Beyond Purple Tomatoes: Combined Strategies Targeting Anthocyanins to Generate Crimson, Magenta, and Indigo Fruit

Abstract

The range of colours of many flowers and fruits is largely due to variations in the types of anthocyanins produced. The degree of hydroxylation on the B-ring affects the hue of these pigments, causing a shift from the orange end of the visible spectrum to the blue end. Besides colour, this modification can also affect other properties of anthocyanins, including the ability to protect the plant against different stresses or, when included in the human diet, to provide benefits for disease prevention. The level of hydroxylation of the B-ring is determined by the activity of two key hydroxylases, F3′H and F3′5′H, and by the substrate preference of DFR, an enzyme acting downstream in the biosynthetic pathway. We show that, in tomato, a strategy based on fruit-specific engineering of three regulatory genes (AmDel, AmRos1, AtMYB12) and a single biosynthetic gene (AmDFR), together with the availability of a specific mutation (f3′5′h), results in the generation of three different varieties producing high levels of anthocyanins with different levels of hydroxylation. These tomatoes show distinctive colours and mimic the classes of anthocyanins found in natural berries, thus providing unique near-isogenic material for different studies.

Designing a novel tomato ideotype for future cultivation in space manned missions

“…The higher accumulation of anthocyanins in hemizygous plants is in accordance with findings obtained for a fruit-specific anthocyanin-related, purple phenotype induced by Delila and Rosea1 (Del/Ros1) transgenes in hemyzigous MicroTom transformants (Butelli et al, 2008). The Del/Ros1 purple phenotype was, thereafter, usefully transferred into other tomato genetic backgrounds by crossing to obtain gene pyramiding (Butelli et al, 2021) and this approach might be taken in consideration for the MicroTom AN4-P 2 phenotype, as well.…”

A transient expression tool box for anthocyanin biosynthesis in Nicotiana benthamiana

“…In contrast, pelargonidin glucosides were produced less efficiently using the DFR gene from tomato (Figure S2). This result is consistent with previous observations that indicate that the tomato DFR has a substrate preference for dihydromyricetin (Bovy et al, 2002;Butelli et al, 2021), although it can also act on dihydroquercetin, as shown here.…”

Does Plant Breeding for Antioxidant-Rich Foods Have an Impact on Human Health?

“…Metabolic engineering allows to accumulate anthocyanins up to 2 mg/g FW in Del/Ros1 purple tomatoes, a level comparable to small berries [120] as well as to drive the selective accumulation of other flavonoids. Indeed, the Cathie Martin's research group developed several transgenic tomato lines enriched in polyphenols (Table 2); among them, the "Bronze" fruits showed the highest antioxidant capacity compared to both red and other polyphenol-enriched lines due to the simultaneous presence of different classes of polyphenols (flavonols, anthocyanins, and stilbenoids) [131,132]. SRG1 mutants 5.1× lycopene [126]; "Sun Black" (Aft/Aft atv/atv, peel) 0.2 mg/g DW total carotenoid content [127]; "Bronze" (E8:MYB12, E8:Del/Ros, 35S:StSy) ∼0.55 mg/g DW total carotenoid content [132] Golden potato (cv.…”

“…Desiree) 3600-fold increase in beta carotene to 4.7 mg/100 g DW [146]; 4 mg/100 g DW zeaxanthin in S. tuberosum 4n [147]. From traces to 0.33 mg/100 g beta-carotene [148] eggplant transgenic line EEF48:crtB 0.15 mg g −1 FW of β-carotene [149] Phenolic acids "Sun Black" (Aft/Aft atv/atv, peel) 0.6 mg/g DW of CGA, 8.6 mg/g DW total phenolic content [127]; "Yellow" E8:MYB12 15 mg g −1 DW CQAs equivalent to 22-fold higher levels, respectively [150] 3.35-fold increases on average) [151] Flavonoids (considered as total content) "Sun Black" (Aft/Aft atv/atv, peel) 0.8 mg/g DW rutin [132], "Yellow" (E8:MYB12) 72 mg g −1 DW total flavonols equivalent to 65-fold higher levels [150]; "Bronze" (E8:MYB12, E8:Del/Ros1, 35S:StSy), "Indigo" (E8:MYB12, E8:Del/Ros1) ∼15 and 20 mg/g DW total flavonols, respectively [132] Flavonols (4.50-fold increase on average) [151] Flavonoids (Anthocyanins) "Sun Black" (Aft/Aft atv/atv) 1.2 mg/g DW in fruit peel [127]; Del/Ros1 (E8:Del/Ros1) 14.7 mg g −1 ; "Indigo" (E8:MYB12, E8:Del/Ros1) ∼5-24 mg g −1 DW [131,132]; "Crimson" (E8:Del/Ros1, E8:AmDFR, f3 5 h) 5.3 ± 1.3 mg/g DW, "Magenta" (E8:Del/Ros1, E8:MYB12, E8:AmDFR, f3 5 h) 7.9 ± 2.3 mg/g DW total anthoyanins [131] From 0.4 in wt to 3 ug/100 mg Petunidin (7×); from 0.04 in wt to up to 0.3 ug/100 g Pelargonidin (7×) [152] 4.…”