鼠疫菌致病性和传播性的遗传基础：质粒获得与基因调控重塑

Yersinia pestis is the causative agent of plague, which caused three pandemics in human history, and plague is an important infectious disease that changing the path of human civilization. Compared with its ancestor Yersinia pseudotuberculosis, Y. pestis acquired transmissibility by flea and high pathogenicity. Previous studies demonstrated that these two newly emerging features were related to two newly acquired plasmids (pPCP1 and pMT1). However, its molecular mechanism has not been elucidated. This project is based on the scientific hypothesis that acquisition of plasmids and their interactions, and their roles in remodeling of genetic regulation system played critical roles during Y. pestis evolution. The palsmid-cured strains (pCD1+、pMT1+、pPCP1+、pCD1+-pMT1+、pCD1+-pPCP1+和pMT1+-pPCP1+) and wild type strain (pCD1+-pMT1+-pPCP1+) will be compared in terms of pathogenicity using animal model and study on molecular basis for the role of acquired plasmids on pathogenicity. The role of genetic regulation system remodeling by newly acquired regulators and sRNAs and their molecular mechanisms in determining pathogenicity and transmissibility of Y. pestis will also be elucidated using different kind of methods. Therefore, the objective of this project, molecular basis of plasmids acquisition and genetic regulation remodeling in determining pathogenicity and transmissibility, could be revealed. Our laboratory has focused on genomics, evolution and pathogenicity of Yersinia pestis for more than 10 years and has laid a solid foundation for this project. The success of this project will promote our understanding on the evolutionary process of Y. pestis, and also provide novel idea for studying other bacterial pathogens.

鼠疫菌是导致鼠疫的病原体，历史上曾致三次世界大流行，是改变人类文明进程的重要传染病。鼠疫菌与其祖先假结核菌比，进化了经蚤传播能力和高致病性。前期研究表明，它们与两个新获得质粒（pPCP1和pPMT1）有关。但其分子基础仍未被揭示。本研究基于“质粒获得及其相互作用和对基因调控网络的重塑在鼠疫菌致病性和传播性获得中发挥着关键作用”的科学假设，利用动物模型比较质粒缺失株（pCD1+、pMT1+、pPCP1+、pCD1+-pMT1+、pCD1+-pPCP1+和pMT1+-pPCP1+）和野生株（pCD1+-pMT1+-pPCP1+）的致病性变化，研究外源质粒对鼠疫菌致病性的影响及其分子机制；并研究新获得质粒编码的调控子和sRNA对基因调控网络重塑及其在致病性和传播性中作用的分子机制。从而探究 “质粒获得与基因调控重塑决定鼠疫菌致病性和传播性的分子基础”这一命题。

鼠疫曾经有三次世界大流行，给人类带来深重灾难，也影响着人类的文明进程。其病原菌是鼠疫耶尔森氏菌，与2万年前有假结核耶尔森氏菌演化而来。从其祖先的肠道致病性演化成一种经蚤传播的烈性传染病。在这个过程中，鼠疫菌从祖先菌继承了一个编码三型分泌系统的质粒（pCD1），并额外获得了两个质粒（pPCP1和pMT1），染色体也发生了一些基因突变。前期研究表明，鼠疫菌的毒力是有质粒编码基因和染色体上的基因共同决定的，但未发现染色体上其决定作用的位点。因此，质粒的功能备受重视。pCD1是致病耶尔森氏菌共有的，另外两个质粒编码的基因决定着其跳蚤传播和体内的侵袭性；但对质粒间互作在鼠疫菌演化和致病中的作用机制不甚了解。本课题旨在研究质粒基因及其编码的小RNA，蛋白质的互作，以及染色体基因变异在鼠疫菌毒力进化中的意义进行研究。我们发现了鼠疫菌两个额外质粒的互作及其决定鼠疫菌经蚤传播的机制，如发现pPCP1质粒编码的Pla蛋白降解pMT1质粒编码的鼠毒素（Ymt），而且证明鼠毒素是一种分泌蛋白，揭示了Ymt通过抑制天然和获得性免疫系统而保障鼠疫菌在动物血液中高密度繁殖，以确保跳蚤的传播；发现了多个小RNA和分泌蛋白质在鼠疫菌致病性演化中的作用；揭示了鼠疫菌受环境因素影响的基因变异规律及其影响跳蚤叮咬行为的延伸表型，即寒冷和干旱可以促进鼠疫菌RNA聚合酶亚基基因rpoZ突变，该基因的突变导致鼠疫菌在体外生物膜形成能力增加，推测这可导致该菌在跳蚤前胃中形成生物膜的速度加快，从而可以促进跳蚤传播细菌，鼠疫菌的这种变异保障了在不利条件下鼠疫菌的自身生存能力。这些发现对理解鼠疫菌致病性的进化具有重要意义。