Formation of DNA interstrand cross-links as a marker of Mitomycin C bioreductive activation and chemosensitivity

ABSTRACT DNA interstrand cross-links (ICLs) represent lethal DNA damage, because they block transcription, replication, and segregation of DNA. Because of their genotoxicity, agents inducing ICLs are often used in antitumor therapy. The repair of ICLs is complex and involves proteins belonging to nucleotide excision, recombination, and translesion DNA repair pathways in Escherichia coli, Saccharomyces cerevisiae, and mammals. We cloned and analyzed mammalian homologs of the S. cerevisiae gene SNM1(PSO2), which is specifically involved in ICL repair. Human Snm1, a nuclear protein, was ubiquitously expressed at a very low level. We generated mouse SNM1−/− embryonic stem cells and showed that these cells were sensitive to mitomycin C. In contrast to S. cerevisiae snm1 mutants, they were not significantly sensitive to other ICL agents, probably due to redundancy in mammalian ICL repair and the existence of other SNM1homologs. The sensitivity to mitomycin C was complemented by transfection of the human SNM1 cDNA and by targeting of a genomic cDNA-murine SNM1 fusion construct to the disrupted locus. We also generated mice deficient for murine SNM1. They were viable and fertile and showed no major abnormalities. However, they were sensitive to mitomycin C. The ICL sensitivity of the mammalian SNM1 mutant suggests that SNM1function and, by implication, ICL repair are at least partially conserved between S. cerevisiae and mammals.

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Rescue of mitomycin C- or psoralen-inactivated Micrococcus radiodurans by additional exposure to radiation or alkylating agents

Journal of Bacteriology ◽

10.1128/jb.152.3.976-982.1982 ◽

1982 ◽

Vol 152 (3) ◽

pp. 976-982

Author(s):

M T Hansen

Keyword(s):

Mitomycin C ◽

Uv Radiation ◽

Uv Light ◽

Alkylating Agents ◽

Repair System ◽

Wild Type ◽

Interstrand Cross Links ◽

Cross Links ◽

Cellular Dna ◽

Constitutive Synthesis

The processing of damaged DNA was altered in a mitomycin C-sensitive mutant (mtcA) of Micrococcus radiodurans. Even though the mutant retained resistance to 254-nm UV radiation, it did not, in contrast to the wild-type strain, show any excessive DNA degradation or cell death when incubated with chloramphenicol after sublethal doses of either UV light or mitomycin C. The results suggest the constitutive synthesis of an enzyme system responsible for wild-type proficiency in the repair of mitomycin C-induced damage. An alternative system able to repair damage caused by mitomycin C was demonstrated in the mtcA background. In this strain, additional damage inflicted upon the cellular DNA effected a massive rescue of cells previously inactivated by mitomycin C. Rescue was provoked by ionizing radiation, by UV light, or by simple alkylating agents. Cells treated with psoralen plus near-UV radiation could be rescued only when inactivation was due primarily to psoralen-DNA interstrand cross-links rather than to monoadducts. The rescue of inactivated cells was prevented in the presence of chloramphenicol. These results can be interpreted most readily in terms of an alternative repair system able to overcome DNA interstrand cross-links produced by mitomycin C or psoralen plus near-UV light, but induced only by the more abundant number of damages produced by radiation or simple alkylating agents.

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Lack of correlation between repair of DNA interstrand cross-links and hypersensitivity of hamster cells towards mitomycin C and cisplatin

FEBS Letters ◽

10.1016/s0014-5793(98)01209-5 ◽

1998 ◽

Vol 437 (1-2) ◽

pp. 97-100 ◽

Cited By ~ 8

Author(s):

Florence Larminat ◽

Gilles Cambois ◽

Małgorzata Z. Zdzienicka ◽

Martine Defais

Keyword(s):

Mitomycin C ◽

Interstrand Cross Links ◽

Cross Links

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

Molecular and Cellular Biology ◽

10.1128/mcb.23.2.754-761.2003 ◽

2003 ◽

Vol 23 (2) ◽

pp. 754-761 ◽

Cited By ~ 95

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.

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Electron microscopic study of the membrane glycoproteins in the rat kidney after cisplatin treatment

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100156547 ◽

1989 ◽

Vol 47 ◽

pp. 912-913

Author(s):

S.K. Aggarwal

Keyword(s):

Alkaline Phosphatase ◽

Rat Kidney ◽

Light And Electron Microscopy ◽

Kidney Tissue ◽

Membrane Glycoproteins ◽

Electron Microscopic ◽

Before And After ◽

Interstrand Cross Links ◽

Cross Links

The proposed primary mechanism of action of the anticancer drug cisplatin (Cis-DDP) is through its interaction with DNA, mostly through DNA intrastrand cross-links or DNA interstrand cross-links. DNA repair mechanisms can circumvent this arrest thus permitting replication and transcription to proceed. Various membrane transport enzymes have also been demonstrated to be effected by cisplatin. Glycoprotein alkaline phosphatase was looked at in the proximal tubule cells before and after cisplatin both in vivo and in vitro for its inactivation or its removal from the membrane using light and electron microscopy.Outbred male Swiss Webster (Crl: (WI) BR) rats weighing 150-250g were given ip injections of cisplatin (7mg/kg). Animals were killed on day 3 and day 5. Thick slices (20-50.um) of kidney tissue from treated and untreated animals were fixed in 1% buffered glutaraldehyde and 1% formaldehyde (0.05 M cacodylate buffer, pH 7.3) for 30 min at 4°C. Alkaline phosphatase activity and carbohydrates were demonstrated according to methods described earlier.

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Correction to Nucleotide Excision Repair of Chemically Stabilized Analogues of DNA Interstrand Cross-Links Produced from Oxidized Abasic Sites

Biochemistry ◽

10.1021/bi501162v ◽

2014 ◽

Vol 53 (40) ◽

pp. 6418-6418

Author(s):

Souradyuti Ghosh ◽

Marc M. Greenberg

Keyword(s):

Nucleotide Excision Repair ◽

Excision Repair ◽

Abasic Sites ◽

Nucleotide Excision ◽

Interstrand Cross Links ◽

Cross Links

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DNA interstrand cross-links from modified nucleotides: mechanism and application

Nucleic Acids Symposium Series ◽

10.1093/nass/49.1.57 ◽

2005 ◽

Vol 49 (1) ◽

pp. 57-58 ◽

Cited By ~ 3

Author(s):

Marc M. Greenberg

Keyword(s):

Modified Nucleotides ◽

Interstrand Cross Links ◽

Cross Links

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Increased gene-specific repair of cisplatin interstrand cross-links in cisplatin-resistant human ovarian cancer cell lines

Molecular and Cellular Biology ◽

10.1128/mcb.12.9.3689-3698.1992 ◽

1992 ◽

Vol 12 (9) ◽

pp. 3689-3698

Author(s):

W Zhen ◽

C J Link ◽

P M O'Connor ◽

E Reed ◽

R Parker ◽

...

Keyword(s):

Cell Lines ◽

Ovarian Cancer Cell ◽

Parental Cell ◽

Resistant Cell ◽

Dhfr Gene ◽

Human Ovarian Cancer Cell ◽

Ovarian Cancer Cell Lines ◽

Repair Efficiency ◽

Interstrand Cross Links ◽

Cross Links

We have studied several aspects of DNA damage formation and repair in human ovarian cancer cell lines which have become resistant to cisplatin through continued exposure to the anticancer drug. The resistant cell lines A2780/cp70 and 2008/c13*5.25 were compared with their respective parental cell lines, A2780 and 2008. Cells in culture were treated with cisplatin, and the two main DNA lesions formed, intrastrand adducts and interstrand cross-links, were quantitated before and after repair incubation. This quantitation was done for total genomic lesions and at the level of individual genes. In the overall genome, the initial frequency of both cisplatin lesions assayed was higher in the parental than in the derivative resistant cell lines. Nonetheless, the total genomic repair of each of these lesions was not increased in the resistant cells. These differences in initial lesion frequency between parental and resistant cell lines were not observed at the gene level. Resistant and parental cells had similar initial frequencies of intrastrand adducts and interstrand cross-links in the dihydrofolate reductase (DHFR) gene and in several other genes after cisplatin treatment of the cells. There was no increase in the repair efficiency of intrastrand adducts in the DHFR gene in resistant cell lines compared with the parental partners. However, a marked and consistent repair difference between parental and resistant cells was observed for the gene-specific repair of cisplatin interstrand cross-links. DNA interstrand cross-links were removed from three genes, the DHFR, multidrug resistance (MDR1), and delta-globin genes, much more efficiently in the resistant cell lines than in the parental cell lines. Our findings suggest that acquired cellular resistance to cisplatin may be associated with increased gene-specific DNA repair efficiency of a specific lesion, the interstrand cross-link.

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