Journal List > J Bacteriol Virol > v.46(3) > 1034212

Lim, Choi, Kim, and Seo: Transcriptional Analysis of the iagB within Salmonella Pathogenicity Island 1 (SPI1)

Abstract

HilA is a central regulator of Salmonella pathogenicity island 1 (SPI1), which is necessary for host invasion by Salmonella and induction of gastroenteritis. The iagB lies downstream of hilA and is thought to be co-transcribed with hilA, but iagB expression has not yet been analyzed directly. In this study, iagB expression in various mutant strains was measured to determine whether the expression pattern was similar to that of hilA. A β-galactosidase assay revealed that iagB expression was greater under shaking than standing culture condition. iagB expression was decreased in relA/ spoT and ihfB mutants but not in luxS mutant, in line with previous reports on hilA expression. The hilA and iagB mRNA levels decreased by approximately 2-fold in arcA mutant grown aerobically and increased by approximately 10-fold in fnr mutant grown anaerobically. Although the fold changes in hilA and iagB mRNA level differed in hfq mutant strain, the patterns of timeand Hfq-dependent regulation were similar for both genes. Thus, iagB and hilA exhibited similar expression patterns in various mutational backgrounds and under different growth condition.

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Figure 1.
Transcriptional analysis of iagB under different growth conditions. The S. Typhimurium strain harboring a chromosomal iagB:: lacZY fusion was grown in LB medium under shaking (●) or standing conditions (○). The expression level was determined by measuring the β-galactosidase activity (Miller units) at the indicated times. Data are presented as the mean ± standard error of three independent experiments performed in duplicate.
jbv-46-128f1.tif
Figure 2.
Transcriptional analysis of iagB in different mutant strains. The β-galactosidase activities (Miller units) of chromosomal iagB::lacZY were measured in ΔluxS (A), ΔppGpp (ΔrelAspoT)(B), and Δ ihfB (C) strains at 4 h p.i. (early stationary phase) and 12 h p.i. (late stationary phase) under shaking and standing culture conditions, respectively. Data are presented as the mean ± standard error of three independent experiments performed in duplicate.
jbv-46-128f2.tif
Figure 3.
Transcriptional analysis of hilA and iagB in arcA and fnr mutants. The mRNA levels of hilA and iagB were determined by qRT-PCR. Total RNA was isolated from strains grown for 4 h under shaking and standing culture conditions. The values for the relative expression were determined by defining the mRNA levels from the wild-type strain as 1. The expression levels of the target genes were normalized to 16S rRNA gene. Data are presented as the mean ± standard error of three independent experiments performed in duplicate.
jbv-46-128f3.tif
Figure 4.
Transcriptional analysis of hilA and iagB in hfq mutants. The mRNA levels of hilA and iagB were determined by qRT-PCR. Total RNA was isolated from strains grown for 4 h and 12 h under shaking culture conditions. The values for the relative expression were determined by defining the mRNA levels from the wild-type strain as 1. The expression levels of the target genes were normalized to 16S rRNA gene. Data are presented as the mean ± standard error of three independent experiments performed in duplicate.
jbv-46-128f4.tif
Table 1.
The bacterial strains
Strains Description Reference or source
SL1344 wild type serovar Typhimurium xyl rpsL hisG Lab stock
CS669 14028s iagB:: lacZY, Kan R 21
SR3306 SL1344 Δ luxS 25
JE3999 TR6538 ihfB:: cat, Cam R 35
SHJ2037 14028s relA:: kan, spoT:: cat, Kan R, Cam R 26
NC983 14028s fnr::Tn10, Tet R 29
SY1001 SL1344 iagB:: lacZY, Kan R This study
SY1002 SL1344 Δ luxS, iagB:: lacZY, Kan R This study
SY1004 SL1344 relA:: kan, spoT:: cat, iagB::lacZY, Kan R, Cam R This study
SY1005 SL1344 ihfB:: cat, iagB:: lacZY, Kan R, Cam R This study
SR3560 SL1344 Δ arcA 22
SY1006 SL1344 fnr::Tn10, Tet R 22
SY1101 SL1344 Δ hfq 22
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