Molecular Mechanism of the Biosynthesis of Flavonoids Involved in Leguminous Plant-Microbe Interactions

• Project/Area Number: 07640872
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Single-year Grants
• Section: 一般
• Research Field: 植物生理
• Research Institution: Nihon University
• Principal Investigator: AYABE Shinichi Nihon University ; College of Bioresource Sciences ; Associate Professor, 生物資源科学部, 助教授 (40050679)
• Co-Investigator(Kenkyū-buntansha): AOKI Toshio Nihon University ; College of Bioresource Sciences ; Assistant Professor, 生物資源科学部, 助手 (80287606) ; 古野 哲郎 日本大学, 農獣医学部, 助手 (70202299)
• Project Period (FY): 1995 – 1996
• Project Status: Completed (Fiscal Year 1996)
• Budget Amount: ¥2,000,000 (Direct Cost: ¥2,000,000); Fiscal Year 1996: ¥1,000,000 (Direct Cost: ¥1,000,000); Fiscal Year 1995: ¥1,000,000 (Direct Cost: ¥1,000,000)
• Keywords: Plant-microbe interactions / Leguminosae / Flavonoids / Cloning / Glycyrrhiza echinata / 6'-Deoxychalcone synthase / Cytochrome P450 / O-Methyltransferase / 6'-Deoxychalcone synthase / 生合成 / 酵素

Research Abstract

Flavonoids of leguminous plants play important roles in pathogenic and symbiotic plant-microbe interactions. The reaction mechanisms and physiological functions of the key enzymes in the biosynthesis of these biologically-active flavonoids were explored.
1.6'-Deoxychalcone synthase (DOCS) : Major biologically-active flavonoids are 5-deoxy (iso) flavonoids. They are derived from 6'-deoxychalcone, which is synthesized by the co-action of chalcone synthase (CHS) and a polyketide reductase (PKR). From the cDNA library of elicitor-induced licorice (Glycyrrhiza echinata) cells, cDNAs for both component enzymes were cloned. Catalytic activity of PKR was demonstrated by the combination of the recombinant enzyme and cell-free extracts of anthocyanin-producing dandelion cells. Co-expression of CHS and PKR in E.coli will be beneficial to elucidate the molecular mechanism of DOCS reaction.
2. O-Methyltransferases (OMT) : A methoxychalcone of alfalfa roots is a strong chemical signal toward a soil be cterium to form symbiotic nitrogen-fixing root nodules. A chalcone OMT (CHMT) of alfalfa in methoxychalcone biosynthesis displayd additional licodione methyltransferase (LMT) activity, which is expressed in licorice cells as a part of defense reactions caused by elicitor treatment. From the licorice cDNA library an OMT cDNA was cloned, the translated protein of which had both LMT and CHMT activities. To understand the roles of OMTs in licorice, purification of LMT devoid of CHMT activity is underway.
3.Cytochrome P450s (P450s) : Distinct P450s acting on a common substrate (liquiritigenin) are responsible for the biosynthetic branching to end products with different physiological functions. PCR fragments of P450 cDNAs were isolated from the licorice cDNA library. Further screening yielded full-length P450 cDNAs, amongwhich were cinnamate 4-hydroxylase cDNA and functionally-unknown clones. Their expression in response to elicitor-treatment indicated that some of the clones are involved in plant-microbe interactions.

Research Products

• [Publications] T.Akashi et al.: "A cDNA for polyketide reductase (accession No.D83718) that catalyzes the formation of 6′-deoxychalcone from cultured Glycyrrhiza echinata L. cells (PGR96-023)" Plant Physiology. 111(1). 347- (1996)
  • Description: 「研究成果報告書概要(和文)」より
• [Publications] M.Ichimura et al.: "Enzymatic O-methylation of isoliquiritigenin and licodione in alfalfa and licorice cultures" Phytochemistry. (in press). (1997)
  • Description: 「研究成果報告書概要(和文)」より
• [Publications] M.Haga et al.: "A cDNA for S-adenosyl-L-methionine : isoliquiritigenin / licodione 2′-O-methyltransferase (accession No.D88742) from cultured licorice (Glycyrrhiza echinata L.) cells (PGR97-014)" Plant Physiology. 113(2). 663- (1997)
  • Description: 「研究成果報告書概要(和文)」より
• [Publications] T.Akashi et al.: "Cloning of cytochrome P450 cDNAs from cultured Glycyrrhiza echinata L. cells and their transcriptional activation by elicitor-treatment" Plant Science. (in press). (1997)
  • Description: 「研究成果報告書概要(和文)」より
• [Publications] Akashi, T.et al.: "A cDNA for polyketide reductase (accession No.D83718) that catalyzes the formation of 6'-deoxychalcone from cultured Glycyrrhiza echinata L.cells (PGR96-023)" Plant Physiology. 111 (1). 347 (1996)
  • Description: 「研究成果報告書概要(欧文)」より
• [Publications] Ichimura, M.et al.: "Enzymatic O-methylation of isoliquiritigenin and licodione in alfalfa and licorice cultures." Phytochemistry. (in press). (1997)
  • Description: 「研究成果報告書概要(欧文)」より
• [Publications] Haga, M.et al.: "A cDNA for S-adenosyl-L-methionine : isoliquiritigenin/licodione 2'-O-methyltransferase (accession No.D88742) from cultured licorice (Glycyrrhiza echinata L.) cells (PGR97-014)" Plant Physiology. 113 (2). 663 (1997)
  • Description: 「研究成果報告書概要(欧文)」より
• [Publications] Akashi, T.et al.: "Cloning of cytochrome P450 cDNAs from cultured Glycyrrhiza echinata L.cells and their transcriptional activation by elicitor-treatment" Plant Science. (in press). (1997)
  • Description: 「研究成果報告書概要(欧文)」より
• [Publications] T.Akashi et al.: "A cDNA for polyketide rductase (accession No.D83718) that catalyzes the formation of 6'-deoxychalcone from cultured Glycyrrhiza echinata L.cells (PGR96-023)" Plant Physiology. 111(1). 347- (1996)
• [Publications] M.Ichimura et al.: "Enzymatic O-methylation of isoliquiritigenin and licodione in alfalfa and licorice cultures" Phytochemistry. (in press). (1997)
• [Publications] M.Haga et al.: "A cDNA for S-adenosyl-L-methionine : isoliquiritigenin/licodione 2'-O-methyltransferase (accession No.D88742) from cultured licorice (Glycyrrhiza echinata L.) cells (PGR97-014)" Plant Physiology. (in press). (1997)
• [Publications] T.Akashi et al.: "Cloning of cytochrome P450 cDNAs from cultured Glycyrrhiza echinata L.cells and their transcriptional activation by elicitor-treatment" Plant Science. (in press). (1997)