title: A metabolic gene cluster in Lotus japonicus discloses novel enzyme functions and products in triterpene biosynthesis.
abstract: Genes for triterpene biosynthetic pathways exist as metabolic gene clusters in oat and Arabidopsis thaliana plants. We characterized the presence of an analogous gene cluster in the model legume Lotus japonicus. In the genomic regions flanking the oxidosqualene cyclase AMY2 gene, genes for two different classes of cytochrome P450 and a gene predicted to encode a reductase were identified. Functional characterization of the cluster genes was pursued by heterologous expression in Nicotiana benthamiana. The gene expression pattern was studied under different developmental and environmental conditions. The physiological role of the gene cluster in nodulation and plant development was studied in knockdown experiments. A novel triterpene structure, dihydrolupeol, was produced by AMY2. A new plant cytochrome P450, CYP71D353, which catalyses the formation of 20-hydroxybetulinic acid in a sequential three-step oxidation of 20-hydroxylupeol was characterized. The genes within the cluster are highly co-expressed during root and nodule development, in hormone-treated plants and under various environmental stresses. A transcriptional gene silencing mechanism that appears to be involved in the regulation of the cluster genes was also revealed. A tightly co-regulated cluster of functionally related genes is involved in legume triterpene biosynthesis, with a possible role in plant development.
authors.affiliation: Department of Biochemistry and Biotechnology, University of Thessaly, Ploutonos 26 & Aeolou Str., Larisa, 41221, Greece.
authors.forename: Afrodite
authors.initials: A
authors.lastname: Krokida
authors.affiliation: Department of Biochemistry and Biotechnology, University of Thessaly, Ploutonos 26 & Aeolou Str., Larisa, 41221, Greece.
authors.forename: Costas
authors.initials: C
authors.lastname: Delis
authors.affiliation: Department of Metabolic Biology, John Innes Centre, Colney Lane, Norwich, NR4 7UH, UK.
authors.forename: Katrin
authors.initials: K
authors.lastname: Geisler
authors.affiliation: Department of Biochemistry and Biotechnology, University of Thessaly, Ploutonos 26 & Aeolou Str., Larisa, 41221, Greece.
authors.forename: Constantine
authors.initials: C
authors.lastname: Garagounis
authors.affiliation: Department of Biochemistry and Biotechnology, University of Thessaly, Ploutonos 26 & Aeolou Str., Larisa, 41221, Greece.
authors.forename: Daniela
authors.initials: D
authors.lastname: Tsikou
authors.affiliation: Unidad de Biotecnología, Centro de Investigación Científica de Yucatán, Mérida, Yucatán, México.
authors.forename: Luis M
authors.initials: LM
authors.lastname: Peña-Rodríguez
authors.affiliation: Department of Biochemistry and Biotechnology, University of Thessaly, Ploutonos 26 & Aeolou Str., Larisa, 41221, Greece.
authors.forename: Dimitra
authors.initials: D
authors.lastname: Katsarou
authors.affiliation: Department of Metabolic Biology, John Innes Centre, Colney Lane, Norwich, NR4 7UH, UK.
authors.forename: Ben
authors.initials: B
authors.lastname: Field
authors.affiliation: Department of Metabolic Biology, John Innes Centre, Colney Lane, Norwich, NR4 7UH, UK.
authors.forename: Anne E
authors.initials: AE
authors.lastname: Osbourn
authors.affiliation: Department of Biochemistry and Biotechnology, University of Thessaly, Ploutonos 26 & Aeolou Str., Larisa, 41221, Greece.
authors.forename: Kalliope K
authors.initials: KK
authors.lastname: Papadopoulou
mesh_terms: D003577:Cytochrome P-450 Enzyme System / Q000235:genetics*; D015870:Gene Expression; D018506:Gene Expression Regulation, Plant; D020868:Gene Silencing; D017343:Genes, Plant; D000070116:Lotus / Q000201:enzymology / Q000235:genetics* / Q000378:metabolism; D063245:Plant Development / Q000235:genetics*; D010940:Plant Proteins / Q000235:genetics*; D055170:Plant Root Nodulation / Q000235:genetics; D018517:Plant Roots / Q000254:growth & development; D053204:Root Nodules, Plant / Q000254:growth & development; D014315:Triterpenes / Q000378:metabolism*
chemical_list: D010940:Plant Proteins; D014315:Triterpenes; D003577:Cytochrome P-450 Enzyme System
keywords: Lotus japonicus; cytochrome P450; nodule; transcriptional silencing; triterpene
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