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. 2005 Feb;187(4):1485-92.
doi: 10.1128/JB.187.4.1485-1492.2005.

Host gene expression changes and DNA amplification during temperate phage induction

Affiliations

Host gene expression changes and DNA amplification during temperate phage induction

Jonathan G Frye et al. J Bacteriol. 2005 Feb.

Abstract

Salmonella enterica serovar Typhimurium LT2 harbors four temperate prophages. The lytic cycle of these phages was induced with hydrogen peroxide or mitomycin C. Microarray analysis was used to monitor the increase in phage genome copy number and the changes in RNA expression. Phage gene transcription was classified temporally, and host genes that responded to hydrogen peroxide, mitomycin C, or phage induction were also identified. A region of the serovar Typhimurium LT2 host genome encompassing hundreds of genes, flanking the Fels-1 lambdoid prophage, was amplified manyfold during lytic induction, presumably due to Fels-1 runoff replication prior to excision, a phenomenon termed escape replication. An excisionase (xis) mutant of Fels-1 also induced escape replication but did not get packaged. Gifsy-1, a lambdoid prophage that does not normally produce escape replication, did so after deletion of either its integrase or excisionase genes. Escape replication is probably widespread; large regions of host genome amplification were also observed after phage induction in serovar Typhimurium strains SL1344 and 14028s at the suspected integration site of prophage genomes.

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Figures

FIG. 1.
FIG. 1.
Serovar Typhimurium strains after treatment with 2 mM hydrogen peroxide or 2 μg of mitomycin C per ml. The y axis indicates the ratio of the DNA content of treated cells to the DNA content of cells before treatment on a log10 scale. The x axis indicates the S. enterica serovar Typhimurium LT2 genes in order of position on the chromosome (blue), the pSLT genes (pink), and select CT18 phage genes (green). The origin (Ori) and terminus (Ter) of replication are indicated. (A) LT2 1 h after treatment with peroxide; (B) LT2 2 h aftertreatment with peroxide; (C) LT2 3 h after treatment with peroxide; (D) LT2 Fels-1 xis mutant JF105 (STM0894::kan) 3 h after treatment with mitomycin C; (E) LT2 Gifsy-1 xis and int double mutant JF109 (STM2635-STM2636::kan) 3 h after treatment with mitomycin C.
FIG. 2.
FIG. 2.
Fels phage particles released after mitomycin C treatment. LT2 cells growing in Luria-Bertani medium were treated with mitomycin C for 5 h. Subsequently, cells were separated from the supernatant, and phage were harvested from the supernatant fraction by PEG precipitation. DNA was isolated from both the cells and the supernatants. (A) LT2 cells 5 h after treatment with mitomycin C. The y axis indicates the ratio of the DNA content of treated cells to the DNA content of cells before treatment on a log10 scale. (B) Phage DNA harvested from LT2 supernatants 5 h after treatment with mitomycin C. The y axis indicates the contribution of every gene to the total signal of all genes represented on the microarray (expressed as percentages).
FIG. 3.
FIG. 3.
Serovar Typhimurium strains SL1344 and 14028s after treatment with 2 mM peroxide. The y axis indicates the ratio of DNA content 3 h after treatment to the DNA content of cells before treatment on a log10 scale. The x axis indicates the S. enterica serovar Typhimurium genes in order of position on the chromosome. (A) SL1344; (B) 14028s.
FIG. 4.
FIG. 4.
Gene expression profile of lambdoid prophage Gifsy-1 after mitomycin C treatment. Numbers are from the STM annotation, including some homologues in Gifsy-2 (16). Names are hypothetical, based on homology with other phages. The expression ratios are the ratios compared with an untreated control and are log2 transformed. Red, increase; green, decrease; black, no change. (A) Genes listed in order of position on the serovar Typhimurium chromosome. (B) Hierarchical clustering of phage gene expression as determined by GENESIS. Putative gene expression cluster assignments are indicated. Cluster E, early; cluster DE, delayed early; cluster M, middle; cluster L, late; rep, replication.

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