Rip Proteins in Innate Immune Signaling

细胞因子肿瘤坏死因子、病原体识别Toll样受体及其核苷酸结合 寡聚化结构域(NOD)蛋白部分通过激活 转录因子NF-βB。我们发表的工作揭示了RIP1是一种关键的肿瘤坏死因子-和 TLR3/4、Trif依赖的核因子-βB通路和RIP1的泛素修饰对肿瘤坏死因子-1是必不可少的。 诱导核因子B的活化和细胞因子的产生。我们发现RIP1缺陷细胞的I型受损 干扰素对病毒感染的反应，揭示RIP1参与RIG-I/MDA5抗病毒信号转导。 我们最近的研究揭示了RIP1在泛素化和激活中的一个新的调节作用 干扰素调节因子7(IRF-7)，一种对I型干扰素产生至关重要的转录因子。 因此，我们假设RIP1和潜在的多泛素化的RIP1介导先天抗病毒 通过调节核因子-B和IRF-7的活性进行免疫反应。为了支持这一假设，我们将测试 RIP1缺陷是否会损害体内的先天抗病毒反应并感染RIP1缺陷 巨噬细胞和树突状细胞与多种病毒结合以确定RIP1如何调节 IRF-7的转录活性(目标1)。激活肿瘤坏死因子、TRIF或RIG-I/MDA5通路刺激 RIP1多泛素化，因此我们将测试激活是否需要多泛素化RIP1 核因子-B和/或IRF-7在病毒感染细胞中的表达，并将确定泛素不能修饰 RIP1导致先天抗病毒反应受损(目标2)。同样，我们对相关RIP1的研究 蛋白RIP2发现内源性RIP2在MDP刺激的巨噬细胞中泛素化，这表明 NOD2途径受泛素调控。我们假设多泛素化的RIP2是 NOD2介导的核因子-B活化和先天抗细菌免疫反应 泛素去调控导致NOD2相关的人类炎症性疾病。为了测试这一点 假设，我们将确定RIP2上的关键泛素位点，并将测试多泛素化的RIP2 是NOD介导的NF-B激活所必需的，它将决定NOD2等位基因如何表达 与人类炎症性疾病相关影响RIP2的募集和多泛素化 (目标3)。总而言之，我们的研究表明，先天免疫反应是由泛素调节的， 负责RIP蛋白泛素修饰的酶可能是 有针对性地治疗传染病或慢性炎症性疾病。

The cytokine TNF, the pathogen recognition Toll-like receptors (TLRs) and the nucleotide binding oligomerization domain (NOD) proteins mediate host defense against infection in part by activating the transcription factor NF-¿B. Our published work reveals Rip1 as a critical mediator of the TNF- and TLR3/4, Trif-dependent NF-¿B pathways and the ubiquitin modification of Rip1 is essential for TNF- induced NF-¿B activation and cytokine production. We find Rip1-deficient cells impaired in their type I interferon response to viral infection, revealing that Rip1 contributes to Rig-I/Mda5 anti-viral signaling. Our recent studies reveal a novel regulatory role for Rip1 in the ubiquitination and activation of the interferon regulatory factor 7 (IRF-7), a transcription factor critical for type I interferon production. Therefore, we hypothesize that Rip1 and potentially polyubiquitinated Rip1 mediate anti-viral innate immune responses by regulating NF-¿B and IRF-7 activity. To support this hypothesis, we will test whether a Rip1-deficiency impairs innate anti-viral responses in vivo and will infect Rip1-deficient macrophages and dendritic cells with multiple classes of viruses to determine how Rip1 regulates the transcriptional activity of IRF-7 (Aim 1). Activation of the TNF, Trif or Rig-I/Mda5 pathways stimulates Rip1 polyubiquitination, hence we will test whether polyubiquitinated Rip1 is required for the activation of NF-¿B and/or IRF-7 in virally infected cells and will determine whether an inability to ubiquitin modify Rip1 results in impaired innate anti-viral responses (Aim 2). Similarly, our studies on the related Rip1 protein Rip2 find endogenous Rip2 polyubiquitinated in MDP-stimulated macrophages, suggesting that the Nod2 pathway is ubiquitin-regulated. We hypothesize that polyubiquitinated Rip2 is required for Nod2-mediated NF-¿B activation and for innate anti-bacterial immune responses and predict that ubiquitin deregulation contributes to NOD2-associated human inflammatory diseases. To test this hypothesis, we will identify the critical ubiquitin site on Rip2 and will test whether polyubiquitinated Rip2 is required for Nod-mediated NF-¿B activation and will determine how expression of NOD2 alleles associated with human inflammatory diseases affect the recruitment and polyubiquitination of Rip2 (Aim3). Collectively, our studies suggest that innate immune responses are ubiquitin regulated, raising the possibility that the enzymes responsible for the ubiquitin modification of Rip proteins may be targeted therapeutically to treat infectious disease or chronic inflammatory disease.