scholarly journals Effect of nucleotide excision repair in human cells on intrachromosomal homologous recombination induced by UV and 1-nitrosopyrene.

1990 ◽  
Vol 10 (8) ◽  
pp. 3945-3951 ◽  
Author(s):  
N P Bhattacharyya ◽  
V M Maher ◽  
J J McCormick
Keyword(s):  
Homologous Recombination ◽  
Thymidine Kinase ◽  
Nucleotide Excision Repair ◽  
Mammalian Cells ◽  
Excision Repair ◽  
Dna Lesions ◽  
Repair Capacity ◽  
Nucleotide Excision ◽  
Specific Increase ◽  
Simplex Virus

To study the role of nucleotide excision repair in the induction of intrachromosomal homologous recombination in mammalian cells, we introduced a plasmid containing a substrate for recombination into three human cell lines that differ in their repair capacity and compared the frequency of recombination induced by UV radiation and by 1-nitrosopyrene. One strain had a normal capacity for nucleotide excision repair, the second exhibited an intermediate rate of repair, and the third, derived from a patient with xeroderma pigmentosum, had no ability to repair UV- or 1-nitrosopyrene-induced DNA damage. The endogenous thymidine kinase genes in these cell strains had been inactivated, and the cells contained an integrated copy of a plasmid carrying duplicated copies of the herpes simplex virus type 1 thymidine kinase (Htk) gene, each inactivated by an 8-base-pair XhoI site inserted at a unique site. A functional tk gene can only be generated by a productive recombination event between the two Htk genes. In all three stains, UV and 1-nitrosopyrene induced dose-dependent increases in the frequency of recombinants. However, the doses required to cause a specific increase in recombination in the repair-deficient strains were 10 to 30 times lower than the dose required for the cell strain with a normal capacity for repair. These results strongly suggest that unexcised DNA lesions, rather than excision repair per se, stimulate intrachromosomal homologous recombination. Southern blot analysis of DNA from representative recombinants indicated that in all cases one of the two Htk genes had become wild type (XhoI resistant). The majority (90%) retained the Htk duplication, consistent with nonreciprocal transfer of genetic information (gene conversion).

Effect of nucleotide excision repair in human cells on intrachromosomal homologous recombination induced by UV and 1-nitrosopyrene

1990 ◽  
Vol 10 (8) ◽  
pp. 3945-3951
Author(s):  
N P Bhattacharyya ◽  
V M Maher ◽  
J J McCormick
Keyword(s):  
Homologous Recombination ◽  
Thymidine Kinase ◽  
Nucleotide Excision Repair ◽  
Mammalian Cells ◽  
Excision Repair ◽  
Dna Lesions ◽  
Repair Capacity ◽  
Nucleotide Excision ◽  
Specific Increase ◽  
Simplex Virus

To study the role of nucleotide excision repair in the induction of intrachromosomal homologous recombination in mammalian cells, we introduced a plasmid containing a substrate for recombination into three human cell lines that differ in their repair capacity and compared the frequency of recombination induced by UV radiation and by 1-nitrosopyrene. One strain had a normal capacity for nucleotide excision repair, the second exhibited an intermediate rate of repair, and the third, derived from a patient with xeroderma pigmentosum, had no ability to repair UV- or 1-nitrosopyrene-induced DNA damage. The endogenous thymidine kinase genes in these cell strains had been inactivated, and the cells contained an integrated copy of a plasmid carrying duplicated copies of the herpes simplex virus type 1 thymidine kinase (Htk) gene, each inactivated by an 8-base-pair XhoI site inserted at a unique site. A functional tk gene can only be generated by a productive recombination event between the two Htk genes. In all three stains, UV and 1-nitrosopyrene induced dose-dependent increases in the frequency of recombinants. However, the doses required to cause a specific increase in recombination in the repair-deficient strains were 10 to 30 times lower than the dose required for the cell strain with a normal capacity for repair. These results strongly suggest that unexcised DNA lesions, rather than excision repair per se, stimulate intrachromosomal homologous recombination. Southern blot analysis of DNA from representative recombinants indicated that in all cases one of the two Htk genes had become wild type (XhoI resistant). The majority (90%) retained the Htk duplication, consistent with nonreciprocal transfer of genetic information (gene conversion).

scholarly journals Involvement of Nucleotide Excision Repair in a Recombination-Independent and Error-Prone Pathway of DNA Interstrand Cross-Link Repair

2001 ◽  
Vol 21 (3) ◽  
pp. 713-720 ◽  
Author(s):  
Xin Wang ◽  
Carolyn A. Peterson ◽  
Huyong Zheng ◽  
Rodney S. Nairn ◽  
Randy J. Legerski ◽  
...  
Keyword(s):  
Homologous Recombination ◽  
Nucleotide Excision Repair ◽  
Mammalian Cells ◽  
Cultured Cells ◽  
Excision Repair ◽  
Repair Pathway ◽  
Cross Link ◽  
Nucleotide Excision ◽  
Interstrand Cross Links ◽  
Cross Links

ABSTRACT DNA interstrand cross-links (ICLs) block the strand separation necessary for essential DNA functions such as transcription and replication and, hence, represent an important class of DNA lesion. Since both strands of the double helix are affected in cross-linked DNA, it is likely that conservative recombination using undamaged homologous regions as a donor may be required to repair ICLs in an error-free manner. However, in Escherichia coli and yeast, recombination-independent mechanisms of ICL repair have been identified in addition to recombinational repair pathways. To study the repair mechanisms of interstrand cross-links in mammalian cells, we developed an in vivo reactivation assay to examine the removal of interstrand cross-links in cultured cells. A site-specific psoralen cross-link was placed between the promoter and the coding region to inactivate the expression of green fluorescent protein or luciferase genes from reporter plasmids. By monitoring the reactivation of the reporter gene, we showed that a single defined psoralen cross-link was removed in repair-proficient cells in the absence of undamaged homologous sequences, suggesting the existence of an ICL repair pathway that is independent of homologous recombination. Mutant cell lines deficient in the nucleotide excision repair pathway were examined and found to be highly defective in the recombination-independent repair of ICLs, while mutants deficient in homologous recombination were found to be proficient. Mutation analysis of plasmids recovered from transfected cells showed frequent base substitutions at or near positions opposing a cross-linked thymidine residue. Based on these results, we suggest a distinct pathway for DNA interstrand cross-link repair involving nucleotide excision repair and a putative lesion bypass mechanism.

scholarly journals Nucleotide Excision Repair- and Polymerase η-Mediated Error-Prone Removal of Mitomycin C Interstrand Cross-Links

2003 ◽  
Vol 23 (2) ◽  
pp. 754-761 ◽  
Author(s):  
Huyong Zheng ◽  
Xin Wang ◽  
Amy J. Warren ◽  
Randy J. Legerski ◽  
Rodney S. Nairn ◽  
...  
Keyword(s):  
Mitomycin C ◽  
Nucleotide Excision Repair ◽  
Mammalian Cells ◽  
Excision Repair ◽  
Double Helix ◽  
Dna Lesions ◽  
Nucleotide Excision ◽  
Interstrand Cross Links ◽  
Cross Links

ABSTRACT Interstrand cross-links (ICLs) make up a unique class of DNA lesions in which both strands of the double helix are covalently joined, precluding strand opening during replication and transcription. The repair of DNA ICLs has become a focus of study since ICLs are recognized as the main cytotoxic lesion inflicted by an array of alkylating compounds used in cancer treatment. As is the case for double-strand breaks, a damage-free homologous copy is essential for the removal of ICLs in an error-free manner. However, recombination-independent mechanisms may exist to remove ICLs in an error-prone fashion. We have developed an in vivo reactivation assay that can be used to examine the removal of site-specific mitomycin C-mediated ICLs in mammalian cells. We found that the removal of the ICL from the reporter substrate could take place in the absence of undamaged homologous sequences in repair-proficient cells, suggesting a cross-link repair mechanism that is independent of homologous recombination. Systematic analysis of nucleotide excision repair mutants demonstrated the involvement of transcription-coupled nucleotide excision repair and a partial requirement for the lesion bypass DNA polymerase η encoded by the human POLH gene. From these observations, we propose the existence of a recombination-independent and mutagenic repair pathway for the removal of ICLs in mammalian cells.

scholarly journals Homologous Recombination but Not Nucleotide Excision Repair Plays a Pivotal Role in Tolerance of DNA-Protein Cross-links in Mammalian Cells

2009 ◽  
Vol 284 (40) ◽  
pp. 27065-27076 ◽  
Author(s):  
Toshiaki Nakano ◽  
Atsushi Katafuchi ◽  
Mayumi Matsubara ◽  
Hiroaki Terato ◽  
Tomohiro Tsuboi ◽  
...  
Keyword(s):  
Homologous Recombination ◽  
Nucleotide Excision Repair ◽  
Mammalian Cells ◽  
Excision Repair ◽  
Pivotal Role ◽  
Nucleotide Excision ◽  
Cross Links

Role of extracellular signal-regulated kinase (ERK) signaling in nucleotide excision repair and genotoxicity in response to As(III) and Pb(II)

2008 ◽  
Vol 80 (12) ◽  
pp. 2735-2750
Author(s):  
Ju-Pi Li ◽  
Chun-Yu Wang ◽  
Yen-An Tang ◽  
Yun-Wei Lin ◽  
Jia-Ling Yang
Keyword(s):  
Signaling Pathways ◽  
Nucleotide Excision Repair ◽  
Mammalian Cells ◽  
Excision Repair ◽  
Erk Signaling ◽  
Erk Activation ◽  
Extracellular Signal Regulated Kinase ◽  
Extracellular Signal ◽  
Nucleotide Excision

Arsenic and lead can induce genetic injuries and epigenetic signaling pathways in cultured mammalian cells. To test whether signaling pathways affect the extent of genetic injuries, we explored the impacts of extracellular signal-regulated kinase 1 and 2 (ERK) on nucleotide excision repair (NER), cytotoxicity, and genotoxicity following sodium arsenite [As(III)] and lead acetate [Pb(II)]. Sustained ERK activation was observed in human cells exposed to As(III) and Pb(II). As(III) inhibited the cellular NER synthesis capability; conversely, Pb(II) stimulated it. ERK activation contributed to the As(III)-induced NER inhibition and micronucleus formation. In contrast, this signal was required for inducing cellular NER activity and preventing mutagenesis following Pb(II). ERK activation by Pb(II) was dependent on protein kinase C (PKCα) that also exhibited anti-mutagenicity. Enforced expression of ERK signaling markedly elevated the cellular NER activity, which was suppressed by As(III). Nonetheless, ERK activation could counteract the cytotoxicity caused by these two metals. Together, the results indicate that pro-survival ERK signaling exhibits dual and opposing impacts on NER process following As(III) and Pb(II) exposures. The findings also suggest that ERK is an important epigenetic signaling in the determination of metal genotoxicity.

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