密度感应系统调节细菌应答反应的发生,这些应答反应与细胞密度有关。通过对比大肠杆菌(Escherichia coli)和志贺氏菌(Shigella spp.)的序列发现,志贺菌属密度感应系统操纵子普遍存在丢失或突变。为研究其密度感应系统的功能,文章利用哈氏弧菌(Vibrio harveyi)BB170作为指示菌,检测弗氏志贺菌(Shigella flexneri)密度感应系统信号分子AI-2,证明其可以分泌有活性的AI-2;其次,采用Golden Gate克隆法将大肠杆菌MG1655的密度感应系统基因克隆至弗氏志贺菌301中,获得密度感应系统回复株301。通过菌落计数表明,在混合培养条件下,密度感应系统基因回复株301比野生株301存在生长优势;通过双向电泳初步比较分析表明,密度感应系统基因可以在志贺菌中表达,并鉴定到了其他一些与应激反应相关的差异表达蛋白, 如Hsp60、GroEL、SodB。
Quorum sensing (QS) regulates the onset of bacterial social responses related to cell density. Comparison between the gene sequences of all components of QS system of Escherichia coli and Shigella strains, shows that the QS system is generally lost or mutated in Shigella. Since AI-2 is produced and processed by the lsr operon, we analyzed the potential function of the lsr operon. We first detected AI-2 in Shigella flexneri 2a strain 301 through the reporter bacteria Vibrio harveyi BB170, indicating that S. flexneri can produce AI-2. Then, the lsr operon of E. coli MG1655 was cloned into S. flexneri using the Golden Gate method. Colony counting experiments showed that the QS system recovery strain had growth advantage over the wild-type strain when they were mixed and cultured. The preliminary comparative proteomics analysis showed that the lsr operon could be expressed and the abundance of stress response proteins also changed when the QS system was introduced into S. flexneri.
[1] El-Sayed AK, Hothersall J, Thomas CM. Quorum-sensing- dependent regulation of biosynthesis of the polyketide antibiotic mupirocin in Pseudomonas fluorescens NCIMB 10586. Microbiology , 2001, 147(8): 2127-2139.
[2] Diggle SP, Crusz SA, Cámara M. Quorum sensing. Curr Bio , 2007, 17(21): R907-R910.
[3] Williams P, Winzer K, Chan WC, Cámara M. Look who's talking: communication and quorum sensing in the bacterial world. Philos Trans R Soc Lond B Biol Sci , 2007, 362(1483): 1119-1134.
[4] Miller MB, Bassler BL. Quorum sensing in bacteria. Annu Rev Microbiol , 2001, 55: 165-199.
[5] Erickson DL, Endersby R, Kirkham A, Stuber K, Vollman DD, Rabin HR, Mitchell I, Storey DG . Pseudomonas aeruginos a quorum-sensing systems may control virulence factor expression in the lungs of patients with cystic fibrosis. Infect Immun , 2002, 70(4): 1783-1790.
[6] Queck SY, Jameson-Lee M, Villaruz AE, Bach THL, Khan BA, Sturdevant DE, Ricklefs SM, Li M, Otto M . RNAIII-independent target gene control by the agr quorum-sensing system: insight into the evolution of virulence regulation in Staphylococcus aureus . Mol Cell , 2008, 32(1): 150-158.
[7] Choi J, Shin D, Ryu S. Implication of quorum sensing in Salmonella enterica serovar typhimurium virulence: the luxS gene is necessary for expression of genes in pathogenicity island. Infect Immun , 2007, 75(10): 4885-4890.
[8] Brito PH, Rocha EPC, Xavier KB, Gordo I . Natural genome diversity of AI-2 quorum sensing in Escherichia coli : conserved signal production but labile signal reception. Genome Biol Evol , 2013, 5(1): 16-30.
[9] Xue T, Zhao LP, Sun HP, Zhou XX, Sun BL. LsrR-binding site recognition and regulatory characteristics in Escherichia coli AI-2 quorum sensing. Cell Res , 2009, 19(11): 1258-1268.
[10] González Barrios AF, Zuo RJ, Hashimoto Y, Yang L, Bentley WE, Wood TK. Autoinducer 2 controls biofilm formation in Eschrichia coli through a novel motility quorum-sensing regulator (MqsR, B3022). J Bacteriol , 2006, 188(1): 305-316.
[11] Herzberg M, Kaye IK, Peti W, Wood TK . YdgG (TqsA) controls biofilm formation in Escherichia coli K-12 through autoinducer 2 transport. J Bacteriol , 2006, 188(2): 587-598.
[12] Bansal T, Jesudhasan P, Pillai S, Wood TK, Jayaraman A. Temporal regulation of enterohemorrhagic Escherichia coli virulence mediated by autoinducer-2. Appl Microbiol Biotechnol , 2008, 78(5): 811-819.
[13] Hegde M, Englert DL, Schrock S, Cohn WB, Vogt C, Wood TK, Manson MD, Jayaraman A . Chemotaxis to the quorum-sensing signal AI-2 requires the Tsr chemoreceptor and the periplasmic LsrB AI-2-binding protein. J Bacteriol , 2011, 193(3): 768-773.
[14] Taga M E, Xavier KB. Methods for analysis of bacterial autoinducer-2 production. Curr Protoc Microbiol , 2005, Chapter 1: Unit1C.1.
[15] Engler C, Kandzia R, Marillonnet S. A one pot, one step, precision cloning method with high throughput capability. PLoS One, 2008, 3(11): e3647.
[16] Xavier KB, Bassler BL. Regulation of uptake and processing of the quorum-sensing autoinducer-2 in Escherichia coli . J Bacteriol , 2005, 187(1): 238-248.
[17] Diggle SP, Gardner A, West SA, Griffin AS. Evolutionary theory of bacterial quorum sensing: when is a signal not a signal? Philos Trans R Soc Lond B Biol Sci , 2007, 362(1483): 1241-1249.
[18] Winzer K, Hardie KR, Williams P. Bacterial cell-to-cell communication: sorry, can't talk now—gone to lunch! Curr Opin Microbiol , 2002, 5(2): 216-222.
