The svpA-srtB locus of Listeria monocytogenes: fur-mediated iron regulation and effect on virulence.
The svpA-srtB locus of Listeria monocytogenes: fur-mediated iron regulation and effect on virulence.
复制标题
单核细胞增生李斯特氏菌的 svpA-srtB 基因座：毛皮介导的铁调节及其对毒力的影响。
DOI：
10.1111/j.1365-2958.2004.04436.x
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发表时间：
2005
影响因子：
3.6
通讯作者：
Charbit,Alain
中科院分区：
文献类型：
--
作者：
Newton,SaleteMC;Klebba,PhillipE;Raynaud,Catherine;Shao,Yi;Jiang,Xiaoxu;Dubail,Iharilalao;Archer,Crystal;Frehel,Claude;Charbit,Alain
在单核细胞增生李斯特菌中，vpA ‐srtBlocus的启动子区包含一个高度保守的Fur盒。我们表征了该位点的铁调节：真实的时间聚合酶链反应分析和抗SvpA免疫印迹显示，当通过向BHI培养基中添加铁螯合剂2，2 ′-联吡啶或通过在铁限制的基本培养基中生长来启动时，作为对铁剥夺的响应，svpA转录和SvpA产生显著增加（分别为80倍和10倍）。当细菌在缺铁条件下生长时，绿色荧光蛋白（GFP）报告基因构建体也显示大肠杆菌（37倍）和单核细胞增生李斯特菌（2 - 3倍）中vpA ‐ srtB启动子的活性增加。单核细胞增生李斯特菌的Δfurmutant组成型合成SvpA，以及融合到vpA ‐ srtB启动子的GFP。细胞分级分离数据显示，在富铁培养基中，野生型SvpA仅分泌到培养上清液中。然而，在缺铁培养基中生长的Δfurderivative和野生型单核细胞增生李斯特菌都将一部分SvpA蛋白（约5%）锚定在肽聚糖上，并产生较低分子量的完全分泌形式的SvpA。 这些数据共同确定了铁的可利用性控制了thesvpA‐srtBlocus的转录（通过Fur介导的调节），以及SvpA与细胞壁的附着（通过SrtB介导的共价键）。SvpA与金黄色葡萄球菌的血红素结合蛋白IsdC具有同源性，并且血红素在溶液中与SvpA结合。然而，四个结构基因和thesvpA‐srtBlocus的启动子的定点缺失并没有损害营养测试中的血红素、血红蛋白或铁色素的利用。我们没有发现强有力的证据来支持vpA ‐srtBlocus参与血红素获得的观点，就像报道的金黄色葡萄球菌的同源物doperon一样。 此外，vpA ‐ srtB突变株在静脉注射小鼠模型系统中没有显示出明显的毒力衰减，但我们发现突变降低了口服给药后小鼠肝脏、脾脏和肠道中单核细胞增生李斯特菌的持久性。
InListeria monocytogenesthe promoter region of thesvpA‐srtBlocus contains a well‐conserved Fur box. We characterized the iron‐regulation of this locus: real‐time polymerase chain reaction analyses and anti‐SvpA immunoblots showed that, in response to iron deprivationsvpAtranscription and SvpA production markedly increased (80‐fold and 10‐fold respectively), when initiated by either the addition of the iron chelator 2,2′‐bipyridyl to BHI media, or by growth in iron‐restricted minimal media. Green fluorescent protein (GFP) reporter constructs also showed increased activity of thesvpA‐srtBpromoter inEscherichia coli (37‐fold) and inL. monocytogenes(two‐ to threefold) when the bacteria were grown in iron‐deficient conditions. A Δfurmutant ofL. monocytogenesconstitutively synthesized SvpA, as well as GFP fused to thesvpA‐srtBpromoter. Cellular fractionation data revealed that in iron‐rich media wild‐type SvpA was exclusively secreted to the culture supernatant. However, both the Δfurderivative and wild‐typeL. monocytogenesgrown in iron‐deficient media anchored a fraction of the SvpA proteins (∼5%) to peptidoglycan, and produced a lower‐molecular weight, wholly secreted form of SvpA. Together these data establish that iron availability controls transcription of thesvpA‐srtBlocus (through Fur‐mediated regulation), and attachment of SvpA to the cell wall (through SrtB‐mediated covalent linkage). SvpA bears homology to IsdC, a haemin‐binding protein ofStaphylococcus aureus,and haemin bound to SvpA in solution. However, site‐directed deletions of four structural genes and the promoter of thesvpA‐srtBlocus did not impair haemin, haemoglobin or ferrichrome utilization in nutrition tests. We did not find strong evidence to support the notion that thesvpA‐srtBlocus participates in haemin acquisition, as was reported for the homologousisdoperon ofS. aureus. Furthermore, thesvpA‐srtBmutant strains showed no significant attenuation of virulence in an intravenous mouse model system, but we found that the mutations reduced the persistence ofL. monocytogenesin murine liver, spleen and intestines after oral administration.