Camptothecin biosynthetic genes in hairy roots of Ophiorrhiza pumila: Cloning, characterization and differential expression in tissues and by stress compounds

• Published in: Plant and cell physiology Vol. 44; no. 4; pp. 395 - 403
• Main Authors: Yamazaki, Y. (Chiba Univ. (Japan)), Sudo, H, Yamazaki, M, Aimi, N, Saito, K
• Format: Journal Article
• Language: English
• Published: Japan Oxford University Press 01-04-2003; Oxford Publishing Limited (England)
• Subjects: AB086168 (OpTDC) and AB086169 (OpCPR); Acetates - pharmacology; Adenine - analogs & derivatives; Adenine - pharmacology; ANTINEOPLASTIC AGENTS; Aromatic-L-Amino-Acid Decarboxylases - genetics; Aromatic-L-Amino-Acid Decarboxylases - metabolism; Base Sequence; Benzyl Compounds; BIOSYNTHESIS; Camptothecin - biosynthesis; Carbon-Nitrogen Lyases - genetics; Carbon-Nitrogen Lyases - metabolism; Cloning, Molecular; Cyclopentanes - pharmacology; CYTOCHROME P450; DNA, Complementary - chemistry; DNA, Complementary - genetics; Escherichia coli - genetics; Escherichia coli - metabolism; GENE EXPRESSION; Gene Expression Regulation, Enzymologic - drug effects; Gene Expression Regulation, Plant - drug effects; Keywords: Anti-tumor alkaloid — Camptothecin — NADPH:cytochrome P450 reductase — Ophiorrhiza pumila — Strictosidine synthase — Tryptophan decarboxylase; Kinetin; Molecular Sequence Data; NADPH-Ferrihemoprotein Reductase - genetics; NADPH-Ferrihemoprotein Reductase - metabolism; Naphthaleneacetic Acids - pharmacology; OXIDOREDUCTASES; Oxylipins; Phylogeny; Plant Roots - drug effects; Plant Roots - genetics; Plant Roots - metabolism; Purines; ROOT HAIRS; RUBIACEAE; Rubiaceae - drug effects; Rubiaceae - genetics; Rubiaceae - metabolism; Salicylic Acid - pharmacology; Sequence Analysis, DNA; The nucleotide sequences in this paper have been submitted to DDBJ/GenBank/EMBL under accession numbers AB060341 (OpSTR)
• ISSN: 0032-0781; 1471-9053
• DOI: 10.1093/pcp/pcg051

Camptothecin derivatives are clinically used antitumor compounds that biogenetically belong to a group of monoterpenoid indole alkaloids (TIA). We have already established a hairy root culture of Ophiorrhiza pumila (Rubiaceae) that produces camptothecin. The present study describes the cloning and characterization of cDNAs encoding strictosidine synthase (OpSTR; EC 4.3.3.2) and tryp-tophan decarboxylase (OpTDC; EC 4.1.1.28), two key enzymes in the biosynthesis of TIA from hairy roots of O. pumila. We also isolated the cDNA coding for NADPH: cytochrome P450 reductase (OpCPR; EC 1.6.2.4) that is presumed to be indirectly involved in camptothecin synthesis. The recombinant OpSTR and OpTDC proteins exhibit STR and TDC activities, respectively, when expressed in Escherichia coli. The tissue-specific and stress-inducible expression patterns of OpSTR and OpTDC were quite similar, unlike those of OpCPR. The high expression of OpSTR and OpTDC observed in hairy roots, roots and stems were closely correlated with STR protein accumulation as observed by immunoblot analysis. Plant stress compounds like salicylic acid repressed expression of OpSTR and OpTDC, suggesting coordinate regulation of these genes for camptothecin biosynthesis.