Epithelial specific ubiquitination events

描述（申请人提供）：肠道炎症反应在很大程度上是通过合成上调分泌的细胞因子和其他炎症效应分子来介导的。这些炎症介质在转录水平上通过DNA结合转录因子（如NF-kB）的作用被激活。NF-kB本身通过一个快速的翻译后途径被激活，该途径涉及抑制剂分子IkB的顺序磷酸化、泛素化和降解。独特的组织，肠上皮功能密切接触的各种原核菌群。这些细菌对宿主上皮细胞的有害和有益的影响越来越引起人们的兴趣。在这个提议中，我们描述了肠道共生细菌菌株，通过影响限速生化步骤，泛素化，抑制关键NF-kB调节因子的激活。泛素化是一种可诱导的共价修饰，在蛋白质转换和降解中具有明确的作用，并在信号转导、内吞运输和其他细胞功能中发挥着新的作用。细菌及其产物可能影响IkB泛素化酶和潜在的其他参与上皮信号传导的蛋白质激活所必需的共价修饰。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，可能起着去泛素化酶的作用。我们推测这些蛋白将对真核促炎途径和细胞凋亡的控制有深远的影响。

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.