SphK1-TRAF2-cIAP interactions in NF-kB activation, colitis and associated cancer

描述（由申请人提供）：结直肠癌是最致命的癌症之一，也是发达国家的主要死亡原因。不幸的是，对于许多患有慢性肠道炎症的人来说，结肠炎与结肠炎相关癌（CAC）的发病机制之间存在着密切的联系。最具特征的CAC促进因子之一是促炎细胞因子TNF-¿及其下游主转录因子NF- B。因此，针对NF-？B活化途径作为一种治疗或预防CAC进展的手段越来越受到研究人员的欢迎。最近，我们的实验室发现生物活性鞘脂鞘鞘醇-1-磷酸（S1P）和产生它的酶鞘鞘醇激酶1 （SphK1）连接持久性NF-？B活化对慢性肠道炎症和CAC的发展。S1P和SphK1一直与TNF-¿的作用有关。在典型的tnf依赖性细胞内信号级联导致NF-？B激活，一系列复杂的蛋白质相互作用和翻译后修饰调节信号传播并最终导致基因转录。其中，受体相互作用激酶-1、TNF受体相关因子2 （TRAF2）和细胞凋亡抑制剂1和2 （cIAP1/2）发挥重要作用。然而，SphK1参与NF-？B通路仍不清楚。重要的是，SphK1和TRAF2已被证明在CAC患者组织以及该疾病动物模型的结肠和肿瘤中升高。我的初步结果表明，SphK1、TRAF2和cIAP1/2之间的蛋白-蛋白相互作用对它们的细胞稳定很重要。因此，本提案将检验的假设是SphK1-TRAF2-cIAP1/2复合物在NF-？B通过蛋白质与蛋白质的直接相互作用来激活，调节每种蛋白质的稳定性、活性和功能。我还认为，SphK1-TRAF2-cIAP1/2复合物的协同稳定有助于NF- 2持续激活的前馈放大机制。B在结肠炎的慢性炎症和CAC的发展中很重要，并为SphK1和TRAF2在结肠炎和CAC中的上调提供了解释。在第一个目标中，我将阐明SphK1-TRAF2-cIAP1/2共稳定的分子机制，并研究它们各自的酶活性的共调节作用。第二个目的是研究SphK1-TRAF2-cIAP1/2相互作用在NF-？B活化和相关的生物反应。最后，第三个目的是研究SphK1和TRAF2的细胞自主功能及其在结肠炎和CAC小鼠模型中的相互作用。该研究将为理解SphK1对慢性炎症和CAC的影响提供一个范式转变，SphK1对慢性炎症和CAC的影响一直围绕着S1P的形成及其对细胞增殖、保护细胞凋亡和存活的多效性作用，并有望阐明SphK1在慢性炎症和CAC的病因学中的作用，并提供新的治疗策略来干扰NF-？这种致命疾病的B途径

DESCRIPTION (provided by applicant): Colorectal cancer is one of the deadliest forms of cancer and a leading cause of death in developed countries. Unfortunately, for many with chronic bowel inflammation there is a strong link between colitis and the pathogenesis of colitis associated cancer (CAC). One of the best characterized CAC contributing factors is the pro-inflammatory cytokine TNF-¿ and its downstream master transcription factor NF-?B. Hence, targeting the NF-?B activation pathway as a means to treat or prevent CAC progression is gaining popularity among researchers. Recently, our laboratory discovered that the bioactive sphingolipid sphingosine-1-phosphate (S1P) and the enzyme that produces it, sphingosine kinase 1 (SphK1), link persistent NF-?B activation to chronic intestinal inflammation and development of CAC. S1P and SphK1 have long been implicated in the actions of TNF-¿. In the canonical TNF-dependent intracellular signaling cascade leading to NF-?B activation, a complex series of protein interactions and post-translational modifications modulate signal propagation and culminate in gene transcription. Among many others, the receptor interacting kinase-1, the TNF receptor associated factor 2 (TRAF2), and the cellular inhibitor of apoptosis 1 and 2 (cIAP1/2) play important roles. However, the molecular mechanism of the involvement of SphK1 in the NF-?B pathway has remained unclear. Importantly, both SphK1 and TRAF2 have been shown to be elevated in the tissues of patients with CAC and in the colons and tumors of animal models of the disease. My preliminary results suggest that protein-protein interactions between SphK1, TRAF2, and cIAP1/2 are important for their cellular stabilization. Hence, the hypothesis that will be examined in this proposal is that a SphK1-TRAF2-cIAP1/2 complex plays an important role in NF-?B activation by direct protein-protein interactions that regulate the stabiliy, activity and function of each protein. I also suggest that co-stabilization of the SphK1-TRAF2-cIAP1/2 complex contributes to the feed forward amplification mechanism for persistent activation of NF-?B important in chronic inflammation in colitis and development of CAC, and provides an explanation for the upregulation of SphK1 and TRAF2 in colitis and CAC. In the first aim I will elucidate the molecular mechanisms of SphK1-TRAF2-cIAP1/2 co-stabilization and examine co-regulatory effects on their respective enzymatic activities. The second aim will investigate the function of SphK1-TRAF2-cIAP1/2 interactions in the regulation of NF-?B activation and associated biological responses. Finally, the third aim examines the cell autonomous functions of SphK1 and TRAF2 and their interactions in murine models of colitis and CAC. This proposal will provide a paradigm shift in understanding the influence of SphK1 on chronic inflammation and CAC, which has historically revolved around formation of S1P and its pleiotropic effects on cellular proliferation, protection from apoptosis, and survival, and hopefully, clarify the role of SphK1 in the etiology of chronic inflammation and CAC and provide new therapeutic strategies to interfere with upregulation of the NF-?B pathway in this deadly disease.