Investigation of the biosynthesis of 3‐deoxyanthocyanins in Sinningia cardinalis

3‐Deoxyanthocyanins provide bright orange‐red colours to flowers of some members of the Gesneriaceae, including sinningia ( Sinningia cardinalis ). We examined 3‐deoxyanthocyanin biosynthesis in sinningia, in particular, the expression of key flavonoid biosynthetic genes and the activities of the encoded proteins. Two abundant 3‐deoxyanthocyanins, luteolinidin 5‐ O ‐glucoside and apigeninidin 5‐ O ‐glucoside, three flavone glycosides, luteolin 7‐ O ‐glucoside, luteolin 7‐ O ‐glucuronide and apigenin 7‐ O ‐glucuronide, and the cinnamic acid verbascoside were identified in sinningia petal tissue. Small amounts of a 3‐hydroxyanthocyanin were also detected in a limited region of the petal. cDNA clones for three flavonoid enzymes, flavanone 3‐hydroxylase (F3H), dihydroflavonol 4‐reductase/flavanone 4‐reductase (DFR/FNR) and anthocyanidin synthase (ANS), were isolated from a sinningia cDNA library made from petal RNA and used to measure transcript abundance during petal development. Only very low levels of F3H transcript were detected, while DFR/FNR transcript was highly abundant. ANS transcript levels were intermediate between these two. The F3H cDNA was shown to encode a functional F3H protein by complementation of the phenotype of an Antirrhinum majus F3H mutant. The recombinant DFR/FNR had activity against both flavanone and dihydroflavonol substrates to a comparable extent. The results suggest a mechanism of 3‐deoxyflavonoid biosynthesis in sinningia similar to that reported for Zea mays , in which lack of F3H activity allows action of the DFR/FNR on flavanone substrates and production of flavan‐4‐ols. These are then likely converted to 3‐deoxyanthocyanins through the action of the ANS and subsequent glucosylation.