| Project/Area Number |
11640650
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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 |
植物生理
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| Research Institution | KYUSHU UNIVERSITY |
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Principal Investigator |
SHIMAZAKI Kenichiro KYUSHU UNIVERSITY, Faculty of science, Prof., 大学院・理学研究院, 教授 (00124347)
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| Co-Investigator(Kenkyū-buntansha) |
TOSHINORI Kinoshita KYUSHU UNIVERSITY, Faculty of science, assist., 大学院・理学研究院, 助手 (50271101)
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| Project Period (FY) |
1999 – 2000
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| Project Status |
Completed (Fiscal Year 2000)
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| Budget Amount *help |
¥3,900,000 (Direct Cost: ¥3,900,000)
Fiscal Year 2000: ¥1,000,000 (Direct Cost: ¥1,000,000)
Fiscal Year 1999: ¥2,900,000 (Direct Cost: ¥2,900,000)
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| Keywords | Stomata / 14-3-3 protein / blue light response / H^+-ATPase / protein phosphosylation / 孔辺細胞 / 青色光反応 / 14-3-3タンパク質 / リン酸化 / ソラマメ / 青色光 / H^+-ATPase |
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| Research Abstract |
The plasma membrane H^+-ATPase is activated by blue light with concomitant binding of the 14-3-3 protein to the C-terminus in guard cells. Since several isoforms of the 14-3-3 protein are expressed in plants, we determined which isoform (s) bound to the H^+-ATPase in vivo. Four cDNA clones (vf14-3-3a, vf14-3-3b, vf14-3-3c and vf14-3-3d) encoding 14-3-3 proteins were isolated from Vicia guard cells. Northern analysis revealed that mRNAs encoding vf14-3-3a and vf14-3-3b proteins were expressed predominantly in guard cells. The 14-3-3 protein that bound to the H^+-ATPase in guard cells had the same molecular mass as the recombinant vf14-3-3a protein. The H^+-ATPase immunoprecipitated from mesophyll cell protoplasts (MCPs), which had been stimulated by fusicoccin, co-precipitated with the 32.5-kDa 14-3-3 protein, although three 14-3-3 isoproteins were found in MCPs. Digestions of the bound 14-3-3 protein and recombinant vf14-3-3a with CNBr gave the identical migration profiles on SDS-PAGE, but that of vf14-3-3b gave a different profile. Mass profiling of trypsin-digested 14-3-3 protein bound to the H^+-ATPase gave the predicted peptide masses of vf14-3-3a. Far Western analysis revealed that the H^+-ATPase had a higher affinity for vf14-3-3a than for vf14-3-3b. These results suggest that the 14-3-3 protein that bound to the plasma membrane H^+-ATPase in vivo is vf14-3-3a and that it may play a key role in the activation of H^+-ATPase in guard cells.
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