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DNA Repair (Amst) ; 5(1): 129-37, 2006 Jan 05.
Article in English | MEDLINE | ID: mdl-16257588

ABSTRACT

Lesions that transiently block DNA synthesis generate replication intermediates with recombinogenic potential. In order to investigate the mechanisms involved in lesion-induced recombination, we developed an homologous recombination assay involving the transfer of genetic information from a plasmid donor molecule to the Escherichia coli chromosome. The replication blocking lesion used in the present assay is formed by covalent binding of the carcinogen N-2-acetylaminofluorene to the C8 position of guanine residues (G-AAF adducts). The frequency of recombination events was monitored as a function of the number of lesions present on the donor plasmid. These DNA adducts are found to trigger high levels of homologous recombination events in a dose-dependent manner. Formation of recombinants is entirely RecA-dependent, the RecF and RecBCD sub-pathways accounting for about 2/3 and 1/3, respectively. Inactivation of recG stimulates recombinant formation about five-fold. In a recG background, the RecF pathway is stimulated about four-fold, while the contribution of the RecBCD pathway remains constant. In addition, in the recG strain, a recombination pathway that accounts for about 30% of the recombinants and requires genes that belong to both RecF and RecBCD pathways is revealed.


Subject(s)
DNA-Binding Proteins/metabolism , Escherichia coli Proteins/metabolism , Escherichia coli/genetics , Recombination, Genetic , 2-Acetylaminofluorene/pharmacology , DNA Damage/genetics , DNA-Binding Proteins/genetics , Escherichia coli/drug effects , Escherichia coli Proteins/genetics , Exodeoxyribonuclease V/genetics , Exodeoxyribonuclease V/metabolism , Gene Expression Regulation, Bacterial , Guanine/metabolism , Plasmids/genetics , Rec A Recombinases/genetics , Rec A Recombinases/metabolism , Signal Transduction
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