IDENTIFICATION OF A SECOND LOCUS IN DROSOPHILA MELANOGASTER REQUIRED FOR EXCISION REPAIR

Genetics ◽  
1982 ◽  
Vol 100 (2) ◽  
pp. 239-257
Author(s):  
J B Boyd ◽  
R D Snyder ◽  
P V Harris ◽  
J M Presley ◽  
S F Boyd ◽  
...  
Keyword(s):  
Excision Repair ◽  
Single Strand ◽  
X Rays ◽  
Dna Breaks ◽  
Strand Breaks ◽  
Single Strand Breaks ◽  
Pyrimidine Dimers ◽  
Biochemical Assays ◽  
X Irradiation

ABSTRACT The mus(2)201 locus in Drosophila is defined by two mutant alleles that render homozygous larvae hypersensitive to mutagens. Both alleles confer strong in vivo somatic sensitivity to treatment by methyl methanesulfonate, nitrogen mustard and ultraviolet radiation but only weak hypersensitivity to X-irradiation. Unlike the excision-defective mei-9 mutants identified in previous studies, the mus(2)201 mutants do not affect female fertility and do not appear to influence recombination proficiency or chromosome segregation in female meiocytes.—Three independent biochemical assays reveal that cell cultures derived from embryos homozygous for the mus(2)  D1 allele are devoid of detectable excision repair. 1. Such cells quantitatively retain pyrimidine dimers in their DNA for 24 hr following UV exposure. 2. No measurable unscheduled DNA synthesis is induced in mutant cultures by UV treatment. 3. Single-strand DNA breaks, which are associated with normal excision repair after treatment with either UV or N-acetoxy-N-acetyl-2-aminofluorene,* are much reduced in these cultures. Mutant cells possess a normal capacity for postreplication repair and the repair of single-strand breaks induced by X-rays.

scholarly journals Sensitive CometChip assay for screening potentially carcinogenic DNA adducts by trapping DNA repair intermediates

2019 ◽  
Vol 48 (3) ◽  
pp. e13-e13 ◽  
Author(s):  
Le P Ngo ◽  
Norah A Owiti ◽  
Carol Swartz ◽  
John Winters ◽  
Yang Su ◽  
...  
Keyword(s):  
Comet Assay ◽  
Dna Adducts ◽  
Excision Repair ◽  
Environmental Chemicals ◽  
Single Strand ◽  
Strand Breaks ◽  
Single Strand Breaks ◽  
Bulky Dna Adducts ◽  
Dna Strand

Abstract Genotoxicity testing is critical for predicting adverse effects of pharmaceutical, industrial, and environmental chemicals. The alkaline comet assay is an established method for detecting DNA strand breaks, however, the assay does not detect potentially carcinogenic bulky adducts that can arise when metabolic enzymes convert pro-carcinogens into a highly DNA reactive products. To overcome this, we use DNA synthesis inhibitors (hydroxyurea and 1-β-d-arabinofuranosyl cytosine) to trap single strand breaks that are formed during nucleotide excision repair, which primarily removes bulky lesions. In this way, comet-undetectable bulky lesions are converted into comet-detectable single strand breaks. Moreover, we use HepaRG™ cells to recapitulate in vivo metabolic capacity, and leverage the CometChip platform (a higher throughput more sensitive comet assay) to create the ‘HepaCometChip’, enabling the detection of bulky genotoxic lesions that are missed by current genotoxicity screens. The HepaCometChip thus provides a broadly effective approach for detection of bulky DNA adducts.

scholarly journals DNA DAMAGE AND REPAIR IN EUKARYOTIC CELLS

Genetics ◽  
1974 ◽  
Vol 78 (1) ◽  
pp. 139-148
Author(s):  
R B Painter
Keyword(s):  
Ionizing Radiation ◽  
Excision Repair ◽  
Double Strand Breaks ◽  
Single Strand ◽  
Premature Aging ◽  
Cross Linking ◽  
Base Damage ◽  
Strand Breaks ◽  
Single Strand Breaks ◽  
Post Replication Repair

ABSTRACT Damage in DNA after irradiation can be classified into five kinds: base damage, single-strand breaks, double-strand breaks, DNA-DNA cross-linking, and DNA-protein cross-linking. Of these, repair of base damage is the best understood. In eukaryotes, at least three repair systems are known that can deal with base damage: photoreactivation, excision repair, and post-replication repair. Photoreactivation is specific for UV-induced damage and occurs widely throughout the biosphere, although it seems to be absent from placental mammals. Excision repair is present in prokaryotes and in animals but does not seem to be present in plants. Post-replication repair is poorly understood. Recent reports indicate that growing points in mammalian DNA simply skip past UV-induced lesions, leaving gaps in newly made DNA that are subsequently filled in by de novo synthesis. Evidence that this concept is oversimplified or incorrect is presented.—Single-strand breaks are induced by ionizing radiation but most cells can rapidly repair most or all of them, even after supralethal doses. The chemistry of the fragments formed when breaks are induced by ionizing radiation is complex and poorly understood. Therefore, the intermediate steps in the repair of single-strand breaks are unknown. Double-strand breaks and the two kinds of cross-linking have been studied very little and almost nothing is known about their mechanisms for repair.—The role of mammalian DNA repair in mutations is not known. Although there is evidence that defective repair can lead to cancer and/or premature aging in humans, the relationship between the molecular defects and the diseased state remains obscure.

scholarly journals A nucleotide resolution map of Top2-linked DNA breaks in the yeast and human genome

2019 ◽  
Vol 10 (1) ◽  
Author(s):  
William H. Gittens ◽  
Dominic J. Johnson ◽  
Rachal M. Allison ◽  
Tim J. Cooper ◽  
Holly Thomas ◽  
...  
Keyword(s):  
Dna Cleavage ◽  
Genome Stability ◽  
Dna Double Strand Breaks ◽  
Dna Breaks ◽  
Strand Breaks ◽  
Single Strand Breaks ◽  
Human Genomes ◽  
Nucleotide Resolution

Abstract DNA topoisomerases are required to resolve DNA topological stress. Despite this essential role, abortive topoisomerase activity generates aberrant protein-linked DNA breaks, jeopardising genome stability. Here, to understand the genomic distribution and mechanisms underpinning topoisomerase-induced DNA breaks, we map Top2 DNA cleavage with strand-specific nucleotide resolution across the S. cerevisiae and human genomes—and use the meiotic Spo11 protein to validate the broad applicability of this method to explore the role of diverse topoisomerase family members. Our data characterises Mre11-dependent repair in yeast and defines two strikingly different fractions of Top2 activity in humans: tightly localised CTCF-proximal, and broadly distributed transcription-proximal, the latter correlated with gene length and expression. Moreover, single nucleotide accuracy reveals the influence primary DNA sequence has upon Top2 cleavage—distinguishing sites likely to form canonical DNA double-strand breaks (DSBs) from those predisposed to form strand-biased DNA single-strand breaks (SSBs) induced by etoposide (VP16) in vivo.

Less Repair of Pyrimidine Dimers and Single-Strand Breaks in Genes by Scid Cells

1999 ◽  
Vol 264 (3) ◽  
pp. 878-882 ◽  
Author(s):  
Rudaina H. Alrefai ◽  
E.Jeffrey Beecham ◽  
Vilhelm A. Bohr ◽  
Patricia J. Gearhart
Keyword(s):  
Single Strand ◽  
Strand Breaks ◽  
Single Strand Breaks ◽  
Pyrimidine Dimers ◽  
Scid Cells

scholarly journals Artificial and Solar UV Radiation Induces Strand Breaks and Cyclobutane Pyrimidine Dimers in Bacillus subtilis Spore DNA

2000 ◽  
Vol 66 (1) ◽  
pp. 199-205 ◽  
Author(s):  
Tony A. Slieman ◽  
Wayne L. Nicholson
Keyword(s):  
Bacillus Subtilis ◽  
Uv Irradiation ◽  
Double Strand Breaks ◽  
Single Strand ◽  
Complex Spectrum ◽  
Cyclobutane Pyrimidine Dimers ◽  
Strand Breaks ◽  
Single Strand Breaks ◽  
Pyrimidine Dimers ◽  
Solar Uv

ABSTRACT The loss of stratospheric ozone and the accompanying increase in solar UV flux have led to concerns regarding decreases in global microbial productivity. Central to understanding this process is determining the types and amounts of DNA damage in microbes caused by solar UV irradiation. While UV irradiation of dormant Bacillus subtilis endospores results mainly in formation of the “spore photoproduct” 5-thyminyl-5,6-dihydrothymine, genetic evidence indicates that an additional DNA photoproduct(s) may be formed in spores exposed to solar UV-B and UV-A radiation (Y. Xue and W. L. Nicholson, Appl. Environ. Microbiol. 62:2221–2227, 1996). We examined the occurrence of double-strand breaks, single-strand breaks, cyclobutane pyrimidine dimers, and apurinic-apyrimidinic sites in spore DNA under several UV irradiation conditions by using enzymatic probes and neutral or alkaline agarose gel electrophoresis. DNA from spores irradiated with artificial 254-nm UV-C radiation accumulated single-strand breaks, double-strand breaks, and cyclobutane pyrimidine dimers, while DNA from spores exposed to artificial UV-B radiation (wavelengths, 290 to 310 nm) accumulated only cyclobutane pyrimidine dimers. DNA from spores exposed to full-spectrum sunlight (UV-B and UV-A radiation) accumulated single-strand breaks, double-strand breaks, and cyclobutane pyrimidine dimers, whereas DNA from spores exposed to sunlight from which the UV-B component had been removed with a filter (“UV-A sunlight”) accumulated only single-strand breaks and double-strand breaks. Apurinic-apyrimidinic sites were not detected in spore DNA under any of the irradiation conditions used. Our data indicate that there is a complex spectrum of UV photoproducts in DNA of bacterial spores exposed to solar UV irradiation in the environment.

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