Incision by UvrABC excinuclease is a step in the path to mutagenesis by psoralen crosslinks in Escherichia coli.

Sladek, Frances M.; Melian, Agustin; Howard-Flanders, Paul

doi:10.1073/pnas.86.11.3982

Cited by 32 publications

(27 citation statements)

References 49 publications

(32 reference statements)

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“…RPA binds to triplexes with high affinity, with or without XPA, whereas XPA binds only in conjunction with RPA, but contributes to specificity by selectively diminishing nonspecific binding and͞or aggregation of RPA to undamaged DNA. Psoralen monoadducts and crosslinks are, at least in part, recognized and repaired by NER in human cells (39)(40)(41)(42)(43)(44)(45)(46). Psoralen monoadducts are somewhat helix-stabilizing, whereas crosslinks are helix-destabilizing (47,48).…”

Section: Discussionmentioning

confidence: 99%

Human XPA and RPA DNA repair proteins participate in specific recognition of triplex-induced helical distortions

Vásquez

Christensen

et al. 2002

Proc. Natl. Acad. Sci. U.S.A.

111

128

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Nucleotide excision repair (NER) plays a central role in maintaining genomic integrity by detecting and repairing a wide variety of DNA lesions. Xeroderma pigmentosum complementation group A protein (XPA) is an essential component of the repair machinery, and it is thought to be involved in the initial step as a DNA damage recognition and͞or confirmation factor. Human replication protein A (RPA) and XPA have been reported to interact to form a DNA damage recognition complex with greater specificity for damaged DNA than XPA alone. The mechanism by which these two proteins recognize such a wide array of structures resulting from different types of DNA damage is not known. One possibility is that they recognize a common feature of the lesions, such as distortions of the helical backbone. We have tested this idea by determining whether human XPA and RPA proteins can recognize the helical distortions induced by a DNA triple helix, a noncanonical DNA structure that has been shown to induce DNA repair, mutagenesis, and recombination. We measured binding of XPA and RPA, together or separately, to substrates containing triplexes with three, two, or no strands covalently linked by psoralen conjugation and photoaddition. We found that RPA alone recognizes all covalent triplex structures, but also forms multivalent nonspecific DNA aggregates at higher concentrations. XPA by itself does not recognize the substrates, but it binds them in the presence of RPA. Addition of XPA decreases the nonspecific DNA aggregate formation. These results support the hypothesis that the NER machinery is targeted to helical distortions and demonstrate that RPA can recognize damaged DNA even without XPA.triple helix ͉ nucleotide excision repair B ecause genome stability is critical for cell survival, extremely sensitive DNA repair mechanisms have evolved to protect the genome from both internal and external assaults. Defects in these repair mechanisms can lead to severe disorders such as xeroderma pigmentosum, ataxia telangiectasia, Fanconi anemia, and Bloom syndrome (reviewed in refs. 1-5). One of the major consequences of these defects is an enhanced predisposition to cancer. It has been reported that 80-90% of human cancers are the result of DNA damage (6).Many types of DNA damage, spontaneous and induced, develop from both endogenous and exogenous sources. Exogenous damages, both chemical and physical, arise from exposure to UV and ionizing radiation and from natural and synthetic chemicals. Cellular metabolic processes lead to internal sources of DNA damage; for example, it is estimated that loss of a purine base (depurination) occurs at a rate of nearly 20,000 events per cell per day (7).In humans, different types of DNA damage are thought to be repaired by one of several mechanisms, including nucleotide excision repair (NER; reviewed in refs. 8 and 9). NER removes covalent DNA lesions and is the only known mechanism for removing bulky DNA adducts in humans. The damaged base is removed by endonucleolytic cleavage of the phosphodiester bond...

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Section: Discussionmentioning

confidence: 99%

Human XPA and RPA DNA repair proteins participate in specific recognition of triplex-induced helical distortions

Vásquez

Christensen

et al. 2002

Proc. Natl. Acad. Sci. U.S.A.

111

128

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“…The ntn jt5 allele was of recessive inheritance and fully penetrant. If TMP mutagenesis induces deletions in the zebrafish genome as in E. coli and C. elegans (Sladek et al, 1989;Yandell et al, 1994;Liu et al, 1999), it would be possible to directly clone deleted DNA segments by subtraction between pooled wild-type and mutant genomes. Alternatively, such whole-genome subtraction may allow us to obtain DNA segments tightly linked to the ntn locus if there were enough RFLPs in the AB strain genome.…”

Section: Resultsmentioning

confidence: 99%

“…TMP, in combination with UV irradiation, forms interstrand crosslinks with DNA double helices and frequently results in deletions, possibly via incomplete nucleotide excision and recombination repair in E. coli and C. elegans (Cimino et al, 1985;Sladek et al, 1989;Yandell et al, 1994). Subtraction of the mutant genome from the wild-type genome should yield deleted genes or tightly linked RFLP markers.…”

Section: Tmp Mutagenesis and Rda Cloningmentioning

confidence: 99%

“…The no tectal neuron (ntn) mutation impaired the development of tectal neurons and eyes, whereas the edawakare mutation affected the arborization of the trigeminal ganglion and RohonBeard sensory neurons. TMP is a DNA crosslinking agent that can frequently induce small deletions in combination with UV irradiation in Escherichia coli and Caenorhabditis elegans (Sladek et al, 1989;Yandell et al, 1994;Liu et al, 1999). A potential advantage of the method would be that one could isolate mutagenized genes directly by whole-genome subtraction using TMP-induced deletions as molecular tags.…”

Section: Introductionmentioning

confidence: 99%

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Identification of chaperonin CCTγ subunit as a determinant of retinotectal development by whole-genome subtraction cloning from zebrafishno tectal neuronmutant

Matsuda

Mishina

2004

Development

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Zebrafish no tectal neuron (ntn) mutant obtained by trimethylpsoralen (TMP) mutagenesis showed defects in tectal neuropil formation and small eyes. We carried out whole-genome subtraction between wild-type and mutant zebrafish embryos using the representational difference analysis (RDA) method. Nineteen subtraction products enabled us to construct genetic and physical maps of the ntn region. Direct selection of cDNAs using a YAC clone encompassing the ntn locus and RT-PCR analysis of transcripts identified a 143 bp deletion in the cct3 gene encoding the γ subunit of chaperonin containing TCP-1 (CCT). Injection of antisense cct3 morpholino oligonucleotides into zebrafish embryos induced characteristic ntn phenotypes including defects in retinal ganglion cell (RGC) differentiation and tectal neuropil formation. Moreover,injection of cct3 mRNA successfully rescued ntn mutant embryos. Our results suggest that RDA is an efficient and widely applicable cloning strategy in zebrafish genetics. The strong expression of the cct3 mRNA started in the entire embryos by 12 hpf and was sustained thereafter, but there were no detectable abnormalities in body patterning and neurogenesis in ntn mutant embryos at 30 hpf. The expression patterns of transcription factor genes ath5 and brn3b that are essential for the development and maintenance of RGCs were indistinguishable between wild-type and ntn mutant embryos, but those of early and late differentiation markers of RGCs, nicotinic acetylcholine receptor β3 and zn5, were diminished in mutant embryos. Immunostaining of acetylated tubulin also revealed the impairment of RGC neurite extension. Thus, the ntnmutation of the cct3 gene impaired the differentiation of retinal neuroepithelial cells to RGCs. Similarly, the expression of brn3b was normal in the tectum of ntn mutants, but tectal neuropil formation was abolished. These results suggest that the γ subunit of chaperonin CCT plays an essential role in retinotectal development.

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“…Studies in Escherichia coli have shown that it causes both single-base changes and deletions (Piette et al 1985;Sladek et al 1989). C. elegans researchers became interested in the potential of ultraviolet trimethylpsoralen (UV/TMP) to generate deletions in worms after the first deletions in this organism were isolated using this mutagen (Yandell et al 1994).…”

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confidence: 99%

Whole-Genome Profiling of Mutagenesis inCaenorhabditis elegans

et al. 2010

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Deep sequencing offers an unprecedented view of an organism's genome. We describe the spectrum of mutations induced by three commonly used mutagens: ethyl methanesulfonate (EMS), N-ethyl-Nnitrosourea (ENU), and ultraviolet trimethylpsoralen (UV/TMP) in the nematode Caenorhabditis elegans. Our analysis confirms the strong GC to AT transition bias of EMS. We found that ENU mainly produces A to T and T to A transversions, but also all possible transitions. We found no bias for any specific transition or transversion in the spectrum of UV/TMP-induced mutations. In 10 mutagenized strains we identified 2723 variants, of which 508 are expected to alter or disrupt gene function, including 21 nonsense mutations and 10 mutations predicted to affect mRNA splicing. This translates to an average of 50 informative mutations per strain. We also present evidence of genetic drift among laboratory wild-type strains derived from the Bristol N2 strain. We make several suggestions for best practice using massively parallel short read sequencing to ensure mutation detection.

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Incision by UvrABC excinuclease is a step in the path to mutagenesis by psoralen crosslinks in Escherichia coli.

Cited by 32 publications

References 49 publications

Human XPA and RPA DNA repair proteins participate in specific recognition of triplex-induced helical distortions

Human XPA and RPA DNA repair proteins participate in specific recognition of triplex-induced helical distortions

Identification of chaperonin CCTγ subunit as a determinant of retinotectal development by whole-genome subtraction cloning from zebrafishno tectal neuronmutant

Whole-Genome Profiling of Mutagenesis inCaenorhabditis elegans

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