1990 Fiscal Year Final Research Report Summary
Intracellular Calcium Ion Signal Transduction System in Plant
| Project/Area Number |
01480010
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| Research Category |
Grant-in-Aid for General Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Research Field |
植物生理学
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| Research Institution | University of Tokyo |
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Principal Investigator |
TORIYAMA Shoshi Univ. of Tokyo, Inst. of Applied Microbiol., Associate Professor, 応用微生物研究所, 助教授 (40013338)
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| Project Period (FY) |
1989 – 1990
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| Keywords | Calcium ion / Phosphoinositides / Phospholipase C / Inositol trisphosphate / Ca^<2+>-pump / Ca^<2+>-channel / Ca^<2+>-dependent protein kinase / Patch-clamp method |
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| Research Abstract |
(1) Enzymes involved in metabolic turnover of inositolphospholipid, phosphatidyl inositol kinase, phosphatidylinositol-monophosphate kinase and phospholipase C (PLC) were localized in the plasma membrane of tobacco culture cells. The kinases were significantly inhibited, whereas PLC was markedly activated by 0.1-1 uM Ca^<2+>. A voltage-dependent outward-rectifying Ca^<2+> current was recorded with an outside-out patch of the tonoplast membrane by patch-clamp method. This Ca^<2+> channel was markedly activated by I uM inositol trisphosphate (IP). These results suggest that IP_3, the product of PLC catalysis causes Ca^<2+> release from the vacuole and results in the increase of cytosolic Ca^<2+> co cncentration. The increased Ca^<2+> inhibits the kinases, lowering the supply of substrate to PLC, and results in the decrease in IP_3 production. By such a way, inositolphospholipid metabolism is under the feedback regulation by Ca^<2+>.
(2) Ca^<2+>-pump ATPase was solubilized from corn leaf plasma membrane, and separated by an ion exchange HPLC. This was reconstituted in a liposome. The proteoliposome actively took up Ca2 by an ATP-dependent manner, suggesting that the Ca^<2+>-ATPase actually functioned as the Ca^<2+>-pump.
(3) Ca^<2+>-dependent protein kinase (CDPK) was purified to 900-fold from a halophilic alga Dunaliella. CDPK was markedly activated by 0.1-1 uM Ca^<2+> and increased its hydrophobicity. The enzyme was soluble in the absense of Ca^<2+>, while bound to the membrane fraction from cells in the presense of Ca^<2+>. A little protein in the soluble fraction was phosphorylated, meanwhile a number of proteins in the membrane fraction were significantly phosphorylated by CDPK. This result suggests that CDPK is activated by the increase of cytoplasmic Ca^<2+> concentration and translocated to the membrane where the substrate proteins are present. The involvement of CDPK in osmoregulation of Dunaliella is expected.
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