Type III restriction is alleviated by bacteriophage (RecE) homologous recombination function but enhanced by bacterial (RecBCD) function.
Previous works have demonstrated that DNA breaks generated by restriction enzymes stimulate, and are repaired by, homologous recombination with an intact, homologous DNA region through the function of lambdoid bacteriophages lambda and Rac. In the present work, we examined the effect of bacteriophage functions, expressed in bacterial cells, on restriction of an infecting tester phage in a simple plaque formation assay. The efficiency of plaque formation on an Escherichia coli host carrying EcoRI, a type II restriction system, is not increased by the presence of Rac prophage-presumably because, under the single-infection conditions of the plaque assay, a broken phage DNA cannot find a homologue with which to recombine. To our surprise, however, we found that the efficiency of plaque formation in the presence of a type III restriction system, EcoP1 or EcoP15, is increased by the bacteriophage-mediated homologous recombination functions recE and recT of Rac prophage. This type III restriction alleviation does not depend on lar on Rac, unlike type I restriction alleviation. On the other hand, bacterial RecBCD-homologous recombination function enhances type III restriction. These results led us to hypothesize that the action of type III restriction enzymes takes place on replicated or replicating DNA in vivo and leaves daughter DNAs with breaks at nonallelic sites, that bacteriophage-mediated homologous recombination reconstitutes an intact DNA from them, and that RecBCD exonuclease blocks this repair by degradation from the restriction breaks.
Handa
Naofumi
N
Laboratory of Social Genome Sciences, Department of Medical Genome Sciences, Graduate School of Frontier Science and Institute of Medical Science, University of Tokyo, 4-6-1 Shirokanedai, Minato-ku, Tokyo 108-8639, Japan.
Kobayashi
Ichizo
I
D010582:Bacteriophage lambda / Q000235:genetics* / Q000502:physiology; D001435:Bacteriophages / Q000235:genetics* / Q000502:physiology; D004260:DNA Repair / Q000502:physiology; D004269:DNA, Bacterial / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism; D015246:Deoxyribonuclease EcoRI / Q000378:metabolism; D015263:Deoxyribonucleases, Type III Site-Specific / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics*; D029968:Escherichia coli Proteins / Q000378:metabolism*; D043211:Exodeoxyribonuclease V / Q000378:metabolism*; D005090:Exodeoxyribonucleases / Q000378:metabolism*; D008780:Methyltransferases / Q000378:metabolism; D039002:Prophages / Q000235:genetics* / Q000502:physiology; D011995:Recombination, Genetic; D010948:Viral Plaque Assay
D004269:DNA, Bacterial; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; C084497:RecT protein, E coli; D008780:Methyltransferases; D005090:Exodeoxyribonucleases; C038225:recE protein, E coli; D043211:Exodeoxyribonuclease V; C468322:exodeoxyribonuclease V, E coli; D015246:Deoxyribonuclease EcoRI; C030432:endodeoxyribonuclease EcoP1; C023451:endodeoxyribonuclease EcoP15I; D015263:Deoxyribonucleases, Type III Site-Specific