同一原噬菌体在两株希瓦氏菌中的作用及H-NS对其切离调控的研究

Through comparative genomic analyses we found that prophages are widely spread in the genomes of bacteria from Shewanella genus. However, the host regulation of these prophages remains largely unknown. Previously we found that host factor H-NS protein is involved in the regulation of prophage CP4-57So which is inserted at ssrA gene of Shewanella oneidensis MR-1 genome. Additionally, another prophage ΦS73181 was found at ssrA gene location in genome of Shewanella putrefaciens W3-18-1. Interestingly, the same prophage ΦS73181 was also found in Shewanella sp. MR-7 strain. However, ΦS73181 was not inserted at ssrA in genome of MR-7. The size and gene contents of prophage ΦS73181 were different from these of CP4-57So. The purpose of this study is to explore whether prophages are regulated by H-NS in other Shewanella bacteria, and to find out this regulation is dependent on the function of prophage or the location of the prophage inserted. Excision of prophage ΦS73181 will be induced from the genome of W3-18-1 and MR-7, respectively, and gene knock-out strategy as well as Real-time qPCR technology will be used to explore the regulation of H-NS on prophages. This project will help to understand the interaction between prophages and their bacteria host, as well as the regulation network of genes that are horizontally transferred among species.

希瓦氏属细菌基因组中存在大量原噬菌体，对该属细菌原噬菌体的研究却鲜有报道。我们前期研究发现模式菌株MR-1的H-NS蛋白可抑制基因组上ssrA位点插入的原噬菌体CP4-57So的切离，随后我们发现W3-18-1的ssrA位点也存在一个原噬菌体ΦS73181，同一原噬菌体也存在于该属MR-7中，但并没有插入在ssrA位点。原噬菌体ΦS73181与CP4-57So的大小和内部基因组成差异显著。本项目研究目的在于探求H-NS对原噬菌体的调控究竟是依赖原噬菌体的种类与功能，还是依赖其在基因组中插入的位置。我们将诱导原噬菌体切离，采用基因缺失和回补等遗传操作手段、运用实时荧光定量PCR、EMSA等技术，检测ΦS73181在W3-18-1和MR-7中的功能以及宿主H-NS对ΦS73181切离的调控。研究将帮助我们进一步了解细菌与噬菌体相互作用的机制，并为细菌间水平基因转移的调控提供有价值的科学证据。

噬菌体与细菌的相互作用,特别是细菌基因组上的原噬菌体与宿主的相互作用，是目前微生物研究领域的热点之一。希瓦氏属细菌基因组中存在大量原噬菌体，对该属细菌原噬菌体的研究却鲜有报道。我们通过比较基因组学分析从54株希瓦式菌株中发现15株细菌的同一基因ssrA位点插入了原噬菌体。通过分析这些原噬菌体的特点，找到了可诱导不同类型原噬菌体切离的基因，通过诱导原噬菌体从基因组上自发切离，利用这一新方法获得原噬菌体缺失型细菌突变株。围绕同属细菌同一基因ssrA位点的原噬菌体的作用，研究原噬菌体对其宿主的功能，并探讨细菌“组蛋白”H-NS对原噬菌体的调控机制。本项目部分解决了申请书提出的：H-NS对原噬菌体的调控是依赖原噬菌体的种类与功能，还是依赖其在基因组中插入的位置这一科学问题，并发现细菌可调控其基因组上原噬菌体来应对外界环境变化的刺激。本项目的研究加深了对细菌和原噬菌体相互作用机制的认识，拓宽了对细菌水平转移基因调控的理解，已发表SCI论文1篇。

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