Dataset Information

Bottlenecks for metabolic engineering of isoflavone glycoconjugates in Arabidopsis.

ABSTRACT: In view of their perceived chemopreventive activities against hormone-dependent cancers, cardiovascular disease, and postmenopausal ailments, there is considerable interest in engineering plants to contain isoflavone phytoestrogens. However, attempts to date have only resulted in low levels of isoflavone accumulation in non-legumes. Introducing soybean isoflavone synthase (IFS) into Arabidopsis thaliana leads to accumulation of low levels of genistein glycosides. Leaves of wild-type A. thaliana contain high levels of similar conjugates of the flavonols quercetin and kaempferol, which could be increased by threefold on introduction of an alfalfa chalcone isomerase transgene. Levels of genistein were not increased by expressing both IFS and alfalfa chalcone isomerase, but levels of flavonol conjugates were reduced to a greater extent than could be accounted for by flux into isoflavone. Introduction of IFS into the tt6/tt3 double mutant blocked in flavonol, and anthocyanin synthesis resulted in high levels of genistein. The bottleneck for constitutive isoflavone production in Arabidopsis is, therefore, competition for flavanone between IFS and endogenous flavonol synthesis, and the flavonol pathway is reciprocally but disproportionately affected by IFS.

SUBMITTER: Liu CJ

PROVIDER: S-EPMC137925 | biostudies-literature | 2002 Oct

REPOSITORIES: biostudies-literature

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Bottlenecks for metabolic engineering of isoflavone glycoconjugates in Arabidopsis.

Liu Chang-Jun CJ Blount Jack W JW Steele Christopher L CL Dixon Richard A RA

Proceedings of the National Academy of Sciences of the United States of America 20021016 22

In view of their perceived chemopreventive activities against hormone-dependent cancers, cardiovascular disease, and postmenopausal ailments, there is considerable interest in engineering plants to contain isoflavone phytoestrogens. However, attempts to date have only resulted in low levels of isoflavone accumulation in non-legumes. Introducing soybean isoflavone synthase (IFS) into Arabidopsis thaliana leads to accumulation of low levels of genistein glycosides. Leaves of wild-type A. thaliana ...[more]

PMID: 12384577

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Project description:Terpenoids are a large and diverse class of plant metabolites that also includes volatile mono- and sesquiterpenes which are involved in biotic interactions of plants. Due to the limited natural availability of these terpenes and the tight regulation of their biosynthesis, there is strong interest to introduce or enhance their production in crop plants by metabolic engineering for agricultural, pharmaceutical and industrial applications. While engineering of monoterpenes has been quite successful, expression of sesquiterpene synthases in engineered plants frequently resulted in production of only minor amounts of sesquiterpenes. To identify bottlenecks for sesquiterpene engineering in plants, we have used two nearly identical terpene synthases, snapdragon (Antirrhinum majus) nerolidol/linalool synthase-1 and -2 (AmNES/LIS-1/-2), that are localized in the cytosol and plastids, respectively. Since these two bifunctional terpene synthases have very similar catalytic properties with geranyl diphosphate (GPP) and farnesyl diphosphate (FPP), their expression in target tissues allows indirect determination of the availability of these substrates in both subcellular compartments. Both terpene synthases were expressed under control of the ripening specific PG promoter in tomato fruits, which are characterized by a highly active terpenoid metabolism providing precursors for carotenoid biosynthesis. As AmNES/LIS-2 fruits produced the monoterpene linalool, AmNES/LIS-1 fruits were found to exclusively produce the sesquiterpene nerolidol. While nerolidol emission in AmNES/LIS-1 fruits was 60- to 584-fold lower compared to linalool emission in AmNES/LIS-2 fruits, accumulation of nerolidol-glucosides in AmNES/LIS-1 fruits was 4- to 14-fold lower than that of linalool-glucosides in AmNES/LIS-2 fruits. These results suggest that only a relatively small pool of FPP is available for sesquiterpene formation in the cytosol. To potentially overcome limitations in sesquiterpene production, we transiently co-expressed the key pathway-enzymes hydroxymethylglutaryl-CoA reductase (HMGR) and 1-deoxy-D-xylulose 5-phosphate synthase (DXS), as well as the regulator isopentenyl phosphate kinase (IPK). While HMGR and IPK expression increased metabolic flux toward nerolidol formation 5.7- and 2.9-fold, respectively, DXS expression only resulted in a 2.5-fold increase.

Dataset Information

Bottlenecks for metabolic engineering of isoflavone glycoconjugates in Arabidopsis.

Publications

Bottlenecks for metabolic engineering of isoflavone glycoconjugates in Arabidopsis.

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