| Project/Area Number |
05670891
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| Research Category |
Grant-in-Aid for General Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Research Field |
内分泌・代謝学
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| Research Institution | UNIVERSITY OF OCCUPATIONAL AND ENVIRONMENTAL HEALTH,JAPAN |
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Principal Investigator |
FUJIHIRA Takashi UOEH,SCH OF MED,ASSISTANT PROFESSOR, 医学部, 助手 (40159124)
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| Co-Investigator(Kenkyū-buntansha) |
ETO Sumiya UOEH,SCH OF MED,PROFESSOR, 医学部, 教授 (90010347)
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| Project Period (FY) |
1993 – 1994
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| Project Status |
Completed (Fiscal Year 1994)
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| Budget Amount *help |
¥2,000,000 (Direct Cost: ¥2,000,000)
Fiscal Year 1994: ¥500,000 (Direct Cost: ¥500,000)
Fiscal Year 1993: ¥1,500,000 (Direct Cost: ¥1,500,000)
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| Keywords | Protein Phosphorylation / Signal Transduction / Thyroid Hormone / Two Dimensional Gel Electrophoresis |
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| Research Abstract |
Rat thyroid cultured cell line, FRTL-5 cells, was labelled by ^<32>P and was analyzed by two dimensional gel electrophoresis (2D-gel) after TSH stimulation. One acidic protein (24kd) was phosphorylated and two neutral proteins (19kd) were dephosphorylated. These reactions were reproduced by not TPA,stimulator for C-kinase, but forskolin, stimulator for adenylate cyclase. Therefore, these proteins existed on the signal transduction cascade of A-kinase and may play an important role for thyroid hormone secretion. These results suggested that not only protein kinase but also protein phosphatase took part in the TSH signal transduction.
NIMl cells, human papillary thyroid carcinoma derived cell line, was regulated on cell growth by interleukin-1 (IL-1). Acidic protein (28kd, pl5.5) 00000000was phosphorylated by IL-1alpha stimulation. It was suggested that this protein existed on the signal transduction cascade of C-kinase.
In FRTL-5 cells, fractionation analysis of the phosphorylated protein suggested that membrane fraction contained the protein dominantly. It was solubilized by Triton X-100 and was partially purified by ion exchange chromatography. Now we are trying to purify the spot using 2D-gel transfer to Problot membrane for microsequencing. In near future, we are going to analyze the peptide sequence on GCG program. If it will be an unknown protein we are going to do a screening of cDNA library by degenerate primers.
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