Identification of a Functional Flavanone 3-Hydroxylase Gene in a Fragrant Wild Cyclamen Species (Cyclamen purpurascens)

Abstract

Although the fragrant wild cyclamen, Cyclamen purpurascens, is widely used as a parent plant for breeding of fragrant cyclamen cultivars, the color variety of fragrant cyclamen is limited. To develop cultivars with a wider range of flower colors, it is crucial to understand the flavonoid biosynthetic pathway involved in flower color in C. purpurascens as a crossbreeding parent. In this study, we isolated the gene (CpurF3H) encoding flavanone 3-hydroxylase, a key enzyme in determining flower color in cyclamen, from C. purpurascens for the first time and investigated its expression and function. The full-length cDNA sequence of CpurF3H obtained from young petals contained a 1,113-bp open reading frame (ORF) encoding 371 amino acids, showing 82.1–88.0% amino acid sequence identity with F3Hs from other plant species. The expression analysis showed that CpurF3H is strongly expressed in young petals, but rapidly decreases in petals following anthesis, suggesting that it functions during the early stages of flower development. Next, in vitro assays showed that recombinant CpurF3H proteins can convert naringenin to dihydrokaempferol. In addition, a complementation assay using the Arabidopsis thaliana f3h mutant confirmed the accumulation of anthocyanins in CpurF3H-transformed plant. Overall, these results strongly suggest that CpurF3H participates in flavonoid biosynthesis and flower coloration in C. purpurascens.