| Project/Area Number |
16017218
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| Research Category |
Grant-in-Aid for Scientific Research on Priority Areas
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| Allocation Type | Single-year Grants |
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| Review Section |
Biological Sciences
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| Research Institution | Chiba University |
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Principal Investigator |
YAMAMOTO Tomoko Chiba University, Graduate School of Pharmaceutical Sciences, Professor, 大学院・薬学研究院, 教授 (60110342)
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| Co-Investigator(Kenkyū-buntansha) |
IOMOYASU Toshifumi Chiba University, Graduate School of Pharmaceutical Sciences, Associate Professor, 大学院・薬学研究院, 助教授 (20323404)
TAKAYA Akiko Chiba University, Graduate School of Pharmaceutical Sciences, Lecturer, 大学院・薬学研究院, 講師 (80334217)
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| Project Period (FY) |
2004 – 2005
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| Project Status |
Completed (Fiscal Year 2005)
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| Budget Amount *help |
¥14,700,000 (Direct Cost: ¥14,700,000)
Fiscal Year 2005: ¥7,300,000 (Direct Cost: ¥7,300,000)
Fiscal Year 2004: ¥7,400,000 (Direct Cost: ¥7,400,000)
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| Keywords | 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 |
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| Research Abstract |
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
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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
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