Control of bacteriophage mu lysogenic repression

The transposable and temperate phage Mu infects Escherichia coli where it can enter the lytic life-cycle or reside as a repressed and integrated prophage. The repressor protein Rep is the key element in the lysis-lysogeny decision. We have analyzed the fate of Rep in different mutants by Western blotting under two conditions that can induce a lysogen: high temperature and stationary phase. We show that, unexpectedly, Rep accumulates under all conditions where the prophage is completely derepressed, and that this accumulation is ClpX-dependent. An analysis of the degradation kinetics shows that Rep is a target of two protease systems: inactivation of either the clpP or lon gene results in a stabilization of Rep. Such a reaction scheme explains the counterintuitive observation that derepression is correlated with high repressor concentration. We conclude that under all conditions of phage induction the repressor is sequestered in a non-active form. A quantitative simulation accounts for our experimental data. It provides a model that captures the essential features of Mu induction and explains some of the mechanisms by which the physiological signals affecting the lysis-lysogeny decision converge onto Rep.     

The Pasteurella multocida toxin is encoded within a lysogenic bacteriophage

Toxigenic strains of Pasteurella multocida produce a 146 kDa toxin (PMT) that acts as a potent mitogen. Sequence analysis of the structural gene for PMT, toxA, previously suggested it was horizontally acquired, because it had a low G + C content relative to the P. multocida genome. To address this, the sequence of DNA flanking toxA was determined. The sequence analysis showed the presence of homologues to bacteriophage tail protein genes and a bacteriophage antirepressor, suggesting that the toxin gene resides within a prophage. In addition to phage genes, the toxA flanking DNA contained a homologue of a restriction/modification system that was shown to be functional. The presence of a bacteriophage was demonstrated in spent medium from toxigenic P. multocida isolates. Its production was increased by mitomycin C addition, a treatment that is known to induce the lytic cycle of many temperate bacteriophages. The genomes of bacteriophages from three different toxigenic P. multocida strains had similar but not identical restriction profiles, and were approximately 45-50 kb in length. The prophages from two of these had integrated at the same site in the chromosome, in a tRNA gene. Southern blot analysis confirmed that these bacteriophages contained the toxA gene.     

Biochemical changes in lysogenic Bacillus stearothermophilus after bacteriophage induction

Welker, N. E. (University of Illinois, Urbana), and L. Leon Campbell. Biochemical changes in lysogenic Bacillus stearothermophilus after bacteriophage induction. J. Bacteriol. 90:1129-1137. 1965.-Cultures of Bacillus stearothermophilus 1503-4R (TP-1) continued to grow at an unaltered rate after induction with mitomycin C (MC). MC-induced cultures exhibited a 2.5-fold increase in cell number before lysis occurred. Prior to lysis, cells were observed to elongate and to contain areas of lesser density. Protein synthesis was slightly inhibited in MC- or ultraviolet light (UV)-induced cultures for a period of 5 to 10 min, and then proceeded at a rate identical to that in the noninduced culture. Ribonucleic acid (RNA) synthesis was not affected by MC induction. UV induction caused RNA synthesis to occur in two stages: in the first stage, the rate of RNA synthesis was one-third that observed in the noninduced culture and lasted for a period of 15 min; the second stage of RNA synthesis then proceeded at a rate identical to that in the noninduced culture. The synthesis of deoxyribonucleic acid (DNA) in an MC- or UV-induced culture occurred in two stages. In the first stage, DNA synthesis in induced cultures occurred at a rate of one-half (MC) and one-third (UV) of that observed in the noninduced culture. The first stage of DNA synthesis in MC- or UV-induced cultures lasted for 25 to 30 min and 15 to 20 min, respectively. In the second stage, the rate of DNA synthesis in MC- or UV-induced cultures occurred at a rate three times that of the noninduced culture. UV induction appeared to have a greater inhibitory effect than MC induction on protein, RNA, and DNA synthesis as well as phage yield. The differential rate (K) of inducible and constitutive alpha-amylase synthesis was inhibited by 75 and 100%, respectively, for a period of 20 min after MC induction. After 20 min, the K values for alpha-amylase synthesis were identical to those obtained in the absence of MC induction. The synthesis of TP-1 phage DNA occurred rapidly and was complete 25 min after MC induction, whereas bacterial DNA was degraded or its rate of synthesis was decreased. During the second stage of DNA synthesis, only bacterial DNA was synthesized, but at a rate greater than that found in the noninduced culture.     

Infection of spheroplasts of Pseudomonas with DNA of bacteriophage PM2

Summary Spheroplasts of Pseudomonas BAL-31/PM2, obtained by treatment of the bacteria with lysozyme, can be infected with purified DNA from bacteriophage PM2. After 4 h of incubation the yield of progeny phage reaches a value of 10⁷-6×10⁷ plaque forming units/g PM2 DNA. The yield increases linearly with the concentration of DNA over at least 3 orders of magnitude.The biological activity of double-stranded circular PM2 DNA containing one or more single-strand breaks per molecule (component II), does not differ significantly from that of intact PM2 DNA (component I). Single-stranded PM2 DNA obtained by denaturation of component II, and the irreversible alkali-denatured form of component I are also infective.     

X-ray sensitivity of Escherichia coli lysogenic for bacteriophage P2.

Summary Strains of Escherichia coli C or K lysogenic for the non-inducible phage P2 show a lower survival following X-ray irradiation as compared to nonlysogenic strains. This difference in X-ray sensitivity is not accompanied by a significant difference in X-ray induced mutability. The capacity of X-irradiated P2 lysogens to multiply any of a number of unirradiated infecting phages is severely impaired. These effects of X-ray treatment can be most simply explained as a consequence of the fact that protein and RNA syntheses are strongly inhibited in P2 lysogens after X-irradiation. All the above events specifically occurring in X-rayed P2 lysogens are dependent on the P2 gene old.     

Quantitative study of sensitive and lysogenic bacteria surviving infection of aStaphylococcus by a temperate bacteriophage

Infection of a culture ofStaphylococcus aureus by a phage left more surviving colony-forming, sensitive bacteria than might be expected from the calculation of P_o according to the Poisson distribution. The discrepancy could at least partly be explained by the heterogeneity in surface area of the colony-forming units before infection. The ratio lysogens/lysed cells shows a tendency to decrease when multiplicity is increased from 6 to 130.The experiments stress the influence of the bacterial density on the adsorption rate constant of the bacteriophage.Single step growth curve experiments gave low burst size values. The latent period was about 30 min. The stationary period began 45 to 50 min after infection.