鼠疫耶尔森氏菌E3泛素连接酶对宿主防御蛋白GBP的泛素化修饰及其生理功能研究

Yersinia pestis (Y. pestis) can cause plague, a potent natural epidemic disease. At present, its pathogenic mechanism is still under-recognized, seriously affecting scientific prevention and control. Through the previous secretory proteomics research of Y. pestis, we screened two virulence proteins Yp_3418 and Yp_3416 with NEL (Novel E3 ligase) domain. Preliminary results have confirmed that they have E3 activity and are capable of degrading multiple interferon-induced guanylate-binding protein (GBP) family members during the infection. The GBP family is an important mechanism recently discovered in the regulation of host-controlled anti-intracellular infections. In this project, we intend to study whether Y. pestis can transport virulence proteins Yp_3418 and Yp_3416 into the host during infection, how to identify and degrade the substrate GBPs through the ubiquitination pathway, and the role of the degradation in the pathogenesis of Y. pestis. This study is expected to reveal the mechanism of Y. pestis resistance to the natural immune defense process involved in GBPs, which is of great significance for understanding its pathogenic mechanism and establishing effective plague prevention and control methods.

鼠疫耶尔森氏菌（简称鼠疫菌）能够引起烈性自然疫源性疾病——鼠疫，目前对其致病机制仍认识不足，严重影响科学防控工作。我们前期鼠疫菌分泌蛋白质组学研究筛选出2个具有NEL型泛素连接酶（E3）结构域的蛋白Yp_3418和Yp_3416。初步证实它们具有E3活性，且能在鼠疫菌感染细胞后降解多个干扰素诱导的鸟苷酸结合蛋白(Guanylate-binding protein, GBP)家族成员。GBP家族蛋白是近年来新发现的参与调控宿主控制抗胞内菌感染的重要机制。本项目中，我们拟研究鼠疫菌在感染过程中是否能将毒力蛋白Yp_3418和Yp_3416分泌入宿主胞内，如何通过泛素化途径靶向识别并降解底物GBPs，及这种降解过程在该菌致病机制中的作用。本研究有望揭示鼠疫菌抵抗GBPs参与的天然免疫防御的机制，对认识其致病机理和建立有效的鼠疫防治手段具有重要意义。

历史上一共发生过三次鼠疫大流行，包括中世纪的黑死病。鼠疫耶尔森菌（简称鼠疫菌）是引起鼠疫的病原菌，在由小肠结肠炎耶尔森菌进化而来的过程中，鼠疫菌已经获得了一种强大的“武器库”来破坏宿主的免疫防御系统。本研究中，我们发现鼠疫菌的两个功能冗余的E3连接酶YspE1和YspE2可以通过III型分泌系统注射到宿主胞质中，并通过泛素化宿主的多种鸟苷酸结合蛋白（GBPs）进行蛋白酶体降解。但是假结核耶尔森菌由于缺乏功能性YspE1/2同源物而没有这种功能。YspE1/2介导的GBP降解显著促进了鼠疫菌在巨噬细胞中的存活，并强烈抑制炎症小体的活化。相比之下，Gbpchr3−/−，chr5−/−型小鼠的巨噬细胞表现出与YspE1 / 2无关的炎症小体活化大大降低，并伴有鼠疫菌的复制显著增强。因此，Gbpchr3−/−，chr5−/−型小鼠对于鼠疫菌更加易感。我们证明了鼠疫菌利用E3连接酶破坏GBP介导的宿主防御，这似乎是鼠疫菌在进化过程中新获得的。