| Project/Area Number |
08457102
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| Research Category |
Grant-in-Aid for Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Immunology
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| Research Institution | Chiba University |
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Principal Investigator |
SAITO Takashi Chiba Univ., Sch.of Med., Professor, 医学部, 教授 (50205655)
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| Co-Investigator(Kenkyū-buntansha) |
ARASE Hisashi Chiba Univ., Sch.of Med., Assistant, 医学部, 助手 (10261900)
OHNO Hiroshi Chiba Univ., Sch.of Med., Associate Professor, 医学部, 助教授 (50233226)
宮武 昌一郎 千葉大学, 医学部, 講師 (30239420)
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| Project Period (FY) |
1996 – 1997
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| Project Status |
Completed (Fiscal Year 1997)
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| Budget Amount *help |
¥7,300,000 (Direct Cost: ¥7,300,000)
Fiscal Year 1997: ¥3,600,000 (Direct Cost: ¥3,600,000)
Fiscal Year 1996: ¥3,700,000 (Direct Cost: ¥3,700,000)
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| Keywords | Jak3 / TCR / Signal transduction / pre-TCR complex / calnexin / CD3gamma / positive / negative selection / T cell differentiation / T細胞分化 / T細胞 / 分化 / 遺伝子欠損マウス / アビディティーモデル / TCR-CD3複合体 |
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| Research Abstract |
We demonstrated last year that Jak3 plays important roles not only in signal transduction of T cell growth but also in T cell development and maintenance of T cell function in periphery. We analyzed whether Jak3 is related to TCR signaling and found that Jak3 is indeed tyrosine-phosphorylated upon TCR-stimulation. Moreover, it was found that Jak3 directly associates with CD3zeta chain when Jak3 and CD3zeta were co-transfected into COS cells. When dominant-negative Jak3 lacking the kinase domain was transfected into antigen-specific T cell hybridoma cells, IL-2 production was markedly reduced in the transfectant upon antigen-stimulation. From these results, Jak3 is an important signal transducer of TCR signals and regulates IL-2 production in T cells.
Analyzing an immature thymocyte cell line expressing pre-TCR complex, we found that pre-TCR complex contains both calnexin and a new 12 kd dimer. To identify this new 12 kd molecule (pTAC12), we collected protein from a huge number of the c
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ells and succeeded partial amino acid analysis of pTAC12. Surprisingly, it was the same sequence as CD3gamma chain. Since pTAC12 is much smaller than CD3gamma, cDNA was isolated by 5'RACE method which should have a related sequence to CD3gamma. To this end, we found that pTAC12 encodes an alternative splicing product of CD3gamma which lacks an exon containing the transmembrane region. Indeed, pTAC12 was reacted by anti-CD3gamma Ab.
Furthermore, in order to analyze regulation of T cell differentiation by TCR signals, we analyzed HY-Tg mice crossed with CD3zeta-deficient mice (HY-Tgzeta-KO) last year. This year, we devised a better system by crossing CD3zeta-KO mice with OVA-specific TCR-Tg mice (DO-Tgzeta-KO) in which the level of TCR and peptide can be changed simultaneously. Whereas T cells in these mice were not selected in the absence of antigen peptide, T cells exhibited positive selection and then negative selection as the concentration of peptide increased. Therefore, we succeeded to show the direct correlation between the strength of TCR signals and T cell selection in a system where the TCR expression level on the cell surface of T cells was reduced by CD3zeta-deficiency. Less
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