2001 Fiscal Year Final Research Report Summary
The role of microtubules in the regulation of the signal transduction through the interaction with the G protein-coupled receptor kinase
| Project/Area Number |
12670118
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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 |
General medical chemistry
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| Research Institution | Sapporo Medical University |
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Principal Investigator |
HATTA Shinichi Sapporo Medical University, Pharmacology, Associate Professor, 医学部, 助教授 (60094223)
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| Co-Investigator(Kenkyū-buntansha) |
HORIO Yoshiyuki Sapporo Medical University, Pharmacology, Professor, 医学部, 教授 (30181530)
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| Project Period (FY) |
2000 – 2001
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| Keywords | microtubules / GRK / Phosphorylation / NG108-15 cells / tubulin / G protein |
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| Research Abstract |
Microtubules have been suggested to be involved in the regulation of the signal transduction through the interaction with several kinases. Recent studies demonstrated that tubulin (Tu) is a G protein-coupled receptor kinase (GRK) 2-binding protein and a GRK2 substrate. This research examined the role of phosphorylated Tu in the regulation of the signal transduction. Western blot analysis of microtubule preparation purified from rat brain revealed an enrichment of both GRK2 and Tu content. Furthermore, GRK2 and β-Tu III, but not GRK5, were detected in the plasma membrane of NG108-15 cells. When NG108-15 cells were incubated with carbachol, the contents of β-Tu III and GRK2 in the membrane fraction of the cells were increased and reached a maximum at 10 min after carbachol stimulation. Heparin, an inhibitor of GRK2, prevented carbachoHnduced transfer of β-Tu III and GRK2 to the membrane. Tu is a GTP-binding protein. GTP-bound Tu has been suggested to participate in the regulation of phospholipase Cβ signal transduction through the interaction and the transfer of GTP between Tu and Gq. Addition of [^<32>P]AAGTP-liganded Tu to NG108-15 cell membranes resulted in incorporation of AAGTP in Gα, indicating the GTP transfer from Tu to Gα. Carbachol increased IP_3 formation in NG108-15 cells and this effect was inhibited by the pretreatment of the cell with heparin. These results indicate that β-Tu III phosphorylated by GRK2 associates to the plasma membrane and participates in the Gq-mediated signal transduction. Thus, it is conceivable that a link between GRK2 and Tu may be important for the regulation of G protein-coupled receptor signal transduction.
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