Studies on Regulatory Mechanism of Salmonella Pathogenesis by Proteome Analysis

蛋白质组分析研究沙门氏菌发病机制

• 批准号: 13470058
• 负责人: YAMAMOTO Tomoko
• 金额: $ 9.15万
• 依托单位: Chiba University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B)
• 财政年份: 2001
• 资助国家: 日本
• 起止时间: 2001 至 2004
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-13470058/
• 关键词: Salmonella; Proteome; Macrophage; Pathogenesis; Apoptosis; Molecular Chaperone; Lon; ClpXP; SPI1; Lonプロテアーゼ; ClpXPプロテアーゼ; 侵入性; 鞭毛; ストレス蛋白質

Intracellular pathogens, including Salmonella, which maintain long-term resistance within host phagocytes elicit a variety of genetic programs to help them adapt to the hostile environmental conditions encountered within the phagosome. To elucidate the genetic programs for Salmonella pathogenesis expressed in host after infection, a proteome analysis of proteins newly induced in Salmonella after phagocytosis by macrophages was performed. Since prominent among these programs was the heat shock response, the, role of molecular chaperones and AAA+ proteases which are members of the heat shock proteins as virulence proteins have been studied.(1)The disruption of ClpXP or Lon of serovar Typhimurium results a loss of virulence to mice. The attenuation could be due to the impaired ability to survive and replicate within macrophage cells, suggesting that ClpXP and Lon are critically important for the systemic Salmonella infection of mice. We found that the ClpXP mutant and Lon mutant persist i … More n the BALB/c mice for long periods of time without causing an overwhelming systemic infection, suggesting a possible candidate of Salmonella live vaccine. We therefore examined whether oral immunization with the lon mutant or clpXP mutant protects mice against subsequent oral challenge with virulent serovar Typhimurium. The results suggested that a single oral immunization of the lon and clpXP mutants should be effective to protect against the colonization of wild-type serovar Typhimurium within the intestinal tract.(2)We demonstrated that depletion of ATP-dependent Lon protease in serovar Typhimurium induces rapid and massive apoptosis in macrophages by a mechanism involving both caspases-1 and -3. This excessive induction of apoptosis was abrogated by disruption of invF, which is required for the expression of the Salmonella pathogenicity island 1(SPI1) genes. Expression of hilA, a central regulator of SPI1 transcription, was repressed in the macrophages after phagocytosis, but this gene was continuously expressed when the Lon mutant grew within the macrophages, so the SPI1 proteins accumulated. Thus, the increase in macrophage apoptosis induced by the Lon mutant could be due to continued expression of SPI1 genes under conditions where they are normally repressed. Once Salmonella has established a systemic infection, excess apoptosis of macrophages cells upon which the organism is reliant would be detrimental to the pathogen. Therefore, the Lon protease may be required to suppress apoptosis sufficiently to allow time for the bacterium to replicate, escape, and invade new macrophages.(3)The DnaK/DnaJ-depleted mutant lost the ability to causes a lethal systemic disease in mice. Macrophage-survival assays revealed that the mutant could not survive or proliferate at all within macrophages. Of further interest are the findings that the mutant could neither invade cultured epithelial cells nor secrete any of the invasion proteins encoded by Salmonella pathogenicity island 1. Less

细胞内病原体，包括沙门氏菌，在宿主吞噬细胞内保持长期抗性，引发各种遗传程序，帮助它们适应吞噬小体内遇到的恶劣环境条件。为了阐明沙门氏菌感染后在宿主中表达的致病基因程序，对沙门氏菌被巨噬细胞吞噬后新诱导的蛋白质进行了蛋白质组分析。由于在这些程序中突出的是热休克反应，因此研究了热休克蛋白中的分子伴侣和AAA+蛋白酶作为毒力蛋白的作用。(1)鼠伤寒沙门氏菌ClpXP或Lon的破坏导致对小鼠的毒力丧失。这种减弱可能是由于巨噬细胞内的生存和复制能力受损所致，这表明ClpXP和Lon在小鼠系统性沙门氏菌感染中至关重要。我们发现ClpXP突变体和Lon突变体持续I…更多的沙门氏菌在BALB/c小鼠体内长期存在，而不会引起压倒性的全身感染，这表明可能是沙门氏菌活疫苗的候选者。因此，我们研究了Lon突变体或ClpXP突变体口服免疫是否能保护小鼠免受强毒血清鼠伤寒沙门氏菌的攻击。结果表明，一次口服免疫Lon和ClpXP突变体可以有效地防止野生型鼠伤寒沙门氏菌在肠道内的定植。(2)我们证明了依赖于ATP的Lon蛋白酶在鼠伤寒沙门氏菌中的缺失通过caspase-1和-3共同参与的机制诱导巨噬细胞快速而大量的凋亡。沙门氏菌致病性岛1(SPI1)基因表达所必需的invF被破坏，从而消除了这种过度的诱导细胞凋亡的作用。巨噬细胞吞噬后，SPI1转录的中心调控因子Hila的表达受到抑制，但当Lon突变体在巨噬细胞内生长时，该基因持续表达，因此SPI1蛋白积累。因此，Lon突变体诱导巨噬细胞凋亡的增加可能是由于SPI1基因在正常情况下被抑制的情况下继续表达。一旦沙门氏菌确立了全身感染，机体所依赖的巨噬细胞过度凋亡将对病原体有害。因此，可能需要Lon蛋白酶来充分抑制细胞凋亡，以便细菌有时间复制、逃逸和入侵新的巨噬细胞。(3)Dna K/Dna J缺失的突变体失去了在小鼠中引起致命性全身疾病的能力。巨噬细胞存活分析显示，该突变体根本不能在巨噬细胞内存活或增殖。更令人感兴趣的是，该突变体既不能入侵培养的上皮细胞，也不能分泌任何由沙门氏菌致病性岛1编码的入侵蛋白。

项目成果

Derepression of Salmonella pathogenicity island 1 genes within macrophages leads to rapid apoptosis via caspase‐1‐ and caspase‐3‐dependent pathways

• DOI: 10.1111/j.1462-5822.2004.00435.x
• 发表时间: 2004-08
• 期刊: Cellular Microbiology
• 影响因子: 3.4
• 作者: A. Takaya;A. Suzuki;Y. Kikuchi;M. Eguchi;E. Isogai;T. Tomoyasu;Tomoko Yamamoto

Takaya A., et al.: "The ATP-dependent Lon protease of Salmonella enterica serovar Typhimurium regulates invasion and expression of genes encoded on Salmonella pathogenicity island"Journal of Bacteriology. 184. 224-232 (2002)

Takaya A. 等人：“鼠伤寒沙门氏菌肠炎血清型的 ATP 依赖性 Lon 蛋白酶调节沙门氏菌致病岛编码基因的侵袭和表达”《细菌学杂志》。

The ATP-dependent Lon protease of Salmonella enterica serovar Typhimurium regulates invasion and expression of genes carried on Salmonella pathogenicity island 1

• DOI: 10.1128/jb.184.1.224-232.2002
• 发表时间: 2002-01-01
• 期刊: JOURNAL OF BACTERIOLOGY
• 影响因子: 3.2
• 作者: Takaya, A;Tomoyasu, T;Yamamoto, T
• 通讯作者: Yamamoto, T

医学大辞典

医学词典

• 发表时间: 2006
• 作者: 末広 寛;Heather Skirton;村上 京子;塚原正人;Skirton H;飯野英親;飯野英親;塚原正人;辻野久美子;松本浩;飯野英親;飯野英親;川内美穂子;斉藤知子;村上京子;末広 寛;辻野久美子;Matsumoto Y;飯野英親;村上京子;飯野英親;中込さと子;塚原正人;塚原正人;塚原 正人;飯野 英親;辻野 久美子;村上 京子;飯野 英親;辻野 久美子;石田喬士;相澤忠範
• 通讯作者: 相澤忠範

Turnover of FlhD and FlhC, master regulator proteins for Salmonella flagellum biogenesis, by the ATP‐dependent ClpXP protease

• DOI: 10.1046/j.1365-2958.2003.03437.x
• 发表时间: 2003-04
• 期刊: Molecular Microbiology
• 影响因子: 3.6
• 作者: T. Tomoyasu;A. Takaya;E. Isogai;Tomoko Yamamoto