSL2 are components of or are physically associated with RNA polymerase II basal transcription factor b (5, 12).Experimental data also show that some NER genes have a role in mitotic recombination. Mutations in RAD1 and RAD10 reduce intrachromosomal recombination between directly repeated sequences and decrease the efficiency of homologous integration of linear DNA fragments and circular plasmids (30,45,47,48). The RAD1 function is also required for elevated recombination in cdc9 and top3 mutants (2, 47) as well as for mitotic recombination stimulated by RNA polymerase I-or II-dependent transcription (59, 67). The effect of rad1 and rad10 mutations on recombination seems to be rather specific in the sense that mitotic, but not meiotic, events are affected in the mutant strains (48). Moreover, only rad1 and rad10, not other NER mutants, showed defects in spontaneous recombination between directly repeated genes (47).A key to understanding a precise role of RAD1 in mitotic recombination came from our previous studies on the genetic control ...

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Single strand and double strand DNA damage-induced reciprocal recombination in yeast. Dependence on nucleotide excision repair andRAD1recombination

Cited by 29 publications

References 47 publications

Processing of Targeted Psoralen Cross-Links in Xenopus Oocytes

Processing of Targeted Psoralen Cross-Links in Xenopus Oocytes

Role of the Rad1 and Rad10 Proteins in Nucleotide Excision Repair and Recombination

RAD1 and RAD10, but Not Other Excision Repair Genes, Are Required for Double-Strand Break-Induced Recombination in Saccharomyces cerevisiae

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