Scopoletin is biosynthesized viaortho‐hydroxylation of feruloyl CoA by a 2‐oxoglutarate‐dependent dioxygenase inArabidopsis thaliana

Abstract

SummaryCoumarins are derived via the phenylpropanoid pathway in plants. The 2H-1-benzopyran-2-one core structure of coumarins is formed via the ortho-hydroxylation of cinnamates, trans/cis isomerization of the side chain, and lactonization. Ortho-hydroxylation is a key step in coumarin biosynthesis as a branch point from lignin biosynthesis; however, ortho-hydroxylation of cinnamates is not yet fully understood. In this study, scopoletin biosynthesis was explored using Arabidopsis thaliana, which accumulates scopoletin and its b-glucopyranoside scopolin in its roots. T-DNA insertion mutants of caffeoyl CoA O-methyltransferase 1 (CCoAOMT1) showed significant reduction in scopoletin and scopolin levels in the roots, and recombinant CCoAOMT1 exhibited 3¢-O-methyltransferase activity on caffeoyl CoA to feruloyl CoA. These results suggest that feruloyl CoA is a key precursor in scopoletin biosynthesis. Ortho-hydroxylases of cinnamates were explored in the oxygenase families in A. thaliana, and one of the candidate genes in the Fe(II)-and 2-oxoglutaratedependent dioxygenase (2OGD) family was designated as F6¢H1. T-DNA insertion mutants of F6¢H1 showed severe reductions in scopoletin and scopolin levels in the roots. The pattern of F6¢H1 expression is consistent with the patterns of scopoletin and scopolin accumulation. The recombinant F6¢H1 protein exhibited orthohydroxylase activity for feruloyl CoA (K m = 36.0 AE 4.27 lM; k cat = 11.0 AE 0.45 sec )1 ) to form 6¢-hydroxyferuloylCoA, but did not hydroxylate ferulic acid. These results indicate that Fe(II)-and 2-oxoglutarate-dependent dioxygenase is the pivotal enzyme in the ortho-hydroxylation of feruloyl CoA in scopoletin biosynthesis.

iTRAQ Protein Profile Analysis of Arabidopsis Roots Reveals New Aspects Critical for Iron Homeostasis

“…Strong up-regulation at the transcript and protein levels was also noted for F6#H1, an oxidoreductase that was shown to be involved in coumarin synthesis by catalyzing the synthesis of 6#-hydroxyferuloyl-CoA from feruloyl-CoA (Kai et al, 2008). Caffeoyl-CoA Figure 5.…”

“…5). Mutations in caffeoyl-CoA O-methyltransferase and in F6#H1 were shown to cause reductions in the levels of scopoletin and its b-glucoside scopolin in the roots (Kai et al, 2008), implying that both enzymes are involved in the biosynthesis of scopoletin. To prove whether scopoletin and/or scopolin levels were in- Figure 6.…”

Isolation and Functional Characterization of CYP71AJ4 Encoding for the First P450 Monooxygenase of Angular Furanocoumarin Biosynthesis

“…6 for a review). However, a dioxygenase has now been cloned from Arabidopsis and expressed; this enzyme converts feruloyl-CoA to 2,4-dihydroxy-5-methoxycinnamoyl-CoA and scopoletin (18). To a lesser extent, it also accepts 4-coumaroylCoA, which yields umbelliferone (Fig.…”

Plant strategies to deal with a combination of drought and high temperatures

“…Among these metabolites, coumarin scopoletin, derived from feruloyl-CoA, is converted into scopolin by glycosylation, and both metabolites are likely to be involved in plant responses to stress (Kai et al 2008) On the whole, our data indicate that Cleopatra, as a susceptible citrus genotype under WS+HS situations (Zandalinas et al 2016), deeply alters its primary metabolism in order to face the damaging effects caused by stress combination by activating glycolysis and TCA cycle to support cellular energy requirements as well as phenylpropanoid-derived scopolin. On the other hand, Carrizo shows reduced modifications in primary metabolism, associated to a lower incidence of stress-induced damage.…”

“…On the other hand, Carrizo accumulated sucrose via gluconeogenesis in response to heat stress and the combination of drought and heat stress, probably related to higher photosynthetic rate observed previously and constituting hence an advantage under these conditions. In addition, phenylpropanoid pathway intermediates are reported to be involved in plant responses to stress (Kai et al 2008). In this sense, whereas Cleopatra leaves accumulated scopolin especially under combined conditions, Carrizo induced the accumulation of sinapic acid and derivatives, direct precursors of lignins.…”