肠上皮将肠道内微生物和肠腔外组织分隔开，它既能耐受肠道共生菌群，又能对致病菌产生免疫应答。但目前我们对肠上皮区分共生菌和致病菌的机制还很不清楚。许多病菌通过三型分泌系统(TTSS)分泌的效应蛋白来阻断宿主功能。我们此前曾报道肠致病大肠杆菌(EPEC)等相关病菌分泌抑制宿z主NF-κB激活的效应蛋白，推测EPEC感染可能诱导NF-κB激活，需要效应蛋白来抑制。初步结果表明EPEC的TTSS通过两种平行且相互独立的途径激活NF-κB。本研究旨在揭示这两种途径的分子机制：1，阐明上皮细胞感知TTSS激活NF-κB的机制；2，查明哪些EPEC效应蛋白能激活NF-κB并明确其作用机制。本研究揭示的新机制，对发展预防和治疗炎性肠病的新策略至关重要，也对了解AE致病菌的感染策略、开发治疗这些感染的新方法具有指导作用，包括EHEC O157:H7导致的溶血性尿毒综合症—西方国家引起儿童肾功能衰竭的主要原因。

The intestinal epithelium separates the vast microbiota from deeper tissue. It should practice tolerance toward apical microbiota, but initiate an inflammatory response upon detection of pathogens. The processes by which epithelial cells differentiate pathogens from commensal bacteria.remain poorly understood. Many pathogens employ a type III secretion system (TTSS) to deliver into infected cells proteins termed effectors that subvert host cell functions. We previously reported that enteropathogenic E. coli (EPEC) and related pathogens produce effectors that inhibit the NF-κB signaling pathway. We hypothesize that EPEC employ these effectors to antagonize NF-κB activation occurred upon infection. Our preliminary results support this prediction and imply that two independent and parallel processes lead NF-κB activation in cells infected with EPEC lacking the anti-NF-κB effectors. The first process involves detection of the TTSS of the infecting bacteria by epithelial cells, leading to NF-κB activation. The second process involves delivery into the host cell of a TTSS effector whose function is to activate NF-κB. We propose to elucidate the two ways epithelial cells respond to EPEC through NF-κB activation by addressing two specific aims. i) to elucidate the TTSS-sensing mechanism of epithelial cells and to reveal how it leads to NF-κB activation, and ii) to identify and characterize the putative EPEC effector(s) that trigger NF-κB activation. This study may expose new basic rules that govern pathogen-commensal discrimination. This knowledge is essential for developing new strategies for prevention and treatment of inflammatory bowel diseases. In addition, the proposed project will also shed light on the principles guiding the infection strategy of EPEC and related pathogens, thus contributing to the development of new approaches to treat infection by these pathogens, including EHEC O157:H7, which can cause the devastating Hemolytic Uremic Syndrome (HUS), the most common cause of pediatric renal failure in western countries.

肠致病性大肠杆菌感染是婴儿腹泻性肠炎的主要原因，在发展中国家有很高的风险导致婴儿死亡。肠致病性大肠杆菌主要感染小肠上皮细胞的顶面，并在那儿定值。肠致病性大肠杆菌通过它的三型分泌系统向宿主上皮细胞中注入多个效应蛋白分子，这些效应分子帮助细菌粘附在上皮细胞的表面并定值。我们的研究发现肠致病性大肠杆菌在感染的宿主细胞中可以引起非常强的NF-kB转录因子的激活，进而诱导强烈的炎症反应。我们进一步研究发现，三型分泌系统在宿主细胞表面的打孔活性本身对诱导NF-kB炎症通路的激活既是必要的，也是充分的，并且这种激活不依赖于三型分泌系统对任何效应蛋白的转运。但重要的是，三型分泌系统本身导致的NF-kB炎症通路的激活会被其分泌的NleB, NleC 和NleE效应蛋白的活性所抑制。因此，只有当细菌中的这三个效应蛋白基因敲除后，我们才能观察到三型分泌系统本身所诱导的NF-kB炎症通路的激活。通过深入的机制研究，我们发现三型分泌系统本身激活NF-kB炎症信号是通过一个非常独特的通路而实现的，该通路不依赖于已知的Toll样和NOD样受体，但同其他已知的NF-kB炎症信号通路一样，也是汇集在TAK1激酶激活这一步。总之， 我们的研究揭示肠上皮细胞具有感知三型分泌系统进而激活NF-kB炎症信号的固有能力，这种能力可以帮助宿主区分是致病菌和非致病菌感染。另外，本项目的研究还发现三型分泌还可以分泌细菌代谢小分子，包括细菌的七炭糖分子，这些细菌特异性的分子也可以激活NF-kB炎症信号通路，这种全新的细菌感知模式是由宿主上皮细胞中的一个特异性的激酶所实现的。