Quantitative Proteomic Analysis of Type III Secretome of Enteropathogenic Escherichia coli Reveals an Expanded Effector Repertoire for Attaching/Effacing Bacterial Pathogens
Quantitative Proteomic Analysis of Type III Secretome of Enteropathogenic Escherichia coli Reveals an Expanded Effector Repertoire for Attaching/Effacing Bacterial Pathogens
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DOI：
10.1074/mcp.m111.013672
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发表时间：
2012-09-01
影响因子：
7
通讯作者：
Finlay, B. Brett
中科院分区：
文献类型：
--
作者：
Deng, Wanyin;Yu, Hong B.;Finlay, B. Brett
III型分泌系统是许多重要的革兰氏阴性细菌病原体的发病机制和毒力的核心，并且阐明分泌机制和鉴定分泌底物对于我们理解其致病机制和开发潜在的治疗方法至关重要。在基于细胞培养的质谱中用氨基酸进行稳定同位素标记是一种定量和高灵敏度的蛋白质组学工具，我们以前曾成功地用于分析啮齿柠檬酸杆菌和鼠伤寒沙门氏菌的III型分泌体。本研究采用稳定同位素标记技术，对致病性大肠杆菌（EPEC）的III型分泌酶组进行了分析，并与肠出血性大肠杆菌（E. coli和C.啮齿类，代表附着和消除细菌病原体的家族。我们不仅证实了所有25个已知的EPEC III型分泌的蛋白和效应物，以前确定的传统的分子和生物信息学技术，但也确定了几个新的III型分泌的蛋白，包括两个新的效应物，C_0814/NleJ和LifA，被证明是易位到宿主细胞。LifA是一种已知的毒力因子，被认为是一种毒素和粘附素，但其分泌和功能机制尚不清楚。LifA的预测分子量为366 kDa，是迄今为止在任何病原体中鉴定的最大的III型效应物。我们进一步证明了Efa 1、ToxB和Z4332（肠出血性大肠杆菌中LifA的同源物）。大肠杆菌）也是III型效应子。本研究全面表征了EPEC的III型分泌组，扩展了III型分泌的附着和消除病原体的效应子库，并为LifA/Efa 1/ToxB/Z4332，一个重要的毒力因子家族的功能模式提供了新的见解。Molecular & Cellular Proteomics 11：10.1074/mcp. M111.013672，692-709，2012.
Type III secretion systems are central to the pathogenesis and virulence of many important Gram-negative bacterial pathogens, and elucidation of the secretion mechanism and identification of the secreted substrates are critical to our understanding of their pathogenic mechanisms and developing potential therapeutics. Stable isotope labeling with amino acids in cell culture-based mass spectrometry is a quantitative and highly sensitive proteomics tool that we have previously used to successfully analyze the type III secretomes of Citrobacter rodentium and Salmonella enterica serovar Typhimurium. In this report, stable isotope labeling with amino acids in cell culture was used to analyze the type III secretome of enteropathogenic Escherichia coli (EPEC), an important human pathogen, which, together with enterohemorrhagic E. coli and C. rodentium, represents the family of attaching and effacing bacterial pathogens. We not only confirmed all 25 known EPEC type III-secreted proteins and effectors previously identified by conventional molecular and bioinformatical techniques but also identified several new type III-secreted proteins, including two novel effectors, C_0814/NleJ and LifA, that were shown to be translocated into host cells. LifA is a known virulence factor believed to act as a toxin as well as an adhesin, but its mechanism of secretion and function is not understood. With a predicted molecular mass of 366 kDa, LifA is the largest type III effector identified thus far in any pathogen. We further demonstrated that Efa1, ToxB, and Z4332 (homologs of LifA in enterohemorrhagic E. coli) are also type III effectors. This study has comprehensively characterized the type III secretome of EPEC, expanded the repertoire of type III-secreted effectors for the attaching and effacing pathogens, and provided new insights into the mode of function for LifA/Efa1/ToxB/Z4332, an important family of virulence factors. Molecular & Cellular Proteomics 11:10.1074/mcp.M111.013672, 692-709, 2012.