RIP2 Caspase-1 Signaling in Macrophages

项目摘要 巨噬细胞释放17 kDa IL-1是一个高度受调控的事件，它超出了巨噬细胞的 转录和合成前体31 kDa proIL-1的能力。此事件以激活 Caspase-1酶，涉及一种称为炎症体的复杂蛋白质组合。炎性小体 成分是存在于植物和动物中的一种古老的固有宿主防御系统的同系物。 本提案试图扩大从IL-1的巨噬细胞生物学中获得的线索。 将这些过程扩展到宿主对脓毒症的基本反应。我们有最近的数据证明 这种细胞内炎性小体复合体的关键成分对毁灭性的 激活定义感染性休克的先天宿主反应。更具体地说，caspase-1，中央 炎症小体的靶标与脓毒症有直接联系。不仅是caspase-1基因敲除的动物 防止脓毒症死亡但caspase-1炎症体复合体的个别成分(例如， Caspase-5、ASC和NALP1)也与脓毒症的结局有关。这些蛋白质相互作用 其他途径包括caspase募集结构域(CARD)和结构相关的吡咯域(PYD)。我们的 假说是，以这些卡片和PYD域为中心的翻译后事件是 脓毒症的发病机制。炎性小体组装不仅调节细胞的加工和激活 炎性细胞因子如IL-1和IL-18，但延伸到对核因子B和宿主细胞凋亡的调节。 目前的提案试图将这些令人兴奋的突破应用于巨噬细胞生物学 应对败血症的挑战。我们知道炎性小体组装中的关键调控事件之一 触发卡/卡和PYD/PYD交互。我们最近已经证明，监管 含卡分子(如RIP2和ASC)可以引导以caspase-1为中心的炎症反应 或向核因子B或向IL-1？处理。有趣的是，这些事件涉及早期的磷酸化。 抑制酪氨酸激酶活性深刻影响炎症小体并保护动物以来的事件 死于败血症。因此，这项提议试图剖析这些事件如何受到调控的分子细节。 中心假设是，决定PRO方向和严重程度的主监管开关 而对败血症攻击的抗炎反应在胞浆中由特定的卡片/卡片和 PYD/PYD互动。我们将剖析调节caspase-1和RIP2能力的机制 相互作用，运输到膜上，然后引导宿主炎症。这些事件最初会诱导NFkB激活 但随后，通过caspase-1的催化激活，可能最终诱导细胞凋亡，下调核因子B的表达 事件。这些研究将提高我们对脓毒症的基本先天宿主反应的理解。 这样做可以发现感染性休克和其他炎症性疾病的新治疗方法 受这个以caspase-1为中心的过程调节。

Project Summary Macrophage release of 17 kDa IL-1¿ is a highly regulated event that extends beyond the macrophage's ability to transcribe and synthesize the precursor, 31 kDa proIL-1¿. This event is centered upon activation of the enzyme caspase-1 and involves a complex protein assemblage termed the inflammasome. Inflammasome components are homologues of an ancient innate host defense system that exists in both plants and animals. The present proposal seeks to expand upon the clues derived from the macrophage biology of IL-1¿ and extend these processes to the basics of the host response to sepsis. We have recent data to demonstrate that key components of this intracellular inflammasome complex contribute significantly to the devastating activation of the innate host response that defines septic shock. More specifically, caspase-1, the central target of the inflammasome has been directly linked to sepsis. Not only are caspase-1 knockout animals protected from sepsis mortality but individual components of the caspase-1 inflammasome complex (e.g., caspase-5, ASC and NALP1) have also been linked to sepsis outcomes. These proteins interact with each other via caspase recruitment domains (CARDs) and structurally related pyrin domains (PYDs). Our hypothesis is that posttranslational events centered upon these CARD and PYD domains are central to the pathogenesis of sepsis. Inflammasome assembly not only regulates the processing and activation of inflammatory cytokines like IL-1¿ and IL-18 but extends to the regulation of NF¿B and host cell apoptosis. The current proposal seeks to apply these exciting breakthroughs in the biology of macrophage function to the challenge of sepsis. We know that one of the key regulatory events in inflammasome assembly is the triggering of CARD/CARD and PYD/PYD interactions. We have recently demonstrated that regulating CARD-containing molecules (e.g., RIP2 and ASC) can direct the caspase-1 centered inflammatory response either toward NF¿B or toward IL-1¿ processing. Interestingly, these events involve an early phosphorylation event since inhibition of tyrosine kinase activity profoundly affects the inflammasome and also protects animals from sepsis. Thus, this proposal seeks to dissect the molecular details of how these events are regulated. The central hypothesis is that the master regulatory switch that determines the direction and severity of the pro and anti-inflammatory responses to septic challenge are controlled in the cytosol by specific CARD/CARD and PYD/PYD interactions. We will dissect the mechanisms that regulate the ability of caspase-1 and RIP2 to interact, traffic to membranes and then direct host inflammation. These events initially induce NFkB activation but subsequently, via caspase-1 catalytic activation, may finally induce apoptosis and down regulation of NF¿B events. These studies will improve our understanding of the basic innate host responses to sepsis and in doing so uncover novel therapeutic approaches to septic shock and other inflammatory disorders that are regulated by this caspase-1-centric process.