Red to Brown: An Elevated Anthocyanic Response in Apple Drives Ethylene to Advance Maturity and Fruit Flesh Browning

Abstract

The elevation of anthocyanin contents in fruits and vegetables is a breeding target for many crops. In some fruit, such as tomato, higher anthocyanin concentrations enhance storage and shelf life. In contrast, highly anthocyanic red-fleshed apples (Malus x domestica) have an increased incidence of internal browning flesh disorder (IBFD). To determine the mechanisms underlying this, ‘Royal Gala’ cultivar apples over-expressing the anthocyanin-related transcription factor (TF) MYB10 (35S:MYB10), which produces fruit with highly pigmented flesh, were compared with standard ‘Royal Gala’ Wild Type (WT) grown under the same conditions. We saw no incidence of IBFD in WT ‘Royal Gala’ but the over-expression of MYB10 in the same genetic background resulted in a high rate of IBDF. We assessed concentrations of potential substrates for IBDF and a comparison of metabolites in these apples showed that anthocyanins, chlorogenic acid, pro-cyanidins, flavon-3-ols, and quercetin were all higher in the MYB10 lines. For the flavol-3-ols sub-group, epicatechin rather than catechin was elevated in MYB10 lines compared with the control fruit. Internal ethylene concentrations were measured throughout fruit development and were significantly higher in 35S:MYB10 lines, and ethylene was detected at an earlier developmental stage pre-harvest. Expression analysis of key genes associated with ethylene biosynthesis (aminocyclopropane-1-carboxylic acid synthase and oxidase; ACS and ACO) and polyphenol oxidase (PPO) showed the potential for increased ethylene production and the mechanism for enhanced PPO-mediated browning. The expression of a transcription factor of the ethylene response factor (ERF) class, ERF106, was elevated in red flesh. Analysis of transcriptional activation by MYB10 showed that this transcription factor could activate the expression of apple ACS, ACO, and ERF106 genes. Our data show a link between the elevation of anthocyanin-related transcription factors and an undesirable fruit disorder. The accelerated advancement of maturity via premature ethylene induction has implications for the breeding and storage of these more highly pigmented plant products.

LjaFGD: Lonicera japonica functional genomics database

“…We determined that many TFs, such as MYB, bHLH, WRKY, and ERF, might regulate key genes in the biosynthesis pathways of chlorogenic acid and luteolin in L. japonica. Previous research suggests that TFs in the MYB and bHLH families might participate in the biosynthesis of chlorogenic acid (Espley et al, 2019;Liu et al, 2019;Qi et al, 2019;Wang et al, 2019). Therefore, we used the motif analysis tool to analyze possible regulatory factors upstream of key enzyme genes and observed that MYB, WRKY, and ERF motifs were significantly enriched.…”

A Critical Review: Recent Advancements in the Use of CRISPR/Cas9 Technology to Enhance Crops and Alleviate Global Food Crises

“…Likewise, the GBSS gene that is responsible for the synthesis of starch was mutated using Cas9 in potatoes. An increase in amylopectin levels was observed in mutant lines [126]. The SnLazy1 locus, known as the Lazy1 ortholog in tomatoes, was manipulated by CRISPR/Cas9 with effective heritability of the snlazy-1 allele, and mutants showed a downward pattern of stem development [127].…”

“…A ground cherry of a wild variety of tomatoes yielded higher and larger fruits [129]. Additionally, in apples, a mutation of PPO was reflected as transgene-free using Cas9, which is appropriate for human consumption [126]. The production of seedless fruit is always the primary goal of any breeding plan.…”

A Revolution toward Gene-Editing Technology and Its Application to Crop Improvement

“…Similarly, the starch synthase GBSS gene has been mutated in potato via CRISPR/Cas9. The mutated lines showed reduced amylose levels and an increased concentration of the amylose/amylopectin ratio [122]. Furthermore, the SnLazy1 locus, which is the tomato ortholog of Lazy1, was edited by CRISPR/Cas9 in Solanum nigra, with successful inheritance of the removal of two separate snlazy1-cr alleles and the production of plants with stem development in a relatively downward direction [123].…”

“…Another group investigated a wild-type species of tomato called groundcherry (Physalis pruinosa) that produces a high yield of large fruits [39]. In addition, the PPO mutation in apples can be considered transgene free using CRISPR/Cas9 and could easily be applicable worldwide [122]. CRISPR/Cas9 is committed to the development of seedless fruits through the modification/mutation of genes responsible for seed formation.…”