title: Rice CYP734As function as multisubstrate and multifunctional enzymes in brassinosteroid catabolism.
abstract: Catabolism of brassinosteroids regulates the endogenous level of bioactive brassinosteroids. In Arabidopsis thaliana, bioactive brassinosteroids such as castasterone (CS) and brassinolide (BL) are inactivated mainly by two cytochrome P450 monooxygenases, CYP734A1/BAS1 and CYP72C1/SOB7/CHI2/SHK1; CYP734A1/BAS1 inactivates CS and BL by means of C-26 hydroxylation. Here, we characterized CYP734A orthologs from Oryza sativa (rice). Overexpression of rice CYP734As in transgenic rice gave typical brassinosteroid-deficient phenotypes. These transformants were deficient in both the bioactive CS and its precursors downstream of the C-22 hydroxylation step. Consistent with this result, recombinant rice CYP734As utilized a range of C-22 hydroxylated brassinosteroid intermediates as substrates. In addition, rice CYP734As can catalyze hydroxylation and the second and third oxidations to produce aldehyde and carboxylate groups at C-26 in vitro. These results indicate that rice CYP734As are multifunctional, multisubstrate enzymes that control the endogenous bioactive brassinosteroid content both by direct inactivation of CS and by the suppression of CS biosynthesis by decreasing the levels of brassinosteroid precursors.
authors.affiliation: Institute for Advanced Research, Nagoya University, Nagoya, Aichi 464-8601, Japan. sakamoto@iar.nagoya-u.ac.jp
authors.forename: Tomoaki
authors.initials: T
authors.lastname: Sakamoto
authors.forename: Ayami
authors.initials: A
authors.lastname: Kawabe
authors.forename: Asako
authors.initials: A
authors.lastname: Tokida-Segawa
authors.forename: Bun-Ichi
authors.initials: B
authors.lastname: Shimizu
authors.forename: Suguru
authors.initials: S
authors.lastname: Takatsuto
authors.forename: Yukihisa
authors.initials: Y
authors.lastname: Shimada
authors.forename: Shozo
authors.initials: S
authors.lastname: Fujioka
authors.forename: Masaharu
authors.initials: M
authors.lastname: Mizutani
mesh_terms: D000818:Animals; D016367:Baculoviridae / Q000235:genetics / Q000378:metabolism; D060406:Brassinosteroids / Q000032:analysis / Q000378:metabolism*; D002460:Cell Line; D002777:Cholestanols / Q000032:analysis / Q000378:metabolism; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics / Q000378:metabolism*; D018076:DNA, Complementary / Q000235:genetics; D015971:Gene Expression Regulation, Enzymologic; D018506:Gene Expression Regulation, Plant; D006900:Hydroxylation; D009154:Mutation; D012275:Oryza / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D010084:Oxidation-Reduction; D010641:Phenotype; D010802:Phylogeny; D030821:Plants, Genetically Modified / Q000201:enzymology / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics; D018749:RNA, Plant / Q000235:genetics; D018411:Spodoptera / Q000821:virology; D013260:Steroids, Heterocyclic / Q000032:analysis / Q000378:metabolism; D013379:Substrate Specificity
chemical_list: D060406:Brassinosteroids; D002777:Cholestanols; D018076:DNA, Complementary; D012333:RNA, Messenger; D018749:RNA, Plant; D013260:Steroids, Heterocyclic; C083545:castasterone; D003577:Cytochrome P-450 Enzyme System; C023623:brassinolide