Molecular mechanisms of intracellular growth of Salmonella

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抗性，引起各种遗传程序，以帮助它们适应吞噬体内遇到的恶劣环境条件。为了阐明沙门氏菌感染后宿主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。由于这些程序中突出的是热休克反应，分子伴侣和AAA+蛋白酶的作用，这是成员的热休克蛋白作为毒力蛋白进行了研究。（1）鼠伤寒血清型C1pXP或Lon的破坏导致对小鼠的毒力丧失。减毒可能是由于巨噬细胞内存活和复制能力受损，表明C1pXP和Lon对小鼠的全身沙门氏菌感染至关重要。我们发现，C1pXP突变体和Lon突变体持续存在， ...更多信息 BALB/c小鼠长期接种，未引起全身感染，提示沙门氏菌活疫苗的可能候选者。因此，我们研究了用lon突变体或clpXP突变体口服免疫是否保护小鼠免受随后用强毒鼠伤寒血清型口服攻击。结果表明，lon和clpXP突变体的单次口服免疫应该有效地保护免受肠道内野生型血清型鼠伤寒的定殖。(2)我们证明，消耗血清型鼠伤寒中的ATP依赖性Lon蛋白酶通过涉及半胱天冬酶-1和-3的机制诱导巨噬细胞快速和大量的凋亡。这种过度诱导的细胞凋亡被废除的破坏invF，这是所需的沙门氏菌致病岛1（SPI1）基因的表达。MIA是SPI1转录的中心调节因子，其表达在吞噬后的巨噬细胞中受到抑制，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，由Lon突变体诱导的巨噬细胞凋亡的增加可能是由于SPI1基因在它们通常被抑制的条件下持续表达。(3)为了确定C1pXP缺陷突变体的毒性丧失的原因，我们表征了在模拟巨噬细胞的细胞内环境的条件下生长的突变体。因此，我们阐明了PagC蛋白的量通过C1pXP-耗尽而增加，并且增加的水平是C1pXP-缺陷细胞的毒力丧失的原因。此外，我们已经发现，PagC蛋白输出的外膜囊泡（OMV）从沙门氏菌细胞。少

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