The target site and role of NDP kinase in cellular signaling networks
| Project/Area Number |
12672149
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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 |
Biological pharmacy
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| Research Institution | Tokyo Metropolitan Institute of Gerontology |
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Principal Investigator |
SHIMADA Nobuko Tokyo Metropolitan Institute of Gerontology, Department of Gene Regulation and Protein Function, Chief, 遺伝子情報部門, 助手 (60158962)
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| Co-Investigator(Kenkyū-buntansha) |
KIMURA Narimichi Tokyo Metropolitan Institute of Gerontology, Department of Gene Regulation and Protein Function, assistant researcher, 遺伝子情報部門, 室長 (60073029)
石嶋 康史 科学技術振興事業団, 科学技術特別研究員
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| Project Period (FY) |
2000 – 2001
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| Project Status |
Completed (Fiscal Year 2001)
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| Budget Amount *help |
¥3,500,000 (Direct Cost: ¥3,500,000)
Fiscal Year 2001: ¥900,000 (Direct Cost: ¥900,000)
Fiscal Year 2000: ¥2,600,000 (Direct Cost: ¥2,600,000)
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| Keywords | NDP kinase / adenylate cyclase / G-protein / PC12D / PACAP / NDPK / 三量体G蛋白 / cAMP / トランスフェクション |
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| Research Abstract |
This study has been done to investigate the functional role of NDPK as a GTP-supplying molecule in cellular signaling system (hormone receptor-G-protein-adenylate cyclase). To this goal, we first utilized dominant negative NDPK-expressing PC12D cells, to see whether the mutant NDPK overexpressed in the cell interferes with the hormonal response. When we compared the PACAP-stimulated cAMP level between PC12D cell clone that was transfected with mutant inactive recombinant NDPK (rNDPKmα or mβ) and control cell clone, the stimulation by PACAP tended to be suppressed in the transfected cell with rNDPKα compared with the control cell. However, this suppression varied from experiment to experiment.
The plasma membranes from PC12D cells contain NDPK activity which acts to form GTP from added GDP, leading to AC activation by PACAP plus GDP. Therefore, we then examined whether the NDPKmα mixed with the plasma membranes can affect the endogenous NDPK and, as a result, alter the cAMP formation stimulated by PACAP. The result showed no obvious effect by NDPKmα. Also, the AC activity in membranes from PC12D-rNDPKmα stimulated by PACAP plus GDP did not differ from that from the contol cells.
As we demonstrated previously, however, the PACAP-stimulated AC activity of the purified membranes was completely dependent on GTP, and the effect of PACAP plus GDP was supressed by UDP as an inhibitor of NDPK. Therefore, it is clear that PC12D AC system is closely associated with NDPK as demonstrated for that of rat liver membrane. It remains to be studied whether the plasma membrane-associated NDPK is derived from the cytosolic one, or it is a novel membrane specific isoform distinct from the cytosolic NDPKs.
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Report
(3 results)
Research Products
(3 results)
