Molecular mechanisms of intracellular growth of Salmonella

沙门氏菌细胞内生长的分子机制

• 批准号: 16017218
• 负责人: YAMAMOTO Tomoko
• 金额: $ 9.41万
• 依托单位: Chiba University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research on Priority Areas
• 财政年份: 2004
• 资助国家: 日本
• 起止时间: 2004 至 2005
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-16017218/
• 关键词: Pathogenesis; Salmonella; Intracellular Growth; Macrophage; マクロファージ内増殖能; ClpXP; PagC; OMV; attenuating virulence factor; 持続感染; アポトーシス; 細胞内寄生菌

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 C1pXP 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 C1pXP and Lon are critically important for the systemic Salmonella infection of mice. We found that the C1pXP mutant and Lon mutant persist in … More 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 clpXPmutants 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 islandl (SPI1) genes. Expression of MIA, 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.(3) To determine the reasons for the loss of virulence of the C1pXP-deficient mutant, we characterized the mutant grown under the conditions that mimic the intracellular environment of macrophages. Consequently, we elucidated that the PagC protein is increased in its amount by C1pXP-depletion and that the increased level is responsible for the loss of virulence of the C1pXP-deficient cells. Furthermore, we have found that the PagC protein is exported by outer membrane vesicle (OMV) from Salmonella cells. Less

包括沙门氏菌在内的细胞内病原体在宿主吞噬细胞内保持长期抵抗力，引发多种遗传程序，帮助它们适应吞噬体内遇到的恶劣环境条件。为了阐明感染后宿主中表达的沙门氏菌发病机制的遗传程序，对沙门氏菌在巨噬细胞吞噬后新诱导的蛋白质进行了蛋白质组分析。由于这些程序中最突出的是热休克反应，因此对作为毒力蛋白的热休克蛋白成员的分子伴侣和AAA+蛋白酶的作用进行了研究。(1)鼠伤寒血清型C1pXP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，这表明 C1pXP 和 Lon 对于小鼠的全身性沙门氏菌感染至关重要。我们发现 C1pXP 突变体和 Lon 突变体在 BALB/c 小鼠体内持续存在很长一段时间，而没有引起压倒性的全身感染，这表明沙门氏菌活疫苗可能是候选者。因此，我们检查了 lon 突变体或 clpXP 突变体的口服免疫是否可以保护小鼠免受随后有毒鼠伤寒血清型的口服攻击。结果表明，lon和clpXP突变体的单次口服免疫应能有效防止野生型鼠伤寒血清型在肠道内定植。(2)我们证明，鼠伤寒血清型中ATP依赖性Lon蛋白酶的耗竭可通过涉及caspase-1和-3的机制诱导巨噬细胞快速和大量凋亡。这种细胞凋亡的过度诱导被 invF 的破坏所消除，invF 是沙门氏菌致病性 island1 (SPI1) 基因表达所必需的。 SPI1转录的核心调节因子MIA的表达在巨噬细胞被吞噬后受到抑制，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 突变体诱导的巨噬细胞凋亡增加可能是由于 SPI1 基因在正常情况下受到抑制的条件下持续表达所致。(3) 为了确定 C1pXP 缺陷突变体毒力丧失的原因，我们对在模拟巨噬细胞细胞内环境的条件下生长的突变体进行了表征。因此，我们阐明 PagC 蛋白的量因 C1pXP 耗尽而增加，并且增加的水平是 C1pXP 缺陷细胞毒力丧失的原因。此外，我们发现 PagC 蛋白是由沙门氏菌细胞的外膜囊泡 (OMV) 输出的。较少的

项目成果

Degradation of the Hi1C and Hi1D regulator proteins by ATP-dependent Lon protease leads to down-regulation of Salmonella pathogenicity island 1 gene expression.

ATP 依赖性 Lon 蛋白酶对 Hi1C 和 Hi1D 调节蛋白的降解导致沙门氏菌致病性岛 1 基因表达下调。

• 发表时间: 2005
• 期刊: Molecular Microbiology 55
• 作者: A.Takaya;Y.Kubota;E.Isogai;T.Yamamoto
• 通讯作者: T.Yamamoto

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

The DnaK/DnaJ chaperone machinery of Salmonella enterica serovar Typhimurium is essential for invasion of epithelial cells and survival in macrophages, leading to systemic infection.

鼠伤寒沙门氏菌的 DnaK/DnaJ 伴侣机制对于上皮细胞的侵袭和巨噬细胞的存活至关重要，从而导致全身感染。

• 发表时间: 2004
• 期刊: Infection and Immunity 72(3)
• 作者: C.Kodama;H.Matsui;A.Takaya et al.
• 通讯作者: A.Takaya et al.

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

master regulator proteins for Salmonella flagellum biogenesis, by the ATP-dependent C1pXP protease.

由 ATP 依赖性 C1pXP 蛋白酶控制沙门氏菌鞭毛生物发生的主调节蛋白。

• 发表时间: 2003
• 期刊: Molecular Microbiology 48
• 作者: T.Tomoyasu;A.Takaya;E.Isogai;T.Yamamoto;Turnover of F1hD;F1hC
• 通讯作者: F1hC