| Project/Area Number |
10670261
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Bacteriology (including Mycology)
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| Research Institution | Ehime University |
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Principal Investigator |
SHINOMIYA Hiroto Ehime University, 医学部, 助教授 (80162618)
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| Project Period (FY) |
1998 – 1999
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| Project Status |
Completed (Fiscal Year 1999)
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| Budget Amount *help |
¥2,700,000 (Direct Cost: ¥2,700,000)
Fiscal Year 1999: ¥1,000,000 (Direct Cost: ¥1,000,000)
Fiscal Year 1998: ¥1,700,000 (Direct Cost: ¥1,700,000)
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| Keywords | pp65 / plastin / host defnse / protein phosphorylation / signal transduction / cytoskeleton / cell adhension / splenomegaly / Mac-1 / PP65 / L-Plastin / Plastinイソフォーム / リン酸化 / 細菌感染 / アクチン結合蛋白 |
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| Research Abstract |
1. Preparation of cDNA and recombinant protein of pp65/plastin family and antibodies againt them : Recombinant L-plastin was produced in E. coli transfected with the cDNA construct in pET vector. Polyclonal antibodyies againt L-plastin were prepared by immunizing rabbits with the recombinant protein. We also cloned the T-plastin gene.
2. Characterization of an LPS-induced serine kinase that phosphorylates pp65/L-plastin : An LPS-stimulated serine kinase that phosphorylates pp65 has been characterized by using peptide substrates. In vivo kinase assay has revealed that the pp65-kinase was stimulated approximately 3-fold in cytosol extracts from LPS-treated macrophages. The enzymatic activity was not dependent on Ca2+ or cAMP, and not inhibited by heparine, a strong inhibitor for casein kinase II. The pp65-kinase activity in extracts from LPS-treated macrophages was preserved after rapid chromatography on a Mono Q column. These results suggest that a soluble serine kinase is rapidly activated by LPS and its properties distinguish it from protein kinases previously described in the literature.
3. Role of pp65/L-plastin in the defense against Salmonella-infection : Jones, S. L., et al. recently clarified that the LPS-induced phosphorylation of the serine-5 of pp65/L-plastin, which was originally determined by us, augmented the adhesiveness of the macrophages through Mac-1 adhesion molecules. We found that Mac-1+ cells were increased in infected-spleens about 25 times as many as normal spleens, and that activities of the pp65-kinase in the cells were also increased. Splenomegaly was found to be important in the protection of the host against Salmonella-infection. Thus, it was suggested that the pp65/L-plastin-Mac-1 system play a pivotal role in the defense againt infections by rapidly accumulationg macrophages to lymphoid organs by modulating the activities of adhesion molecules.
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