Mutations altering genetic recombination and repair of DNA in bacteriophage T4

Abstract

A search was made for sublethal mutants of bacteriophage T4 defective in genetic recombination. T4D + was mutagenized with proflavin, and phage from minute plaques were tested for their sensitivity to ultraviolet (uv) irradiation. Nine uv-sensitive mutants were found, of which two exhibit decreased recombination. The mutations are located in a previously unknown T4 gene-gene w. Seven other minute-plaque-forming, uv sensitive mutants exhibit increased recombination; in each of these seven cases, the mutations were found to be in gene 58 (DNA-delay). The newly located gene w maps between genes 24 and 25. Mutants defective in gene w are sensitive to uv, to methyl methanesulfonate, and to hydroxyurea, although they degrade host DNA normally. Incorporation of [ 3H]thymidine into DNA is normal. A delayed and reduced phage yield appears to be caused by inefficient packaging of DNA in w mutants. This inefficient packaging appears to be caused by failure to form or to maintain a normal complement of concatemeric DNA. Intracellular DNA in 58 − infection was examined for unusual properties which could explain increased recombination. The normal rapidly sedimenting complex containing concatemeric DNA is present, but is formed more slowly than in wild-type infection. Joining of short DNA chains produced by discontinuous synthesis is normal, as is attachment of parental and newly synthesized DNA to a rapidly-sedimenting cell component (presumably membrane). Parental DNA is degraded, apparently by an exonuclease. Both the production of acid-soluble material from parental DNA and the increase in genetic recombination are eliminated by additional mutations in genes 46 and 47. Genetic evidence indicates that both 58 and w mutants are defective in the same uv repair pathway as the uv sensitive T4 mutants x and y; none of the double mutants is more sensitive than each of the respective single mutants. Some of these double mutants, however, have cumulative effects on recombination, indicating a more complicated interaction than a discrete unilinear pathway. The x and y strains currently available were found to contain additional mutations. The original x strain contains several mutations which affect plaque morphology. The original y strain, y 10, contains a mutation which affects plaque morphology, and the y 100 strain contains a mutation which causes the y defect to be lethal. The purified x and y strains make minute plaques and retain the reduced burst size, lowered recombination, uv sensitivity of the originally described mutants.