Activation of the eis gene in a W-Beijing strain of Mycobacterium tuberculosis correlates with increased SigA levels and enhanced intracellular growth.
Activation of the eis gene in a W-Beijing strain of Mycobacterium tuberculosis correlates with increased SigA levels and enhanced intracellular growth.
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结核分枝杆菌 W-Beijing 菌株中 eis 基因的激活与 SigA 水平升高和细胞内生长增强相关。
DOI：
10.1099/mic.0.024638-0
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发表时间：
2009
期刊：
影响因子：
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通讯作者：
Howard,SusanT
中科院分区：
文献类型：
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作者：
Wu,Shiping;Barnes,PeterF;Samten,Buka;Pang,Xiuhua;Rodrigue,Sébastien;Ghanny,Saleena;Soteropoulos,Patricia;Gaudreau,Luc;Howard,SusanT
越来越多的证据表明结核分枝杆菌菌株在致病性和传播性方面存在差异，但对其影响因素知之甚少。我们之前已经表明，主要西格玛因子 SigA 表达的增加介导了 M 的能力。与其他菌株相比，结核菌株 210 在人类单核细胞中生长得更快。菌株 210 是广泛分布的 W-北京 M 家族的一部分。结核菌株并包括临床分离株 TB294。为了鉴定对 SigA 水平变化作出反应并可能增强细胞内生长的基因，我们检查了 TB294-pSigA 中的 RNA 和蛋白质表达模式，TB294-pSigA 是 TB294 的重组菌株，可从多拷贝质粒过度表达 sigA。 TB294-pSigA 肉汤培养物的裂解物含有高水平的 Eis，这是一种已知可调节宿主与病原体相互作用的蛋白质。 DNA微阵列分析表明，在人单核细胞系MonoMac6中生长期间，TB294-pSigA中的eisgene Rv2416c的表达水平比载体对照菌株TB294-pCV高40倍。 TB294-pSigA 中表达升高的其他基因与 TB294-pCV 相比变化要小得多，并且两个菌株之间表达差异的大多数基因在 TB294-pSigA 中的表达降低，包括与 DNA 损伤反应相关的大量基因。实时PCR分析证实eis在单核细胞和肉汤培养物中的TB294-pSigA中以非常高的水平表达，并进一步揭示，与sigA一样，在单核细胞中生长期间，eis在野生型TB294中的表达也比在实验室菌株H37Rv中更高。这些发现表明 SigA 水平升高与去激活之间存在关联，并且染色质免疫沉淀结果证实 SigA 与活 TB294 细胞中的 eis 启动子结合。缺失ei会减少单核细胞中TB294的生长，而补充ei则会逆转这种效应。我们得出结论，SigA调节seis，SigA的上调和ofeis的高表达水平之间存在直接相关性，并且这有助于增强M的临床分离株的能力。结核菌株210在单核细胞中生长。
There is growing evidence that strains ofMycobacterium tuberculosisdiffer in pathogenicity and transmissibility, but little is understood about the contributory factors. We have previously shown that increased expression of the principal sigma factor, SigA, mediates the capacity ofM. tuberculosisstrain 210 to grow more rapidly in human monocytes, compared with other strains. Strain 210 is part of the widespread W-Beijing family ofM. tuberculosisstrains and includes clinical isolate TB294. To identify genes that respond to changes in SigA levels and that might enhance intracellular growth, we examined RNA and protein expression patterns in TB294-pSigA, a recombinant strain of TB294 that overexpressessigAfrom a multicopy plasmid. Lysates from broth-grown cultures of TB294-pSigA contained high levels of Eis, a protein known to modulate host–pathogen interactions. DNA microarray analysis indicated that theeisgene, Rv2416c, was expressed at levels in TB294-pSigA 40-fold higher than in the vector control strain TB294-pCV, during growth in the human monocyte cell line MonoMac6. Other genes with elevated expression in TB294-pSigA showed much smaller changes from TB294-pCV, and the majority of genes with expression differences between the two strains had reduced expression in TB294-pSigA, including an unexpected number of genes associated with the DNA-damage response. Real-time PCR analyses confirmed thateiswas expressed at very high levels in TB294-pSigA in monocytes as well as in broth culture, and further revealed that, likesigA,eiswas also more highly expressed in wild-type TB294 than in the laboratory strain H37Rv, during growth in monocytes. These findings suggested an association between increased SigA levels andeisactivation, and results of chromatin immunoprecipitation confirmed that SigA binds theeispromoter in live TB294 cells. Deletion ofeisreduced growth of TB294 in monocytes, and complementation ofeisreversed this effect. We conclude that SigA regulateseis, that there is a direct correlation between upregulation of SigA and high expression levels ofeis, and thateiscontributes to the enhanced capacity of a clinical isolate ofM. tuberculosisstrain 210 to grow in monocytes.