Camptothecin biosynthetic genes in hairy roots of Ophiorrhiza pumila: cloning, characterization and differential expression in tissues and by stress compounds.
Camptothecin derivatives are clinically used anti-tumor 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 tryptophan 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.
Graduate School of Pharmaceutical Sciences, Chiba University, 1-33 Yayoi-cho, Inage-ku, Chiba, 263-8522 Japan.
Yasuyo
Y
Yamazaki
Hiroshi
H
Sudo
Mami
M
Yamazaki
Norio
N
Aimi
Kazuki
K
Saito
D000085:Acetates / Q000494:pharmacology; D000225:Adenine / Q000031:analogs & derivatives* / Q000494:pharmacology; D001142:Aromatic-L-Amino-Acid Decarboxylases / Q000235:genetics / Q000378:metabolism; D001483:Base Sequence; D001593:Benzyl Compounds; D002166:Camptothecin / Q000096:biosynthesis*; D019759:Carbon-Nitrogen Lyases / Q000235:genetics / Q000378:metabolism; D003001:Cloning, Molecular; D003517:Cyclopentanes / Q000494:pharmacology; D018076:DNA, Complementary / Q000737:chemistry / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D018506:Gene Expression Regulation, Plant / Q000187:drug effects; D007701:Kinetin; D008969:Molecular Sequence Data; D009251:NADPH-Ferrihemoprotein Reductase / Q000235:genetics / Q000378:metabolism; D009280:Naphthaleneacetic Acids / Q000494:pharmacology; D054883:Oxylipins; D010802:Phylogeny; D018517:Plant Roots / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism; D011687:Purines; D019666:Rubiaceae / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism; D020156:Salicylic Acid / Q000494:pharmacology; D017422:Sequence Analysis, DNA
D000085:Acetates; D001593:Benzyl Compounds; D003517:Cyclopentanes; D018076:DNA, Complementary; D009280:Naphthaleneacetic Acids; D054883:Oxylipins; D011687:Purines; C034182:1-naphthaleneacetic acid; C072239:methyl jasmonate; D009251:NADPH-Ferrihemoprotein Reductase; D001142:Aromatic-L-Amino-Acid Decarboxylases; D019759:Carbon-Nitrogen Lyases; C022684:strictosidine synthetase; D000225:Adenine; C480551:benzylaminopurine; D020156:Salicylic Acid; D007701:Kinetin; D002166:Camptothecin