Transcription factor PpNAC1 and DNA demethylase PpDML1 synergistically regulate peach fruit ripening.

Fruit ripening is accompanied by dramatic changes in color, texture, and flavor and is regulated by transcription factors (TFs) and epigenetic factors. However, the detailed regulatory mechanism remains unclear. Gene expression patterns suggest that PpNAC1 (NAM/ATAF1/2/CUC) TF plays a major role in peach (Prunus persica) fruit ripening. DNA affinity purification (DAP)-seq combined with transactivation tests demonstrated that PpNAC1 can directly activate the expression of multiple ripening-related genes, including ACC synthase1 (PpACS1) and ACC oxidase1 (PpACO1) involved in ethylene biosynthesis, pectinesterase1 (PpPME1), pectate lyase1 (PpPL1), and polygalacturonase1 (PpPG1) related to cell wall modification, and lipase1 (PpLIP1), fatty acid desaturase (PpFAD3-1), and alcohol acyltransferase1 (PpAAT1) involved in volatiles synthesis. Overexpression of PpNAC1 in the tomato (Solanum lycopersicum) nor (nonripening) mutant restored fruit ripening, and its transient overexpression in peach fruit induced target gene expression, supporting a positive role of PpNAC1 in fruit ripening. The enhanced transcript levels of PpNAC1 and its target genes were associated with decreases in their promoter mCG methylation during ripening. Declining DNA methylation was negatively associated with increased transcripts of DNA demethylase1 (PpDML1), whose promoter is recognized and activated by PpNAC1. We propose that decreased methylation of the promoter region of PpNAC1 leads to a subsequent decrease in DNA methylation levels and enhanced transcription of ripening-related genes. These results indicate that positive feedback between PpNAC1 and PpDML1 plays an important role in directly regulating expression of multiple genes required for peach ripening and quality formation.

Glycoside hydrolase PpGH28BG1 modulates benzaldehyde metabolism and enhances fruit aroma and immune responses in peach.

Benzaldehyde (BAld) is one of the most widely distributed volatiles that contributes to flavor and defense in plants. Plants regulate BAld levels through various pathways, including biosynthesis from trans-cinnamic acid (free BAld), release from hydrolysis of glycoside precursors (BAld-H) via multiple enzymatic action steps, and conversion into downstream chemicals. Here, we show that BAld-H content in peach (Prunus persica) fruit is up to 100-fold higher than that of free BAld. By integrating transcriptome, metabolomic and biochemical approaches, we identified glycoside hydrolase PpGH28BG1 as being involved in the production of BAld-H through the hydrolysis of glycoside precursors. Overexpressing and silencing of PpGH28BG1 significantly altered BAld-H content in peach fruit. Transgenic tomatoes heterologously expressing PpGH28BG1 exhibited a decrease in BAld-H content and an increase in SA accumulation, while maintaining fruit weight, pigmentation, and ethylene production. These transgenic tomato fruits displayed enhanced immunity against Botrytis cinerea compared to wild type (WT). Induced expression of PpGH28BG1 and increased SA content were also observed in peach fruit when exposed to Monilinia fructicola infection. Additionally, elevated expression of PpGH28BG1 promoted fruit softening in transgenic tomatoes, resulting in a significantly increased emission of BAld compared to WT. Most untrained taste panelists preferred the transgenic tomatoes over WT fruit. Our study suggests that it is feasible to enhance aroma and immunity in fruit through metabolic engineering of PpGH28BG1 without causing visible changes in the fruit ripening process.

Multi-omics analysis unravels chemical roadmap and genetic basis for peach fruit aroma improvement.

Selection of fruits with enhanced health benefits and superior flavor is an important aspect of peach breeding. Understanding the genetic interplay between appearance and flavor chemicals remains a major challenge. We identify the most important volatiles contributing to consumer preferences for peach, thus establishing priorities for improving flavor quality. We quantify volatiles of a peach population consisting of 184 accessions and demonstrate major reductions in the important flavor volatiles linalool and Z-3-hexenyl acetate in red-fleshed accessions. We identify 474 functional gene regulatory networks (GRNs), among which GRN05 plays a crucial role in controlling both red flesh and volatile content through the NAM/ATAF1/2/CUC (NAC) transcription factor PpBL. Overexpressing PpBL results in reduced expression of PpNAC1, a positive regulator for Z-3-hexenyl acetate and linalool synthesis. Additionally, we identify haplotypes for three tandem PpAATs that are significantly correlated with reduced gene expression and ester content. We develop genetic resources for improvement of fruit quality.