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PMID: 11333215 Published · ppublish English Journal Article Research Support, U.S. Gov't, P.H.S.

Repair of topoisomerase-mediated DNA damage in bacteriophage T4.

Genetics ·Vol. 158 ·No. 1 ·2001-05-00 ·Pages 19-28

Stohr BA, Kreuzer KN

Abstract

Type II topoisomerase inhibitors are used to treat both tumors and bacterial infections. These inhibitors stabilize covalent DNA-topoisomerase cleavage complexes that ultimately cause lethal DNA damage. A functional recombinational repair apparatus decreases sensitivity to these drugs, suggesting that topoisomerase-mediated DNA damage is amenable to such repair. Using a bacteriophage T4 model system, we have developed a novel in vivo plasmid-based assay that allows physical analysis of the repair products from one particular topoisomerase cleavage site. We show that the antitumor agent 4'-(9-acridinylamino)methanesulphon-m-anisidide (m-AMSA) stabilizes the T4 type II topoisomerase at the strong topoisomerase cleavage site on the plasmid, thereby stimulating recombinational repair. The resulting m-AMSA-dependent repair products do not form in the absence of functional topoisomerase and appear at lower drug concentrations with a drug-hypersensitive topoisomerase mutant. The appearance of repair products requires that the plasmid contain a T4 origin of replication. Finally, genetic analyses demonstrate that repair product formation is absolutely dependent on genes 32 and 46, largely dependent on genes uvsX and uvsY, and only partly dependent on gene 49. Very similar genetic requirements are observed for repair of endonuclease-generated double-strand breaks, suggesting mechanistic similarity between the two repair pathways.

MeSH Terms
Amsacrine/pharmacology Bacteriophage T4/genetics Base Sequence DNA Damage DNA Primers DNA Repair DNA Topoisomerases, Type II/metabolism DNA, Viral/drug effects,genetics Mutation Plasmids Recombination, Genetic
Chemicals
DNA Primers DNA, Viral Amsacrine DNA Topoisomerases, Type II
Authors & Affiliations
2 authors, click to expand affiliations / ORCID
Stohr B A
Departments of Microbiology and Biochemistry, Duke University Medical Center, Durham, NC 27710, USA.
Kreuzer K N
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Article Info
Journal
Genetics
Abbr.
Genetics
ISSN
0016-6731
DOI
10.1093/genetics/158.1.19
Published
2001-05-00
Pages
19-28
Language
English
Region
United States
NLM ID
0374636
PMCID
PMC1461630
Subset
IM
Grants
NCI NIH HHS · CA60836 · United States
NIGMS NIH HHS · T32-GM07171-26 · United States
External Links
PubMed source DOI: 10.1093/genetics/158.1.19 PMC Full Text View journal details
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