NF-kB /Beta-catenin interaction during intestinal inflammation

描述（申请人提供）：越来越多的证据表明，内源性肠道细菌在炎症性肠病（IBDs）的发病机制中起着至关重要的作用。很明显，肠道菌群有助于提高活化NF-水平。在IBD患者肠黏膜中然而，目前尚不清楚是哪种菌株和/或细菌？产品有助于IBD的发病和/或维持。由于肠道菌群的复杂性，IBD中特定致病微生物的鉴定仍然具有挑战性，并且寻找致病微生物的工作非常激烈。AvrA是某些细菌的致病基因，其编码蛋白通过III型分泌途径插入肠上皮细胞。在之前的研究中，我们证明了AvrA可以稳定？-catenin，它是NF-??的负调节因子上皮细胞通路。我们假设细菌效应物AvrA抑制NF-?？途径并激活？-catenin途径抑制肠道炎症。在这项研究中，我将重点研究AvrA的机制和作用，采用体外和体内两种方法。对于体外研究，我们将用avra充足或缺乏的细菌菌株定殖上皮细胞。我们将确定细菌效应物AvrA实际上是如何调节NF-的？-catenin结合和鉴定NF-调控因子转录后的变化然后呢?AvrA诱导的-catenin通路（如磷酸化、泛素化）。我们将研究AvrA在NF-??的下游靶点（即IL-8， c-myc, cyclin D1）中诱导的表达变化。和? ?连环蛋白通路。在体内研究中，我们将使用发生自发性结肠炎的IL-10-/-小鼠，这已被证明需要结肠细菌定植才能完全表达。这些IL-10-/-小鼠最初将在无菌环境中饲养，然后与大肠杆菌F18或大肠杆菌F18AvrA+单相关，以确定AvrA在体内抑制炎症的功能。阐明AvrA调控NF-??的机制然后呢?-catenin信号通路将为细菌-宿主相互作用如何促进炎症提供新的见解。这些研究与更好地理解炎症性肠病和感染性结肠炎等疾病直接相关。

DESCRIPTION (provided by applicant): Increasing evidence indicates that endogenous enteric bacteria play a crucial role in the pathogenesis of inflammatory bowel diseases (IBDs). It is clear that gut flora contribute to the elevated levels of activated NF-???in the intestinal mucosa of IBD patients. However, it is unclear which bacterial strain and/or bacterial ?products contribute to the pathogenesis and/or maintenance of IBD. Due to the complexity of the gut flora, identification of the specific causative microbial agents in IBD remains challenging, and the search for the causative microbial agents is intense. AvrA is a pathogenic gene of certain bacteria whose encoded protein is inserted into intestinal epithelial cells through a Type III secretory pathway. In previous studies, we demonstrated that AvrA stabilizes ?-catenin, which is a negative regulator of the NF-?? pathway in epithelial cells. We hypothesize that the bacterial effector AvrA inhibits the NF-???pathway and actives the ?-catenin pathway to inhibit intestinal inflammation. In this study, I will focus on the mechanism and effects of AvrA, using both in vitro and in vivo approaches. For in vitro studies, we will colonize epithelial cells with AvrA-sufficient or -deficient bacteria strains. We will determine how the bacterial effector AvrA actually regulates NF-??/?-catenin binding and identify post-transcriptional changes in regulators of the NF-?? and ?-catenin pathways induced by AvrA (e.g., phosphorylations, ubiquitinations). We will investigate expression changes induced by AvrA in downstream targets (i.e., IL-8, c-myc, cyclin D1) of the NF-?? and???-catenin pathways. For in vivo studies, we will employ IL-10-/- mice that develop spontaneous colitis, which has been shown to require colonic bacterial colonization for full expression. These IL-10-/- mice will initially be raised in germ-free environments, and will then be mono-associated with E.coli F18 or E.coli F18AvrA+ to determine the function of AvrA in inhibiting inflammation in vivo. Elucidating the mechanisms of AvrA regulating the NF-?? and ?-catenin signaling pathways will provide new insights into how bacteria-host interactions contribute to inflammation. These studies have direct relevance to a better understanding of diseases such as inflammatory bowel diseases and infectious colitis. Lay language: This proposal is aimed at understanding the mechanism and effects of the bacterial protein AvrA in inhibiting inflammation. We will focus on the mechanism and effects of AvrA in cultured cells colonized with bacterial strains (with or without AvrA expression) and in a mouse colitis model. The information obtained from our research will provide new insights into the way bacteria-host interactions contribute to inflammation. This has direct relevance to a better understanding of diseases such as inflammatory bowel diseases and infectious colitis.