Enzymatic degradation of uracil-containing deoxyribonucleic acid. V. Survival of Escherichia coli and coliphages treated with sodium bisulfite.

A number of mutants of Escherichia coli defective in the ung gene (structural gene for uracil-deoxyribonucleic acid [ura-DNA] glycosylase) are shown to be abnormally sensitive to treatment with sodium bisulfite when compared with congenic ung+ strains. These results provide further evidence that sodium bisulfite causes the deamination of cytosine to uracil in DNA and that ura-DNA glycosylase is required for the repair of U-G mispairs. The effect of the chemical is apparently selective with respect to base damage; coliphages containing cytosine in their DNA are inactivated by treatment with sodium bisulfite, whereas those containing hydroxymethylcytosine are not. ura-DNA glycosylase and the major apurinic-apyrimidinic endonuclease of E. coli may function in the same repair pathway, since the extent of inactivation of a congenic set of strains which are ung xth (structural gene for the major apurinic-apyrimidinic endonuclease of E. coli) or ung xth+ is the same.     

tif-Stimulated deoxyribonucleic acid repair in Escherichia coli K-12

Bacterial survival is significantly increased after ultraviolet irradiation in tif sfi cells, provided that the thermosensitive tif mutation has been expressed at 41 degrees C before irradiation. This tif-mediated "reactivation of ultraviolet irradiated bacteria" needs de novo protein synthesis, as is the case for the tif-mediated reactivation of ultraviolet-irradiated phage lambda. However, in striking contrast to the phage reactivation process, this tif-mediated reactivation is no longer associated with mutagenesis. It also requires the presence of the uvrA+ excision function. These results strongly suggest the existence in Escherichia coli K-12 of a repair pathway acting on bacterial deoxyribonucleic acid which is inducible, error free, and uvr dependent.     

Nature of transforming deoxyribonucleic acid in calcium-treated Escherichia coli.

A study of the reactivation of ultraviolet-irradiated plasmid and phage deoxyribonucleic acid molecules after transformation into Escherichia coli strains indicated that, when double-stranded deoxyribonucleic acid was used as the donor species, it was taken up without conversion to the single-standed form.     

Conditional-lethal deoxyribonucleic acid ligase mutant of Escherichia coli.

A new Escherichia coli deoxyribonucleic acid (DNA) ligase mutant has been identified among a collection of temperature-sensitive DNA replication mutants isolated recently (Sevastopoulos, Wehr, and Glaser, Proc. Natl. Acad. Sci. U.S.A. 74:3485-3489, 1977). At the nonpermissive temperature DNA synthesis in the mutant stops rapidly, the DNA is degraded to acid-soluble material, and cell death ensures. This suggests that the mutant may be among the most ligase-deficient strains yet characterized.     

Medium-dependent variation of deoxyribonucleic acid segregation in Escherichia coli.

The degree to which deoxyribonucleic acid segregates nonrandomly has been investigated for Escherichia coli B/r growing in different media. The degree of nonrandom segregation observed is dependent on the medium, with segregation becoming less random as the growth rate decreases. This indicates that there must be some varying probabilistic component to the segregation process. A probabilistic modification of the Pierucci-Zuchowski model is proposed as well as a probabilistic model, in which it is proposed that deoxyribonucleic acid strands segregate, with a probability greater than 0.5, in the same direction (toward the same pole) as at the previous cell division.     

Genetics of host-controlled restriction and modification of deoxyribonucleic acid in Escherichia coli

Lederberg, Seymour (Brown University, Providence, R.I.). Genetics of host-controlled restriction and modification of deoxyribonucleic acid in Escherichia coli. J. Bacteriol. 91:1029-1036. 1966.-The locus for the host specific restriction and modification of deoxyribonucleic acid in Escherichia coli has been mapped by matings between mutants for these characters in strains K-12, C600, and B. Linkage analysis and kinetics of marker transfer indicate that a single or closely linked multiple chromosomal site located about 4 min counterclockwise to leucine is responsible for these activities. Secondary factors which affect the quantitative level of restriction also were detected. Wild-type recombinants were isolated in crosses between rm(-) (restriction or modification, or both) mutants. The expression in zygotes of the restrictionless character of a rm(-) donor is masked by a separate, physiological impairment of restriction, which results from mating and is independent of the modification state of the donor. The relevance of the restriction character to mating incompatibilities in these and other bacterial strains is considered.     

Kinetics of bacteriophage lambda deoxyribonucleic acid infection of Escherichia coli

Barnhart, Benjamin J. (Los Alamos Scientific Laboratory, University of California, Los Alamos, N.M.). Kinetics of bacteriophage lambda deoxyribonucleic acid infection of Escherichia coli. J. Bacteriol. 90:1617-1623. 1965.-The kinetics of Escherichia coli K-12 infection by phage lambda deoxyribonucleic acid (DNA) were determined. An initial lag of 55 to 80 sec was found to be the time required for infecting DNA to become deoxyribonuclease-insensitive at 33 C. When cell-DNA interactions were stopped by washing away unbound DNA, the already bound DNA continued to infect the cell at rates described by linear kinetics with no apparent lag. Whereas the lag period was relatively insensitive to DNA and cell concentrations, both the lag and the subsequent linear portions of the rate curves were temperature-sensitive. Cell and DNA dose-response curves prescribed hyperbolic functions. Similarities between lambda DNA infection of E. coli and bacterial transformation systems are discussed.     

Temperature-sensitive recA mutant of Escherichia coli K-12: deoxyribonucleic acid metabolism after ultraviolet irradiation.

A mutant of Escherichia coli K-12 temperature sensitive for genetic recombination was investigated and found to carry a mutation that could be cotransduced with cysC and hence could be in the recA gene. To determine whether recA+ can complement this mutation, matings were carried out at 35 and 40 C between Hfr donors that transfer recA+ or recA1 early and recipients carrying wild-type or mutant alleles. It was found that recA+ but not recA1 complements this mutation in zygotic temporary partial diploids. The mutant allele was accordingly designated recA44. A transductant carrying recA44 behaved normally at low temperatures but more like recA- strains at high temperatures with respect to recombinant colony formation in Hfr matings, cell survival, and deoxyribonucleic acid (DNA) synthesis after ultraviolet irradiation, cellular DNA breakdown, and prophage induction when lysogenic for lambda. Alkaline sucrose sedimentation studies of DNA from recA44 cells showed that short DNA molecules synthesized immediately after ultraviolet irradiation increased in molecular weight during subsequent incubation at 32 C but not at 45 C. Hence, recA+ is required for this molecular weight increase. Cells exposed to ultraviolet light synthesized DNA that remained of low molecular weight during a 40-min incubation at 32 C. This material increased in molecular weight in recArut not in recA44 cells during subsequent incubation at 45 C. Thus, the availability of recA+ during the first 40 min at 32 C after irradiation did not obviate the need for recA+ in the subsequent phases of this post-replication repair process.     

Penicillin selection of Escherichia coli deoxyribonucleic acid repair mutants.

A rapid method has been developed for isolation of ultraviolet-sensitive mutants of Escherichia coli, by inducing delay in the growth and/or division of repair-deficienct cells with low fluences of far-ultraviolet radiation, and killing with penicillin the repair-proficient cells, which continue to grow and divide. With this technique, we have achieved about a 3,000-fold enrichment for photoreactivation less (phr) cells and have isolated and characterized three phr mutants.     

Conditional-lethal deoxyribonucleic acid polymerase I mutant of Escherichia coli.

A new deoxyribonucleic acid polymerase I mutant of Escherichia coli was isolated among conditional lethal mutants. Deoxyribonucleic acid replication in the mutant ceased in 20 min after the temperature was raised to 43 degrees C, and reinitiated when cells were further incubated at this temperature.     

Rifampin disrupts conjugal and chromosomal deoxyribonucleic acid metabolism in Escherichia coli K-12 carrying some IncIalpha plasmids.

The effects of rifampin and chloramphenicol on the transfer of ColIdrd-1 have been examined to determined whether transfer requires the synthesis of an untranslated species of ribonucleic acid (RNA), as proposed in models for the transfer of another IncIalpha plasmid, R64drd-11. When RNA synthesis was inhibited throughout mating by rifampin, ColI transfer between dna+ bacteria occurred at the normal rate for about 10 min and then stopped abruptly. Conjugational deoxyribonucleic acid (DNA) synthesis in dnaB mutants indicates that plasmid DNA was made in the rifampin-treated donors to replace the transferred material but the DNA tended to be unstable. In the presence of chloramphenicol, transfer of ColI gradually diminished over a longer period. Rifampin, but not chloramphenicol, was found to have unpredicted effects on chromosomal DNA metabolism in unmated dna+ and dnaB bacteria when they harbor any of three IncIalpha plasmids (ColIdrd-1, R144drd-3, and R64drd-11). Replication of the bacterial chromosome in such cells stopped abruptly about 15 min after the addition of rifampin, and at 41 degrees C, but not 37 degrees C, this was followed by extensive DNA breakdown. These findings suggest that curtailment of IncIalpha plasmid transfer by the drug results from a general disruption of DNA metabolism rather than from inhibition of a species of RNA essential for transfer.