Epithelial specific ubiquitination events

描述（由申请人提供）：肠道炎症反应在很大程度上由分泌的细胞因子和其他炎症效应分子的合成上调介导。这些炎症介质在转录水平上通过DNA结合转录因子如NF-κ B的作用而被激活。NF-kB本身通过快速翻译后途径激活，该途径涉及抑制剂分子IkB的连续磷酸化、泛素化和降解。肠上皮细胞在组织中是独特的，其功能与多种原核植物群密切接触。人们越来越关注这些细菌对宿主上皮细胞的有害和有益的影响。在这个提议中，我们描述了肠道细菌菌株，通过影响限速生化步骤，泛素化抑制关键NF-κ B调节剂的激活。泛素化是一种可诱导的共价修饰，在蛋白质周转和降解中具有明确的作用，并且在信号转导、内吞运输和其他细胞功能中具有新兴作用。细菌及其产物可能影响负责IkB泛素化的酶和参与上皮信号传导的潜在其他蛋白质的活化所必需的共价修饰。在其他工作中，我们已经确定了肠道病原体中的候选效应分子，它们是NFkB的有效抑制剂，并可能作为去泛素化酶。我们假设这些蛋白质将对真核细胞的促炎通路和细胞凋亡的控制产生深远的影响。 我们的总体假设是，肠道细菌，肠道和已知的病原体的寄生虫，已经进化出的机制，通过影响真核泛素系统来调节上皮细胞的信号通路。根据特定的生物学背景，这些相互作用的后果可能是感染性和特发性结肠炎的原因，和/或可能有助于肠道的生理学和整体健康。该提案描述了各种体外和体内实验方法来研究细菌如何影响肠上皮细胞中的泛素化系统，特别是当它涉及NF-κ B和凋亡途径的激活时。阐明这些宿主-微生物相互作用将增加我们对人类小肠结肠炎中涉及的上皮和细菌因素的理解，并可能在特发性炎症性肠病中。

DESCRIPTION (provided by applicant): The intestinal inflammatory response is mediated in large part by synthetic upregulation of secreted cytokines and other inflammatory effector molecules. These inflammatory mediators are activated at the transcriptional level by the action of DNA binding transcription factors such as NF-kB. NF-kB itself is activated by a rapid post translational pathway involving sequential phosphorylation, ubiquitination and degradation of the inhibitor molecule, IkB. Unique among tissues, the intestinal epithelia functions in close contact with a diverse prokaryotic flora. There is increasing interest in the effects, both deleterious and beneficial, of these bacteria on the host epithelia. In this proposal, we describe enteric commensal bacterial strains that repress activation of the key NF-kB regulator by influencing a rate-limiting biochemical step, ubiquitination. Ubiquitination is an inducible covalent modification with a well defined role in protein turnover and degradation, and emerging roles in signal transduction, endocytic trafficking and other cellular functions. Bacteria and their products may influence covalent modifications necessary for activation of the enzymes responsible for ubiquitination of IkB and potentially other proteins involved in epithelial signaling. In other work, we have identified candidate effector molecules in enteric pathogens that are potent inhibitors of NFkB and likely act as deubiquitinating enzymes. We hypothesize these proteins will have profound effects on eukaryotic proinflammatory pathways and control of cellular apoptosis. Our overall hypothesis is that enteric bacteria, both commensal inhabitants of the intestine and known pathogens, have evolved mechanisms to modulate epithelial signaling pathways by influencing the eukaryotic ubiquitin system. Depending the specific biological context, the consequences of these interactions may by causal of infectious and idiopathic colitis, and/or may contribute to the physiology and overall health of the intestine. This proposal describes a variety of in vitro and in vivo experimental methods to study how bacteria influence the ubiquitination system in the intestinal epithelia, especially as it relates to the activation of the NF-kB and apoptotic pathways. Elucidation of these host-microbe interactions will increase our understanding of the epithelial and bacterial factors involved in human enterocolitis, and potentially in idiopathic inflammatory intestinal disease.