泛素连接酶Peli1调控巨噬细胞中NLRP3炎症小体活化的机制研究

Inflammasomes play a key role in the initial and progressive process among various inflammation-related diseases. Ubiquitination is an important way to regulate the immune homeostasis. Compared with the priming and activation mechanisms of the inflammasome signaling pathway, post-translational ubiquitination mechanisms controlling inflammasome activation have not been clearly addressed. We found that the E3 ubiquitin ligase Peli1 can regulate the NLRP3 inflammasome activation. Peli1 deficiency decreases the secretion of IL-1β and IL-18 with modulating Caspase-1 cleavage and has a reduction in ASC speck formation in cultured macrophages. Consistently, Peli1-/- mice show lower IL-1β secretion in the sepsis model of intraperitoneal LPS injection. Our project is expected to reveal the mechanism of Peli1-mediated regulate the activation of NLRP3 inflammasome. In this project, the following research will be carried out: we used both in vivo and vitro experiments to confirm that Peli1 can regulate the activation of NLRP3 inflammasome; we will screen the target proteins, ubiquitination involving in Peli1 regulated NLRP3 signaling pathway, investigate their function; we will reveal the mechanism of Peli1-mediated regulate the activation of NLRP3inflammasome. Solid evidences from this project will be helpful for understanding the molecular mechanism mediating the activation of NLRP3 inflammasome and providing potential targets for the diagnosis and treatment of inflammation-related diseases.

炎症小体作为固有免疫的重要成分在炎症相关疾病的发生发展中发挥关键作用。泛素化修饰作为表观调控的一种方式对于调控胞内信号发挥重要作用，然而泛素化修饰对炎症小体活化的调控机制仍知之甚少。我们预实验发现干预E3泛素连接酶Peli1可显著抑制炎症反应中IL-1β的分泌，减缓小鼠急性炎症的发生发展；干预Peli1可显著抑制巨噬细胞NLRP3炎症小体的活化；通过免疫共沉淀实验发现Peli1能够与NLRP3相互作用。据此我们提出假说：Peli1能够通过泛素化靶蛋白NLRP3，参与调控NLRP3炎症小体的活化。本项目拟开展以下研究：利用Peli1基因敲除小鼠，明确Peli1对炎症小体活化的调控作用；确定Peli1作用的靶蛋白；阐明Peli1与靶蛋白相互作用的结构域和泛素化修饰类型。本研究将有助于揭示Peli1调控NLRP3炎症小体活化的分子机制，为炎症相关疾病的诊治提供新思路。

炎症小体作为固有免疫的重要成分在炎症相关疾病的发生发展中发挥关键作用。泛素化修饰作为表观调控的一种方式对于调控胞内信号发挥重要作用，然而泛素化修饰对炎症小体活化的调控机制仍知之甚少。我们首先通过peli1基因敲除小鼠和对照组小鼠构建败血症和腹膜炎模型发现：干预E3泛素连接酶Peli1可显著抑制炎症反应中IL-1β的分泌，减缓小鼠急性炎症的发生发展，这提示Peli1的缺失会抑制炎症小体的活化；然后我们通过体外培养Peli1基因敲除小鼠和对照组小鼠的骨髓来源巨噬细胞实验表明，干预Peli1可显著抑制巨噬细胞中NLRP3炎症小体的活化；进一步通过免疫共沉淀实验机制研究发现Peli1能够与炎症小体相关蛋白ASC相互作用介导ASC的K63连接的泛素化。该研究首次发现泛素连接酶Peli1能够介导ASC泛素化调控NLRP3炎症小体的激活，这将为NLRP3相关炎症疾病的诊治提供了潜在的新靶点和策略。