Structure and expression of genes encoding the enzymes of phosphatidylcholine metabolism
| Project/Area Number |
03454149
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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 |
General medical chemistry
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| Research Institution | Gunma University |
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Principal Investigator |
YAMASHITA Satoshi Gunma University, Department of Biochemistry, Professor, 医学部, 教授 (50025623)
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| Co-Investigator(Kenkyū-buntansha) |
SUGIMOTO Hiroyuki Gunma University, Department of Biochemistry, Assistant, 医学部, 助手
UCHIDA Tsutomu Gunma University, Department of Biochemistry, Assistant, 医学部, 助手 (00160276)
HOSAKA Kohei Gunma University, Department of Biochemistry, Instructor, 医学部, 講師 (70108992)
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| Project Period (FY) |
1991 – 1992
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| Project Status |
Completed (Fiscal Year 1992)
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| Budget Amount *help |
¥6,600,000 (Direct Cost: ¥6,600,000)
Fiscal Year 1992: ¥1,300,000 (Direct Cost: ¥1,300,000)
Fiscal Year 1991: ¥5,300,000 (Direct Cost: ¥5,300,000)
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| Keywords | phospholipase D / lysophospholipase / phosphatidic acid / activation / inhibition / enzyme purification / gene expression / yeast phospholipid synthesis / ホスファチジルコリン代謝 / コリンキナ-ゼ / ホスホリパ-ゼC / プロモ-タ- / イノシト-ル・コリン調節 |
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| Research Abstract |
1. Activation of phosphatidylcholine phospholipase D We identified a novel activator of phospholipase D, phosphatidic acid, which is the product of phospholipse D as well. This finding is consistent with the fact that phosphatidic and diacylglycerol formed by various agonist stimulation are maintained at high levels for extended periods of time.
2. Purification of lysophospholipase and inhibition by phosphatidic acid Lysophosphatidylcholine is a physiologically and phamacologically active lipid. Lysopholipase is the one factor which determines the cellular content of this lipid. We purified the enzyme by 214,000-fold from rat liver supernatant and obtained a homogeneous preparation with an apparent molecular weight of 60,000 as determined by SDS- polyacrylamide electrophoresis. The enzyme exhibited transacylase activity as well as hydrolytic activity, indicating the formation of acyl-enzyme as the intermediate. The enzyme activity was strongly inhibited by arachidonic acid and phosphati
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dic acid, the products of phospholipase A_2 and phospholipase D, respectively. Thus, when cells are stimulated by phospholipase D-linked agonists, the phosphatidic acid released from phosphatidylcholine is thought to inhibit lysophospholipase and increase the lysophosphatidylcholine level in the cells, eventually potentiating the effects of the agonists. Thus lysophospholipase may play a role in the signal transduction pathway of the cell.
3. Control of the expression of the genes in yeast phospholipid synthesis Many of the yeast genes of yeast phospholipid synthesis are coordinately regulated by inositol and choline. We constructed various deletions and point mutations in the upstream regions of these genes and examined the effect of these mutations on the expression and control of the genes by gene-transfer assay. The study established that the consensus CATRTGAA sequence is responsible for the regulation of these genes. Novel genes, RIC1 and SCS1, were cloned and sequenced. These genes, very likely to encode transcription factors, were required for the full expression of phospholipid-synthesizine enzymes. Less
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Research Products
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