Sperandio V, Torres A G, Girón J A, Kaper J B
Center for Vaccine Development and Department of Microbiology and Immunology, University of Maryland School of Medicine, Baltimore, Maryland 21201, USA.
J Bacteriol. 2001 Sep;183(17):5187-97. doi: 10.1128/JB.183.17.5187-5197.2001.
Enterohemorrhagic Escherichia coli (EHEC) O157:H7 is responsible for outbreaks of bloody diarrhea and hemolytic-uremic syndrome in many countries. EHEC virulence mechanisms include the production of Shiga toxins (Stx) and formation of attaching and effacing (AE) lesions on intestinal epithelial cells. We recently reported that genes involved in the formation of the AE lesion were regulated by quorum sensing through autoinducer-2, which is synthesized by the product of the luxS gene. In this study we hybridized an E. coli gene array with cDNA synthesized from RNA that was extracted from EHEC strain 86-24 and its isogenic luxS mutant. We observed that 404 genes were regulated by luxS at least fivefold, which comprises approximately 10% of the array genes; 235 of these genes were up-regulated and 169 were down-regulated in the wild-type strain compared to in the luxS mutant. Down-regulated genes included several involved in cell division, as well as ribosomal and tRNA genes. Consistent with this pattern of gene expression, the luxS mutant grows faster than the wild-type strain (generation times of 37.5 and 60 min, respectively, in Dulbecco modified Eagle medium). Up-regulated genes included several involved in the expression and assembly of flagella, motility, and chemotaxis. Using operon::lacZ fusions to class I, II, and III flagellar genes, we were able to confirm this transcriptional regulation. We also observed fewer flagella by Western blotting and electron microscopy and decreased motility halos in semisolid agar in the luxS mutant. The average swimming speeds for the wild-type strain and the luxS mutant are 12.5 and 6.6 microm/s, respectively. We also observed an increase in the production of Stx due to quorum sensing. Genes encoding Stx, which are transcribed along with lambda-like phage genes, are induced by an SOS response, and genes involved in the SOS response were also regulated by quorum sensing. These results indicate that quorum sensing is a global regulatory mechanism for basic physiological functions of E. coli as well as for virulence factors.
肠出血性大肠杆菌(EHEC)O157:H7在许多国家引发了血性腹泻和溶血尿毒综合征的暴发。EHEC的毒力机制包括志贺毒素(Stx)的产生以及在肠道上皮细胞上形成紧密黏附并抹去(AE)损伤。我们最近报道,参与AE损伤形成的基因受群体感应调控,通过由luxS基因产物合成的自诱导物-2来实现。在本研究中,我们用从EHEC菌株86 - 24及其同基因luxS突变体提取的RNA合成的cDNA与大肠杆菌基因芯片进行杂交。我们观察到404个基因受luxS调控至少5倍,约占芯片基因的10%;与luxS突变体相比,野生型菌株中这些基因有235个上调,169个下调。下调的基因包括一些参与细胞分裂的基因以及核糖体和tRNA基因。与这种基因表达模式一致,luxS突变体比野生型菌株生长得更快(在杜尔贝科改良伊格尔培养基中,代时分别为37.5分钟和60分钟)。上调的基因包括一些参与鞭毛表达与组装、运动性和趋化性的基因。使用操纵子::lacZ融合到I类、II类和III类鞭毛基因,我们能够证实这种转录调控。通过蛋白质免疫印迹和电子显微镜观察,我们还发现luxS突变体中的鞭毛较少,并且在半固体琼脂中的运动晕圈减小。野生型菌株和luxS突变体的平均游动速度分别为12.5和6.6微米/秒。我们还观察到由于群体感应导致Stx产量增加。编码Stx的基因与类λ噬菌体基因一起转录,由SOS反应诱导,并且参与SOS反应的基因也受群体感应调控。这些结果表明群体感应是大肠杆菌基本生理功能以及毒力因子的一种全局调控机制。