Listeria monocytogenes σB regulates stress response and virulence functions
Listeria monocytogenes σB regulates stress response and virulence functions
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DOI：
10.1128/jb.185.19.5722-5734.2003
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发表时间：
2003-10-01
影响因子：
3.2
通讯作者：
Wiedmann, M
中科院分区：
文献类型：
--
作者：
Kazmierczak, MJ;Mithoe, SC;Wiedmann, M
虽然已经在芽孢杆菌属、李斯特菌属和葡萄球菌属的不同物种中鉴定了应激响应性替代σ因子σ（B），但σ（B）调节子仅在B中被广泛表征。枯草芽孢杆菌我们结合生物计算和微阵列为基础的战略，以确定西格玛（B）依赖的基因在兼性细胞内病原体单核细胞增生李斯特菌。基于隐马尔可夫模型（HMM）的搜索在菌株EGD-e基因组序列中鉴定了170个候选σ（B）依赖性启动子序列。这些数据被用来开发一个专门的，208个基因的微阵列，其中包括166个基因的HMM预测的西格玛（B）依赖性启动子的下游，以及选定的毒力和应激反应基因。从野生型和DeltasigB无效突变体L.单核细胞增多性细胞生长至稳定期或暴露于渗透胁迫（0.5 M KCl）。微阵列分析鉴定了在这些实验中使用的条件下具有统计学显著的sigma（B）依赖性表达的总共55个基因，在任一胁迫条件下野生型中的表达比sigB突变体高至少1.5倍（51个基因在野生型中显示出至少高2.0倍的表达）。在55个表现出sigma（B）依赖性表达的基因中，54个之前有类似于sigma（B）启动子共有序列的序列。cDNA末端的快速扩增-PCR用于确认8个所选启动子区域的子集的sigma（B）依赖性。值得注意的是，sigma（B）依赖的L.通过这种HMM/微阵列策略鉴定的单核细胞增多症基因包括两种应激反应基因（例如，gadB，ctc，和谷胱甘肽还原酶基因lmo 1433）和毒力基因（例如，inlA、inlB和bsh）。我们的数据表明，除了调节在环境胁迫条件下生存的重要基因的表达外，sigma（B）还有助于调节L.单核细胞增多症。这些发现强烈地表明，σ（B）对L有贡献。单核细胞增生基因表达。
While the stress-responsive alternative sigma factor sigma(B) has been identified in different species of Bacillus, Listeria, and Staphylococcus, the sigma(B) regulon has been extensively characterized only in B. subtilis. We combined biocomputing and microarray-based strategies to identify sigma(B)-dependent genes in the facultative intracellular pathogen Listeria monocytogenes. Hidden Markov model (HMM)-based searches identified 170 candidate sigma(B)-dependent promoter sequences in the strain EGD-e genome sequence. These data were used to develop a specialized, 208-gene microarray, which included 166 genes downstream of HMM-predicted sigma(B)-dependent promoters as well as selected virulence and stress response genes. RNA for the microarray experiments was isolated from both wild-type and DeltasigB null mutant L. monocytogenes cells grown to stationary phase or exposed to osmotic stress (0.5 M KCl). Microarray analyses identified a total of 55 genes with statistically significant sigma(B)-dependent expression under the conditions used in these experiments, with at least 1.5-fold-higher expression in the wild type over the sigB mutant under either stress condition (51 genes showed at least 2.0-fold-higher expression in the wild type). Of the 55 genes exhibiting sigma(B)-dependent expression, 54 were preceded by a sequence resembling the sigma(B) promoter consensus sequence. Rapid amplification of cDNA ends-PCR was used to confirm the sigma(B)-dependent nature of a subset of eight selected promoter regions. Notably, the sigma(B)-dependent L. monocytogenes genes identified through this HMM/microarray strategy included both stress response genes (e.g., gadB, ctc, and the glutathione reductase gene lmo1433) and virulence genes (e.g., inlA, inlB, and bsh). Our data demonstrate that, in addition to regulating expression of genes important for survival under environmental stress conditions, sigma(B) also contributes to regulation of virulence gene expression in L. monocytogenes. These findings strongly suggest that sigma(B) contributes to L. monocytogenes gene expression during infection.