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Abstract In this paper, we explored the influence of lipase on cuticle wax and anthocyanin metabolism in blueberries. The results demonstrated a progressive alteration in wax crystal structure, transitions from rod‐like to short rod‐like and flaky, and even disappearance of the cuticle wax. Gas chromatography–mass spectrometry analysis revealed a reduction in esters, aldehydes, long‐chain alkanes, and total wax content following lipase treatment of blueberry fruits. Additionally, ultra high performance liquid chromatography–mass spectrometry analysis showed significant decreases in peonidin, petunidin, cyanidin, and delphinidin, accompanied by an increase in malvidin content due to lipase treatment. Subsequent investigations uncovered that lipase treatment inhibited the activities of key anthocyanin synthetic enzymes, namely phenylalanine ammonia‐lyase (PAL), chalcone isomerase (CHI), and dihydroflavonol reductase (DFR). Moreover, lipase treatment downregulated the expression of synthesis‐related genes, including PAL, 4‐coumarate: CoA ligase, chalcone synthase (CHS), CHI, and DFR. In summary, our findings indicate that lipase disrupts the integrity of cuticle wax structure, inhibits anthocyanin synthesis, and reduces anthocyanin content. Accordingly, our studies suggest that inhibiting lipase could offer a novel approach to preserving the freshness and anthocyanin concentration of blueberries.

This study aimed to unraveling the color evolution and metabolic pathways of pelargonidin-3-O-glucoside (P3G) during lactic acid fermentation of the strawberry juice color simulation system. The results revealed that fermentation with both Lactobacillus plantarum and Lactobacillus acidophilus caused a decline in pH of the strawberry juice color simulation system and significantly accelerated the decrease in P3G content. The CIELAB space model pointed out that parameters a ⁎ and b ⁎ of the group treated with both lactic acid bacteria and P3G initially increased to a peak level and then gradually decreased, shifting the overall color towards orange and then gradually fading. Furthermore, untargeted metabolomics results revealed that P3G was progressively degraded and converted to pyruvate, methylparaben, 3,4-dihydroxybenzoic acid, p-anisic acid, and terephthalic acid, affecting the metabolic pathways of glycolysis, d-amino acids, benzoate degradation, aromatic compounds degradation, and aminobenzoate degradation in lactic acid bacteria., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier Ltd.)

Anthocyanins are susceptible to degradation owing to environmental factors. Combining them with proteins can improve their stability; however, the interaction mechanism is difficult to elucidate. This study used multispectral and molecular dynamics simulations and molecular docking methods to investigate the interaction mechanism between peanut protein isolate (PPI) and cyanidin-3-O-glucoside (C3G). The UV absorption peak and PPI turbidity increased, while the fluorescence intensity decreased with greater C3G content. Protein secondary structure changes suggested that PPI and C3G coexisted in spontaneous covalent and non-covalent interactions via static quenching. The complex structures were stable over time and C3G stably bound to the peanut globulin Ara h 3 cavity through hydrogen bonding and hydrophobic interactions. Furthermore, PPI enhanced the C3G antioxidant activity and bioaccessibility by increasing its retention rate during in-vitro simulated digestion. This study elucidates the binding mechanism of PPI and C3G and provides insight into applications of the complex in food development., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Background: Soybean reddening during storage and transportation has caused great concern due to the serious economic loss. However, the mechanism of reddening has not been clearly elucidated. In this study, metabolomics was employed to investigate the reasons for soybean reddening during storage., Results: The results of multivariate statistical analysis showed that the metabolite level of red soybean was significantly different from that of normal soybean. The differentially expressed metabolites were mainly enriched by biosynthesis of secondary metabolites and amino acid metabolism. Metabolism analysis showed that the biosynthesis of cyanidin and betalains was enhanced in reddening soybean. In addition, it was found that phenolic and flavonoid compounds decreased, while quinones, furans and 5-hydroxymethylfurfural increased in reddening soybeans compared to normal soybeans., Conclusion: The upregulation of cyanidin and betalains was the main reason for soybean reddening. Besides, the oxidation of phenols and flavonoids, as well as Maillard reaction, also contributed to the color change. © 2024 Society of Chemical Industry., (© 2024 Society of Chemical Industry.)

The metabolic processes of fruits induce water vapor condensation in packaging, creating a high-humidity microenvironment that exacerbates the proliferation of microorganisms and causes food spoilage. Therefore, a Fe/Zr bimetallic organic framework (Fe-MOF-801) packaging (i.e., FeMGCF) with high binding stability to transpiration water was prepared based on transition metal Fe doping strategy, and the mechanism of its enhanced binding stability towards transpiration water was investigated. The results indicate that Fe enhances the interaction between MOFs and H 2 O by promoting the charge depletion at the O sites coordinated with Zr, thereby increasing the aggregation tendency of H 2 O in the T 1 and T 2 cavities (characterized by strong hydrogen bonding) of Fe-MOF-801, which increased the binding energy of MOFs to H 2 O by 29.6 %. Accordingly, the application of FeMGCF packaging extended the shelf life of tomatoes by 6 days and delayed the degradation of lycopene and anthocyanins during postharvest tomato preservation., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier Ltd.)

The deposition of anthocyanin plays a crucial role in fruit pigmentation and serves as the primary determinant of pear quality. Various factors influence the synthesis of anthocyanins, with salicylic acid playing a significant role among them. However, the mechanism by which salicylic acid affects anthocyanin synthesis remains unclear. Our study identifies the transcription factor PyWRKY40 as a pivotal regulator of SA-mediated anthocyanin synthesis in the nucleus. The negative regulatory function of this factor lies in its ability to suppress anthocyanin synthesis, thereby exerting an influence on fruit coloration. We have confirmed the direct binding of PyWRKY40 to the PyDFR promoter through Y1H and EMSA experiments. The findings elucidate a signaling regulatory module, PyWRKY40-PyDFR, which is responsive to SA and enhances fruit pigmentation by modulating anthocyanin metabolism. This insight offers a viable approach to enhancing fruit coloration and improving the overall quality of pear fruits., Competing Interests: Declaration of Competing Interest The authors declare no conflict of interest., (Copyright © 2024. Published by Elsevier B.V.)

Introduction: The scarcity of naturally available sources for blue colorants has driven reliance on synthetic alternatives. Nevertheless, growing health concerns have prompted the development of naturally derived blue colorants, which remains challenging with limited success thus far. Anthocyanins (ACNs) are known for providing blue colors in plants, and metal complexation with acylated ACNs remains the primary strategy to generate stable blue hues. However, this approach can be costly and raise concerns regarding potential metal consumption risks., Objectives: Our study aims to introduce a metal-free approach to achieve blue coloration in commonly distributed non-acylated 3-glucoside ACNs by exploring their interactions with proteins and unveiling the underlying mechanisms., Methods: Using human serum albumin (HSA) as a model protein, we investigated the structural influences of ACNs on their blue color generation using visible absorption spectroscopy, fluorescence quenching, and molecular simulations. Additionally, we examined the bluing effects of six proteins derived from milk and egg and identified the remarkable roles of bovine serum albumin (BSA) and lysozyme (LYS)., Results: Our findings highlighted the importance of two or more hydroxyl or methoxyl substituents in the B-ring of ACNs for generating blue colors. Cyanidin-, delphinidin- and petunidin-3-glucoside, featuring two neighboring hydroxyl groups in the B-ring, exhibited blue coloration when interacting with HSA or LYS, driven primarily by favorable enthalpy changes. In contrast, malvidin-3-glucoside, with two methoxyl substituents, achieved blue coloration through interactions with HSA or BSA, where entropy change played significant roles., Conclusion: Our work, for the first time, demonstrates the remarkable capability of widely distributed 3-glucoside ACNs to generate diverse blue shades through interactions with certain proteins. This offers a promising and straightforward strategy for the production of ACN-based blue colorants, stimulating further research in this field., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier B.V.)

Background: Drying is a critical post-harvest process for medicinal plants, which are typically high in moisture and microorganisms. To prevent spoilage and quality loss, it is essential to dry these plants promptly. The drying method significantly impacts the levels of secondary metabolites and the organoleptic characteristics of medicinal plants. This study aimed to investigate the effects of various drying methods on the total phenolics, flavonoids, anthocyanins, antioxidant activity, and phenolic acids in caper (Capparis spinosa L.) fruits. The experiment was performed using a completely randomized design with three replications and included 11 treatments: shade drying, sun drying, oven drying (at 50 °C, 60 °C, and 70 °C), microwave drying (at 300 W, 600 W, and 900 W), freeze-drying, salt-drying, and a fresh plant sample as a control., Results: Among the drying methods tested, microwave drying consistently produced the highest levels of flavonoids, anthocyanins, and antioxidant activity, regardless of wattage. Specifically, the highest total phenol content was observed in samples dried at 900 W microwave, 600 W microwave, and 70 °C in the oven (5.3, 5.37, and 5.31 mg GAE/g DW, respectively). Drying at 600 W microwave yielded the highest levels of caffeic, cinnamic, ferulic, vanillic, and protocatechuic acids (13.03, 3.85, 4.28, 9.73, and 5.6 µg/g, respectively) while drying at 900 W microwave also resulted in elevated levels of caffeic, ferulic, protocatechuic, and p-coumaric acids. The 70 °C oven drying method also showed high levels of caffeic, cinnamic, ferulic, vanillic, and protocatechuic acids. Freeze-drying achieved the highest levels of rosmarinic, gallic, and m-coumaric acids (320.17, 175.3, and 12.99 µg/g, respectively), while shade drying produced high levels of p-hydroxybenzoic, cinnamic, ferulic, m-coumaric, protocatechuic, and p-coumaric acids., Conclusions: Overall, microwave drying (especially at 600 W), oven drying at 70 °C, and freeze-drying emerged as effective alternatives to traditional drying methods. These methods not only preserved the color, texture, and taste of the fruits but also enhanced their bioactive compound levels., Competing Interests: Declarations. Ethics approval and consent to participate: The plant materials utilized in this study were C. spinosa fruits collected from their natural habitats in Khorramabad, Iran. Plant material was sampled on private land with the owner’s permission. We adhered to all relevant institutional, national, and international guidelines and legislation during the collection process. All procedures were carried out accordnig to the guidelines. Consent for publication: Not applicable. Guideline statement: The authors confirm that all methods were carried out following relevant guidelines and regulations. Clinical trial number: Not applicable. Competing interests: The authors declare no competing interests., (© 2025. The Author(s).)

Anthocyanins not only serve as critical pigments determining floral hues but also play essential roles in attracting insects for pollination, feeding animals and mitigating abiotic stress. However, the molecular mechanisms underlying the regulation of flower color in sesame has not yet been reported. In this study, an F 2 population was constructed by crossing 'Ganzhi 9' (purple-flowered) with 'BS377' (white-flowered). Genetic analysis revealed that purple flower is controlled by a single locus named as SiFC (Sesamum indicum flower color). Using the BSA-seq approach, SiFC was preliminarily identified on chromosome 6, which was further mapped to a 473 kb interval using Kompetitive Allele Specific PCR (KASP) marker analysis. Moreover, functional annotation, expression profiling, and sequence analyses confirmed that the SibHLHA (Sesame10992) was the most likely candidate gene for SiFC. In addition, SibHLHA, highly homologous to AtTT8 (a key regulator in the anthocyanin synthesis pathway), was found to interact with WER-like or TTG1 proteins, enhancing anthocyanin accumulation in tobacco leaves. Furthermore, an SNP in the second exon of Sibhlha (BS377 variant) was found to alter the encoding amino acids, which affected Sibhlha binding to MYB protein and showed low anthocyanin in tobacco leaves compared with SibHLHA binding with WER-like or TTG1 proteins. These findings not only deepen our understanding of the molecular mechanisms controlling sesame corolla color, but also provide valuable insights for developing ornamental and consumable sesame varieties., Competing Interests: Declarations. Conflict of interest: All these authors declare that they have no conflicts of interest., (© 2025. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Betalains, a distinctive group of nitrogen-containing pigments exclusive to the Caryophyllales order, possess diverse biological activities such as antioxidant, anti-inflammatory, and antimicrobial properties, making them highly valuable for applications in food, nutraceutical, and pharmaceutical industries. This Review provides a comprehensive analysis of betalain biosynthesis, structural diversity, and ecological significance, highlighting their roles in enhancing stress resilience, adaptation mechanisms, and plant evolutionary strategies. The evolutionary development of betalains is explored, revealing their emergence through gene duplication events and providing insights into their mutual exclusivity with anthocyanins. This study utilizes comparative genetics and advanced molecular tools to uncover the intricate regulatory networks involving transcription factors such as MYB, bHLH, WRKY, and SPL, which govern betalain biosynthesis. Furthermore, the Review discusses innovative transgenic studies that introduce betalains into non-native species, demonstrating their potential to enhance stress tolerance and boost agricultural productivity. While significant progress has been made in understanding betalain biosynthesis pathways, the evolutionary relationships with anthocyanins and the specific ecological functions of betalains in plants remain areas of ongoing exploration. Future research directions include integrating chemotaxonomic studies, molecular phylogenetics, and multiomics approaches to unravel the full spectrum of betalain functions and regulatory mechanisms. Such studies are essential to deepening our understanding of these vibrant pigments and their evolutionary implications, offering new opportunities for biotechnological innovations and sustainable agricultural practices. This Review stands out by combining genetic, ecological, and evolutionary perspectives, providing novel insights into the multifunctionality of betalains and their potential to drive future advancements in plant science and biotechnology.

Background: Red Actinidia arguta has recently become highly popular because of its red appearance resulting from anthocyanin accumulation, and has gradually become an important breeding direction. However, regulators involved in anthocyanin biosynthesis have not been fully characterized in A. arguta., Results: Here, we demonstrated that a key R2R3-MYB transcription factor, AaMYB61-like, plays a crucial role in A. arguta anthocyanin biosynthesis. The RT-qPCR results revealed that transient overexpression of AaMYB61-like in A. arguta fruit at 90-100 DAFB significantly promoted anthocyanin biosynthesis, as did the gene expression levels of AaCHS, AaCHI, AaF3H, AaLDOX, and AaF3GT, whereas the result of VIGS revealed the opposite results in A. arguta fruit at 105-115 DAFB. A transcriptional activation assay indicated that AaMYB61-like exhibited transcriptional activation activity. Y1H and LUC assays revealed that AaMYB61-like activates the promoters of AaCHS, AaLDOX, and AaF3GT. In addition, AabHLH137 was found to be related to fruit color from the transcriptome data. We demonstrated that AaMYB61-like promotes anthocyanin biosynthesis by interacting with AabHLH137 via Y2H, BiFC, and Agrobacterium-mediated co-transformation., Conclusions: Our study not only reveals the functions of AaMYB61-like and AabHLH137 in anthocyanin regulation, but also broadly enriches color regulation theory, establishing a foundation for clarifying the molecular mechanism of fruit coloration in kiwifruit., Competing Interests: Declarations. Ethics approval and consent to participate: This study did not include human or animal subjects. Consent for publication: Not applicable. Competing interests: The authors declare no competing interests., (© 2025. The Author(s).)

Gloriosa L. possesses exceptional ornamental value, with its floral hues exhibiting a wide range of variations. In this study, we employed sophisticated colorimetry, Ultra Performance Liquid Chromatography-Tandem Mass Spectrometry (UPLC-MS/MS), and transcriptome sequencing to investigate the phenotypic expression of tepal colors, the composition of carotenoids and anthocyanins, and the differential gene expression in four Gloriosa varieties during their full bloom phase. Our findings revealed that the redness of the tepals, indicated by higher a * values, increased with the intensity of the red hue, while lighter colors corresponded to higher L * values. Metabolomic analysis identified 50 carotenoids and 60 anthocyanins. It was observed that carotenoids primarily influence the yellow and orange color of Gloriosa tepals, with β-carotene, lutein, and zeaxanthin being the predominant carotenoids. Anthocyanins serve as the principal coloring agents in the orange, red and purple tepals of Gloriosa. High levels and proportions of cyanidins and pelargonidins are key contributors to the formation of red and purple tepals, while high levels and proportions of peonidins also play a significant role in purple coloration. In contrast, the presence of high levels and proportions of pelargonidins alone is a crucial factor in the formation of orange tepals. Transcriptomic data unearthed 57 and 92 candidate differentially expressed genes (DEGs) belong to carotenoid and anthocyanin biosynthesis pathway, respectively, with PSY, PDS, DFR, and ANS genes considered as critical genes for the differential accumulation of pigments of Gloriosa tepals. Weighted gene co-expression network analysis (WGCNA) revealed significant co-expression patterns between 217 transcription regulatory factors (TFs) and 8 carotenoid biosynthesis genes, and between 194 TFs and 41 anthocyanin biosynthesis genes. qRT-PCR verified the expression patterns of four carotenoid biosynthesis-related genes, eight anthocyanin biosynthesis-related genes, and three transcription regulatory factors. It was found that Cluster-121969.6 (MYB) gene is specifically expressed in the tepals of the four varieties (compared to stems and leaves) and shows a high consistency with the trend of anthocyanin content changes. This research provides new insights into the mechanisms underlying the formation of diverse floral colors in Gloriosa tepals., Competing Interests: Declarations. Ethics approval and consent to participate: Not applicable. Consent for publication: Not applicable. Competing interests: The authors declare no competing interests., (© 2025. The Author(s).)

Background: Anthocyanin is an important class of water-soluble pigments that are widely distributed in various tissues of plants, and it not only facilitates diverse color changes but also plays important roles in various biological processes. Maize silk, serving as an important reproductive organ and displaying a diverse range of colors, plays an indispensable role in biotic resistance through its possession of anthocyanin. However, the copy numbers, characteristics, and expression patterns of genes involved in maize anthocyanin biosynthesis are not fully understood. In this study, gene numbers, distribution, structure, cis-elements of the anthocyanin synthetic gene family were identified, and then the potential transcriptional factors were predicted by two analyzed methods. Finally, genes involved in maize silk pigment were screened by un-targeted metabolism analysis., Results: Ten gene families involved in the maize anthocyanin biosynthesis pathway were identified, and 142 synthetic genes were obtained. These anthocyanin biosynthetic genes have high copy numbers and are normally clustered on chromosomes. The promoters of these synthetic genes contain various cis-elements and the gene expression patterns and transcriptional regulatory networks were analyzed. These genes are distributed on different chromosomes and gene expression patterns vary across different tissues in maize. Specifically, these genes often exhibit higher expression in the stem, leaves, and seeds. Ten highly expressed genes in silks were identified. Based on un-targeted metabolites detection in the silks of four maize representative inbred lines with different colors, two main differential anthocyanin components were identified. Furthermore, the gene expression patterns of the ten highly expressed genes and their potential interacting transcriptional factors were analyzed across the four inbred lines., Conclusions: The results in this study show a through picture of maize anthocyanin synthetic genes, and the structure and function of genes related to anthocyanin biosynthesis in maize could be further investigated., Competing Interests: Declarations. Ethics approval and consent to participate: Not applicable. Consent for publication: Not applicable. Competing interests: The authors declare no competing interests. Clinical trial number: Not applicable., (© 2025. The Author(s).)

Background: Synchronous degradation between anthocyanin and vitamin C was found in fruit and vegetable juice matrices. To investigate whether the C-ring of anthocyanin is the key site of this interaction, cyanidin with four different C-ring modifications (3-glucosylation, 3,5-diglucosylation, 6″-malonylation, pyranylation) was added to vitamin C-containing apple juice, and the changes of anthocyanin retention, vitamin C retention, color, antioxidative activity and differential metabolites were analyzed., Results: The anthocyanin retention was in the order of pyranylation >6″-malonylation >3,5-diglucosylation >3-glucosylation. The vitamin C retention was in the order of 6″-malonylation > pyranylation >3,5-diglucosylation >3-glucosylation. The order of color stability was the same as that of anthocyanin retention, and the order of antioxidative activity was opposite to that of vitamin C retention. The results showed that modification at the C-ring limited the activity of anthocyanin, and suggested that the C-ring was one of the key sites for anthocyanin and vitamin C interaction. The shared differential metabolite of all apple juice matrices added with different anthocyanins was trans-hinokiresinol, which was likely generated from anthocyanin skeleton reacted with certain compounds in apple juice., Conclusion: This study showed that modification of the anthocyanin C-ring could affect the anthocyanin and vitamin C interaction to some extent, which provided valuable insights for the application of anthocyanin C-ring modification in shelf-life quality control of typical fruit and vegetable beverages with the coexistence of anthocyanin and vitamin C. © 2024 Society of Chemical Industry., (© 2024 Society of Chemical Industry.)

Background: Anthocyanins are susceptible to degradation due to external factors. Despite the potential for improved anthocyanin stability with whey protein isolate (WPI), the specific effects of individual components within WPI on the stability of anthocyanins have yet to be studied extensively. This study investigated the interaction of WPI, β-lactoglobulin (β-Lg), bovine serum albumin (BSA), and lactoferrin (LF) with cyanidin-3-O-glucoside (C3G), and also considered their effects on stability., Results: Fluorescence analysis revealed static quenching effects between C3G and WPI, β-Lg, BSA, and LF. The binding constants were 1.923 × 10 3 L · mol⁻¹ for WPI, 24.55 × 10 3 L · mol⁻¹ for β-Lg, 57.25 × 10 3 L · mol⁻¹ for BSA, and 1.280 × 10 3 L · mol⁻¹ for LF. Hydrogen bonds, van der Waals forces, and electrostatic attraction were the predominant forces in the interactions between C3G and WPI and between C3G and BSA. Hydrophobic interaction was the main binding force in the interaction between C3G and β-Lg and between C3G and LF. The binding of C3G with WPI, β-Lg, BSA, and LF was driven by different thermodynamic parameters. Enthalpy changes (∆H) were -38.76 kJ · mol⁻¹ for WPI, -17.59 kJ · mol⁻¹ for β-Lg, -16.09 kJ · mol⁻¹ for BSA, and 39.50 kJ · mol⁻¹ for LF. Entropy changes (∆S) were -67.21 J · mol⁻¹·K⁻¹ for WPI, 3.72 J · mol⁻¹·K⁻¹ for β-Lg, 37.09 J · mol⁻¹·K⁻¹ for BSA, and 192.04 J · mol⁻¹·K⁻¹ for LF. The addition of C3G influenced the secondary structure of the proteins. The decrease in the α-helix content suggested a disruption and loosening of the hydrogen bond network structure. The presence of proteins enhanced the light stability and thermal stability (stability in the presence of light and heat) of C3G. In vitro simulated digestion experiments demonstrated that the addition of proteins led to a delayed degradation of C3G and to improved antioxidant capacity., Conclusion: The presence of WPI and its components enhanced the thermal stability, light stability, and oxidation stability of C3G. Preheated proteins exhibited a more pronounced effect than unheated proteins. These findings highlight the potential of preheating protein at appropriate temperatures to preserve C3G stability and bioactivity during food processing. © 2024 Society of Chemical Industry., (© 2024 Society of Chemical Industry.)

Background: Genetic breeding is essential to develop grapevine genotypes adapted to warm climates and resistant to pathogens. Traditionally cultivated Vitis vinifera is susceptible to biotic and abiotic stresses. Winemakers and consumers, however, perceive wines from non-vinifera or hybrid cultivars as inferior. In this study, sensory analyses and comprehensive metabolite profiling by targeted and untargeted approaches were used to investigate the oenological potential of wines from grapes of genotypes developed throughout four breeding cycles to improve climate adaptation, sugar contents and berry color., Results: Novel genotypes had higher yields and the wines exhibited increased contents of polyphenols, including anthocyanins. Volatile monoterpenes in the wines decreased throughout breeding cycles in the absence of selective pressure. Polyphenol contents were higher in intermediate wines, with hydroxytyrosol contents reaching up to three times reported values. Mouthfeel attributes astringency, leafy taste, flavor and body, and persistency showed significant correlation with untargeted features. Supervised model-based analyses of the metabolome effectively discriminate wines from distinct genetic origins., Conclusion: Taken together, the results demonstrate the potential of novel grapevine genotypes to a more sustainable viticulture and quality wine production in warm climates. Comprehensive metabolite profiling of the wines reveals that genotype clustering is dependent on the chemical class and that traits not submitted to selective pressure are also altered by breeding. Supervised multivariate models were effective to predict the genetic origin of the wines based on the metabolic profile, indicating the potential of the technique to identify biomarkers for wines from sustainable genotypes. © 2024 Society of Chemical Industry., (© 2024 Society of Chemical Industry.)

Background: Wheat seeds display different colors due to the types and contents of anthocyanins, which is closely related to anthocyanin metabolism. In this study, a transcriptomic and metabolomic analysis between white and purple color wheat pericarp aimed to explore some key genes and metabolites involved in anthocyanin metabolism., Results: Two wheat cultivars, a white seed cultivar Shiluan02-1 and purple seed cultivar Hengzi151 were used to identify the variations in differentially expressed genes (DEGs) and differentially accumulated flavonoids (DAFs). Based on metabolomic data, 314 metabolites and 191 DAFs were identified. Chalcone, flavonol, pro-anthocyanidin and anthocyanidin were the most differentially accumulated flavonoid compounds in Hengzi151. 2610 up-regulated and 2668 down-regulated DEGs were identified according to transcriptomic data. The results showed that some structural genes in anthocyanin synthesis pathway were prominently activated in Hengzi151, such as PAL, CAD, CHS and so on. Transcription factors (TFs) of MYB, bHLH, WD40 and some other TFs probably involved in regulating anthocyanin biosynthesis were identified. Some genes from hormone synthetic and signaling pathways that may participate in regulating anthocyanin biosynthesis also have been identified., Conclusions: Our results provide valuable information on the candidate genes and metabolites involved in the anthocyanin metabolism in wheat pericarp., Competing Interests: Declarations. Ethics approval and consent to participate: All experimental research and field studies on plants in our study complies with Chinese institutional, national, and international guidelines and legislation. Consent for publication: Not applicable. Competing interests: The authors declare no competing interests., (© 2025. The Author(s).)

Stropharia rugosoannulata , or wine-cap Stropharia, is a well-known edible mushroom cultivated globally. The pileipellis color is a crucial quality attribute of S. rugosoannulata , exhibiting significant variation throughout its developmental stages. However, the pigment types and regulatory mechanisms behind color variation remain unclear. The metabolome analysis found that the anthocyanin biosynthesis pathway was significantly enriched and anthocyanins accumulated steadily in fruiting bodies during three developmental stages. The pileipellis pigment was extracted, and HPLC-MS confirmed the presence of anthocyanins. Notably, significant differences in anthocyanin content were observed among the various colored varieties. Thus, anthocyanins contribute to the pileipellis color of S. rugosoannulata . Through further investigation, this study elucidated, for the first time, the relationship between the "SrNFYA- SrDRF2 " regulatory module and anthocyanin accumulation. Combined multiomics assays and HPLC analysis revealed that auxin functions as a signaling molecule that regulates the accumulation of anthocyanins in the pileipellis. Subsequently, the hub gene of anthranilate synthase for auxin synthesis was identified as SrTRP1, and the transcription factor SrMYB1 was verified as a regulator of SrTRP1 , influencing auxin accumulation. These findings provide a valuable resource for the targeted enhancement of the quality of S. rugosoannulata .

Background: The quality of soybeans is reflected in the seed coat color, which indicates soybean quality and commercial value. Researchers have identified genes related to seed coat color in various plants. However, research on the regulation of genes related to seed coat color in soybeans is rare., Methods: In this study, four lines of seed coats with different colors (medium yellow 14, black, green, and brown) were selected from the F 2:5 population, with Beinong 108 as the female parent and green bean as the male parent, and the dynamic changes in the anthocyanins in the seed coat were stained with 4-dimethylaminocinnamaldehyde (DMACA) during the grain maturation process (20 days from grain drum to seed harvest). Through RNA-seq of soybean lines with four different colored seed coats at 30 and 50 days after seeding, we can further understand the key pathways and gene regulation modules between soybean seed coats of different colors., Results: DMACA revealed that black seed coat soybeans produce anthocyanins first and have the deepest staining. Clustering and principal component analysis (PCA) of the RNA-seq data divided the eight samples into two groups, resulting in 16,456 DEGs, including 5359 TFs. GO and KEGG enrichment analyses revealed that the flavonoid biosynthesis, starch and sucrose metabolism, carotenoid biosynthesis, and circadian rhythm pathways were significantly enriched. We also conducted statistical and expression pattern analyses on the differentially expressed transcription factors. Based on weighted gene coexpression network analysis (WGCNA), we identified seven specific modules that were significantly related to the four soybean lines with different seed coat colors. The connectivity and functional annotation of genes within the modules were calculated, and 21 candidate genes related to soybean seed coat color were identified, including six transcription factor (TF) genes and three flavonoid pathway genes., Conclusions: These findings provide a theoretical basis for an in-depth understanding of the molecular mechanisms underlying differences in soybean seed coat color and provide new genetic resources.

R2R3-MYB transcription factors function as the master regulators of the phenylpropanoid pathway in which both lignin and anthocyanin are produced. In poplar, R2R3-MYB transcription factor PdMYB118 positively regulates anthocyanin production to change leaf color. However, the molecular mechanism by which it controls different branches of the phenylpropanoid pathway still remains poorly understood. Here, we reported that in addition to anthocyanin synthesis, lignin deposition and xylem differentiation were regulated by PdMYB118 through inhibiting PagKNAT2/6b gene expression. The transgenic poplar plants overexpressing PdMYB118 accumulated more xylem, lignin and anthocyanin. Transcriptome and reverse transcription quantitative PCR analyses revealed that the expression of PagKNAT2/6b gene which inhibited lignin deposition and xylem differentiation was significantly down-regulated in transgenic poplar plants. Subsequent dual-luciferase reporter and yeast-one-hybrid assays demonstrated that PdMYB118 directly inhibited the transcription of PagKNAT2/6b by binding to the AC elements in its promoter region. Further experiments with transgenic poplar plants overexpressing PagKNAT2/6b demonstrated that overexpression of PagKNAT2/6b in the PdMYB118 overexpression background rescued lignin accumulation and xylem width to the same level of wild type plants. The findings in this work suggest that PdMYB118 is involved in the lignin deposition and xylem differentiation via modulating the expression of PagKNAT2/6b, and the PdMYB118- PagKNAT2/6b model can be used for the genetic breeding of new woody tree with high lignin production., Competing Interests: Declaration of Competing Interest The authors declare no conflict of interest., (Copyright © 2024 Elsevier B.V. All rights reserved.)

DNA methylation plays a crucial role in regulating gene expression, thereby affecting the growth and development of organisms. The application of 5-azacytidine (5-azaC) serves as a potent regulator of DNA methylation levels by inhibiting DNA methyltransferase activity, which subsequently impacts organismal growth and development. In this study, we explored the effects of varying concentrations of 5-azaC on the growth and fruit quality attributes of Merlot grapes. Our findings indicate that treatment with 5-azaC accelerates fruit coloration in grape berries, particularly under conditions of suboptimal solar irradiance. Although 5-azaC treatments at various concentrations suppressed the accumulation of glucose and fructose, the accumulation of organic acids was more influenced by climatic factors than by 5-azaC exposure. Notably, high concentrations (200 μM) of 5-azaC significantly enhanced anthocyanin content, while subhigh concentrations (100 μM) had a contrasting effect. Genome-wide methylation profiling revealed that treatment with 5-azaC reduced CpG methylation levels, thereby affecting the transcriptional regulation of key genes involved in pertinent metabolic pathways. Expression levels of C4H, PAL, NAD-IDH, and SAI were markedly downregulated following treatment with 5-azaC. Collectively, our results demonstrate that 5-azaC modulates grape DNA methylation, stimulates grape growth and development, promotes anthocyanin accumulation, but also concurrently diminishes the levels of sugars and organic acids in mature grape fruits., Competing Interests: Declaration of competing interest All authors disclosed no relevant relationships., (Copyright © 2024. Published by Elsevier Masson SAS.)

Flower color change, a common phenomenon that is important in pollination ecology, has intrigued scientists for decades. While previous flower color studies have mainly focused on color diversity among different plant species, our focus is on unraveling the mechanism of post-anthesis color change (PACC) and the molecular basis for its presence and absence, respectively, in two closely related species of Lotus, Lotus filicaulis and Lotus japonicus MG20. Metabolomic analysis reveals anthocyanins as the key metabolites responsible for the observed PACC. Differential expression of anthocyanin biosynthetic and transport genes causes the variation in PACC between the two Lotus species. Crucially, the significant upregulation of a functionally characterized MYB regulator, LfPAP1, is linked to the accumulation of anthocyanins and visible color alterations in L. filicaulis flowers. Notably, we uncover a nucleotide polymorphism in the initiation codon of LjPAP1. Although this mutation does not affect transcription, we show that it has a major effect in attenuating protein translation, reducing its capacity to activate anthocyanin biosynthesis, and leading to a failure of PACC in L. japonicus MG20. Our study sheds light on mechanisms of PACC phenomenon and highlights the potential for mutations in initiation sequences to generate phenotypic differences between species in evolution., (© 2024 Society for Experimental Biology and John Wiley & Sons Ltd.)

Plant-derived biostimulants have gained attention in agricultural practices for their potential to enhance crop quality and resilience. In this study, we investigated the effects of applying a maize gluten-derived protein hydrolysate at the soil level in vineyards on berry quality in a table grape variety, the Black Magic early table grapevine, during veraison. Our results demonstrate significant improvements in various parameters 14 days after application, including increased anthocyanin levels, enhanced sugar accumulation, and larger berry diameter while maintaining berry firmness. Transcriptomic analysis revealed mechanisms underlying these effects, highlighting the biostimulant's ability to expedite ripening processes while selectively modulating genes associated with cell wall metabolism, thus explaining the observed preservation of berry firmness. Furthermore, the treatment with a gluten-derived protein hydrolysate enhanced the grapevine's resilience to abiotic and biotic stresses, and several related genes were affected. This study sheds light on the potential of plant-derived biostimulants in grapevine cultivation, emphasizing the need for further research to elucidate their mechanisms and optimize agricultural practices., (© 2024 Scandinavian Plant Physiology Society.)

5-Aminolevulinic acid (ALA), a key biosynthetic precursor of tetrapyrrole compounds, significantly induces anthocyanin accumulation in apple (Malus × domestica Borkh.) as well as other fruits. Although the molecular mechanisms of ALA-induced anthocyanin accumulation have been reported, it remains unknown whether the metabolism of ALA is involved in ALA-induced anthocyanin accumulation. Here, we found that MdFC2, a gene encoding ferrochelatase (MdFC2), which catalyzes the generation of heme from protoporphyrin lX (PPIX), may play an important role in ALA-induced apple anthocyanin accumulation. Exogenous ALA induced the MdFC2 expression as well as anthocyanin accumulation in apple leaves, calli, and isolated fruits. MdFC2 overexpression in apple leaves or calli significantly enhanced anthocyanin accumulation as well as the expression of genes involved in anthocyanin biosynthesis, while RNA interference MdFC2 inhibited anthocyanin accumulation and the expression of genes involved in anthocyanin biosynthesis. When 2,2'-dithiodipyridine, an inhibitor of MdFC2, was added, ALA-induced anthocyanin accumulation was blocked. These results suggest that ALA-induced anthocyanin accumulation of apple may be regulated by heme or its biosynthesis, among which MdFC2 or MdFC2 may play a critical positive regulatory role. This finding provides a novel insight to explore the mechanisms of ALA-regulating physiological processes and better application of ALA in high-quality fruit production., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier GmbH. All rights reserved.)

Chilling stress seriously affects the growth and yield of tomato. Anthocyanin is a typical chilling-induced metabolite with strong antioxidant activity and photoprotective capacity. Here, we found that anthocyanin was also involved in ascorbic acid biosynthesis and glycolysis under chilling stress. SlAN2 is an important positive gene in anthocyanin biosynthesis. The results of physiological indicators showed that SlAN2 overexpression lines (A189) had a greater ability to tolerate cold stress than wild-type (WT) plants. Conjoint analysis of transcriptomics and metabonomics of A189 lines and WT plants was used to analyze the metabolic difference and the cold resistance mechanisms caused by anthocyanin under chilling stress. The anthocyanin accumulated more in A189 than that in WT under chilling stress at 4 °C for 24 h, which led to hexoses and ascorbic acid increased significantly. Results indicate that SlAN2 overexpression reduces the expression of key enzyme genes in glycolytic pathway such as phosphofructokinase (PFK) and pyruvate kinase (PK) genes, weakens glycolysis ability, and promotes accumulation of hexoses in A189 lines at 4 °C for 24 h compared with wild lines. Additionally, ascorbic acid content is increased by up-regulated the genes of ascorbate peroxidase (APX) and dehydroascorbate reductase (DHAR). The increased hexose content can reduce cell osmotic potential, freezing point and synthesize more ascorbic acid, while the increased ascorbic acid content can enhance the ability to scavenge reactive oxygen species, so improves the cold resistance of tomato. The glycolysis and ascorbic acid metabolism pathway mediated by SlAN2 provides a new insight for the molecular mechanism of anthocyanins in improving the cold resistance of tomato and provides a new theoretical basis for cultivating new cold-tolerant tomato varieties., Competing Interests: Declaration of competing interest The authors declare that they have no competing interests., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Crabapple is a valuable tree species in gardens due to its captivating array of flower and leaf colors, rendering it a favored choice in landscaping. The economic and ornamental values of Malus crabapple are closely associated with the biosynthesis of anthocyanin, a pigment responsible for its vibrant hues. The intricate regulation of anthocyanin biosynthesis involves the concerted activity of various genes. However, the specific mechanism governing this process in crabapple warrants in-depth exploration. In this study, we explored the inhibitory role of MsMYB62-like in anthocyanin biosynthesis. We identified MsDFR and MsANS as two downstream target genes of MsMYB62-like. These genes encode enzymes integral to the anthocyanin biosynthetic pathway. The findings demonstrate that MsMYB62-like directly binds to the promoters of MsDFR and MsANS , resulting in the downregulation of their expression levels. Additionally, our observations indicate that the plant hormone cytokinins exert a suppressive effect on the expression levels of MsMYB62-like , while concurrently upregulating MsDFR and MsANS . This study reveals that the MsMYB62-like- MsDFR / MsANS module plays an important role in governing anthocyanin levels in Malus crabapple. Notably, the regulatory interplay is modulated by the plant hormone cytokinins.

Fruit ripening is a highly-orchestrated process that requires the fine-tuning and precise control of gene expression, which is mainly governed by phytohormones, epigenetic modifiers, and transcription factors. How these intrinsic regulators coordinately modulate the ripening remains elusive. Here we report the identification and characterization of FvALKBH10B as an N 6 -methyladenosine (m 6 A) RNA demethylase necessary for the normal ripening of strawberry (Fragaria vesca) fruit. FvALKBH10B is induced by phytohormone abscisic acid (ABA), and ABA-Responsive Element Binding Factor 3 (FvABF3), a master regulator in ABA signaling, is responsible for this activation. FvALKBH10B mutation leads to a delay in fruit ripening and causes global m 6 A hypermethylation of 1859 genes. Further analyses show that FvALKBH10B positively modulates the mRNA stability of SEPALLATA3 (FvSEP3) encoding a transcription factor via m 6 A demethylation. In turn, FvSEP3 targets numerous ripening-related genes including those associated with biosynthesis of ABA and anthocyanin and regulates their expression. Our findings uncover an FvABF3-FvALKBH10B-FvSEP3 cascade in controlling fruit ripening in strawberry and provide insights into the complex regulatory networks involved in this process., Competing Interests: Competing interests: The authors declare no competing interests., (© 2024. The Author(s).)

Cinnamomum camphora, a key multifunctional tree species, primarily serves in landscaping. Leaf color is crucial for its ornamental appeal, undergoing a transformation to red that enhances the ornamental value of C. camphora. However, the molecular mechanisms underlying this transformation remain largely unexplored. In this study, green leaf (GL), color turning red leaf (RL) and whole red leaf (WRL) were obtained to measure pigment contents, while GL and RL were analyzed for transcriptomic alterations. A decline in chlorophyll content and a rise in anthocyanins were observed during the transition from green to red leaves. Using LC MS/MS, 11 types of anthocyanins showed significant accumulative differences, with cyanidin-3,5-O-diglucoside exhibiting the greatest disparity. Comparative RNA-seq identified 22,948 genes against reference genes, revealing 544 novel genes. Of these, 3,222 genes were up-regulated and 7,391 genes were down-regulated when the FPKM mean value > 1 in at least one group. The ribosome was identified as the most abundant KEGG term, with a substantial number of down-regulated differentially expressed genes (DEGs). The results indicated a downward trend in protein content, with GL exhibiting the highest protein concentration. 22, 4, and 29 DEGs were associated with chlorophyll biosynthesis, chlorophyll degradation, and anthocyanin biosynthesis, respectively. Most DEGs related to chlorophyll biosynthesis were down-regulated. SGR and SGRL, which are associated with chlorophyll degradation, exhibited opposite differential expression, resulting in a significant decrease in chlorophyll content in RL. The significantly up-regulated genes ANS and UFGT are advantageous for anthocyanin biosynthesis, contributing to the red coloration observed. Additionally, differential expression was noted in 40 R2R3-MYBs. Two MYB90 (Ccam01G003512 and Ccam01G003515) homologs of AtMYB113 were also identified showed high levels of up-regulation in RL. These findings suggest a strong correlation between pigment metabolism and transcriptome data, elucidating the mechanism that leads to the red coloration of leaves in C. camphora., Competing Interests: Declarations. Competing interests: The authors declare no competing interests., (© 2024. The Author(s).)

Anthocyanins affect quality in fruits such as grape (Vitis vinifera). High temperatures reduce anthocyanin levels by suppressing the expression of anthocyanin biosynthesis genes and decreasing the biosynthetic rate. However, the regulatory mechanisms that coordinate these 2 processes remain largely unknown. In this study, we demonstrate that high-temperature-mediated inhibition of anthocyanin biosynthesis in grape berries depends on the auxin and endoplasmic reticulum (ER) stress pathways. Inactivation of these pathways restores anthocyanin accumulation under high temperatures. We identified and characterized FAR-RED ELONGATED HYPOCOTYL3 (FHY3), a high-temperature-modulated transcription factor that activates multiple anthocyanin biosynthesis genes by binding to their promoters. The auxin response factor VvARF3 interacts with VvFHY3 and represses its transactivation activity, antagonizing VvFHY3-induced anthocyanin biosynthesis. Additionally, we found that the ER stress sensor VvbZIP17 represses anthocyanin biosynthesis. VvFHY3 suppresses VvbZIP17 activity by directly binding to the VvbZIP17 promoter to repress its transcription and by physically interacting with VvbZIP17 to block its DNA binding ability. Furthermore, AUXIN RESPONSE FACTOR 3 (ARF3) interferes with the VvFHY3-VvbZIP17 interaction, releasing VvbZIP17 to activate the unfolded protein response and further suppress anthocyanin production. Our results unravel the VvARF3-VvFHY3-VvbZIP17 regulatory module, which links the auxin and ER stress pathways to coordinately repress anthocyanin structural gene expression and biosynthesis under high-temperature stress., Competing Interests: Conflict of interest statement. The authors declare no conflict of interests., (© The Author(s) 2024. Published by Oxford University Press on behalf of American Society of Plant Biologists. All rights reserved. For commercial re-use, please contact reprints@oup.com for reprints and translation rights for reprints. All other permissions can be obtained through our RightsLink service via the Permissions link on the article page on our site—for further information please contact journals.permissions@oup.com.)

Temperature is a critical abiotic factor affecting rice (Oryza sativa L.) yields, and cold stress at the seedling stage can inhibit plant growth or even be fatal. Antioxidants such as anthocyanins accumulate in a variety of plants during cold stress, but the underlying mechanisms are not well understood. Here, we report that rice TRANSPARENT TESTA GLABRA 1 (OsTTG1), a major regulator of anthocyanin biosynthesis in rice, responds to short- and long-term cold stress at both the transcriptional and protein levels. Metabolomic and transcriptomic data indicate that OsTTG1 activates the expression of anthocyanidin synthase (OsANS) genes under cold stress. Our data also suggest that OsTTG1 forms a MYB-bHLH-WD (MBW) complex with Basic helix-loop-helix 148 (OsbHLH148) and Myb-related S3 (OsMYBS3), and this complex activates the expression of Dehydration-responsive element-binding protein 1 (OsDREB1) and OsANS genes. Together, our findings reveal the mechanisms by which OsTTG1 coordinates both anthocyanin biosynthesis and the expression of cold-responsive genes in colored rice, providing genetic resources for future cold resistance breeding in rice., Competing Interests: Conflict of interest statement. The authors declare no conflicts of interest for this work., (© The Author(s) 2024. Published by Oxford University Press on behalf of American Society of Plant Biologists. All rights reserved. For commercial re-use, please contact reprints@oup.com for reprints and translation rights for reprints. All other permissions can be obtained through our RightsLink service via the Permissions link on the article page on our site—for further information please contact journals.permissions@oup.com.)

Rapeseed (Brassica napus L.) possesses substantial economic value as an oil, vegetable, and forage crop, while also exhibiting notable ornamental characteristics. Recent advances in flower colour breeding have significantly enhanced the visual appeal of rapeseed, with anthocyanins identified as the primary contributor to the development of red, purple, and pink flowers. However, the mechanisms underlying the synthesis and regulation of anthocyanins during petal coloration in rapeseed are still poorly understood. This research combined miRNA and mRNA expression data from four different color phases, along with degradome analysis, to discover important miRNA-mRNA modules responsible for controlling the accumulation of anthocyanin in purple-flowered rapeseed. In the process of petal development, a grand sum of 247 miRNAs (including 223 known and 24 novel miRNAs) were effectively detected, with 64 of them displaying differential expression patterns. Degradome sequencing was used to conduct a comprehensive analysis of 152 targets for the differential expression miRNAs. Out of these, 108 miRNA-mRNA modules exhibit contrasting expression patterns. Some miRNAs and their corresponding targets have additionally been discovered, potentially playing a role in governing the buildup of anthocyanin in purple-flowered rapeseed. The regulatory modules miR156-SPL9 and miR828-PAP2 composed of miR156b and miR828 and their targets may play a key role in this process. The results offer a thorough analysis of miRNAs linked to the regulation of anthocyanin in B. napus, offering valuable understanding into the regulatory processes that govern miRNA-mediated anthocyanin production in Brassica crops., Competing Interests: Declarations. Ethics approval and consent to participate: This study including the collection on plants material complies with relevant institutional, national, and international guidelines and legislation. Consent for publication: Not applicable. Competing interests: The authors declare no competing interests., (© 2024. The Author(s).)

Main Conclusion: This study revealed a substrate-level synthesis of pigment cyanidin-3-O-glucoside and the redirection of metabolomic flux in the flavonoid/anthocyanin biosynthesis pathway in poplar adventitious roots (ARs) induced by stem canker pathogens. Recently, we observed a novel allometry on poplar stems, with copious colorful adventitious roots (ARs) induced by fungal canker pathogens. Here, we reveal chemical, physiological, and molecular mechanisms of AR coloration in poplar-pathogens (Valsa sordida/Botrosphaeria dothidea) interaction system using our phloem girdling-inoculation system. Light-induced coloration in ARs: red/rosy under sunlight, and milky white under shading. Chemical and metabolomic analyses indicated that numerous (93 in all 110) and high relative intensities/contents of flavonoids metabolites (mainly including flavonols, flavones, and anthocyanins class) accumulate in red ARs, some flavones and anthocyanins metabolites all contribute to the color of poplar ARs, and cyanidin-3-O-glucoside is the most abundant colorant. Integrated analysis of metabolomic and transcriptomic analysis suggested that sunlight exposure redirected the metabolomic flux from the flavonoid biosynthesis pathway to the flavonols and flavones branch pathways, induced by the upregulation of FLS (flavonol synthase/flavanone 3-hydroxylase) and other structural genes. The anthocyanins metabolomic analysis and the downregulation of the ANS (anthocyanin synthase) gene illustrated a retard of metabolomic flux from leucoanthocyanidins to anthocyanidins. Metabolomic results and the upregulation of the gene BZ1 (Bronze 1, anthocyanin 3-O-glucosyltransferase) illustrated that sunlight triggered a rapid biosynthesis of anthocyanin metabolites in poplar ARs, which based on the substrate level of anthocyanidins. Transcriptomic and RT-qPCR analyses showed that transcriptional factor MYB113, HY5 (Elongated hypocotyl 5), and COP1 (Ring-finger protein CONSTITUTIVE PHOTOMORPHOGENIC1) genes positively regulated the expression of the flavonoid/anthocyanin biosynthesis structural genes (such as the BZ1, FLS and LAR gene) in both sunlight-exposed red ARs and white ARs after light-exposure, suggesting sunlight induces anthocyanins biosynthesis through the interaction between "MBW" complex and COP1-HY5 module. Moreover, one SPL gene (squamosa promoter-binding-like protein gene, target of miR156, and one component of miR156-SPL module) was down-regulated in sunlight-exposed poplar ARs, implying the biosynthesis flavonoid/anthocyanin be regulated at the posttranscriptional level. This study provides a potential AR experimental system for research on flavonoid/anthocyanin biosynthesis in tree species., Competing Interests: Declarations. Conflict of interest: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Anthocyanin regulation can be fruitfully explored from a diverse perspective by studying distantly related model organisms. Land plants pioneers faced a huge evolutionary leap, involving substantial physiological and genetic changes. Anthocyanins have evolved alongside these changes, becoming versatile compounds capable of mitigating terrestrial challenges such as drought, salinity, extreme temperatures and high radiation. With the accessibility of whole-genome sequences from ancient plant lineages, deeper insights into the evolution of key metabolic pathways like phenylpropanoids have emerged. Despite understanding the function of anthocyanins under stress, gaps remain in uncovering the precise metabolic and regulatory mechanisms driving their overproduction under stressful conditions. For example, the regulatory effect of reactive oxygen species (ROS) on well-known transcription factors like MYBs is not fully elucidated. This manuscript presents an evolutionary analysis of the anthocyanin biosynthetic pathway to elucidate key genes. CINNAMATE 4-HYDROXYLASE (C4H) and CHALCONE ISOMERASE (CHI2) received particular attention. C4H exposes remarkable differences between aquatic and land plants, while CHI2 demonstrates substantial variation in gene copy number and sequence similarity across species. The role of transcription factors, such as MYB, and the involvement of ROS in the regulation of anthocyanin biosynthesis are discussed. Complementary gene expression analyses under abiotic stress in Arabidopsis thaliana, Selaginella moellendorffii, and Marchantia polymorpha reveal intriguing gene-stress relationships. This study highlights evolutionary trends and the regulatory complexity of anthocyanin production under abiotic stress, providing insights and opening avenues for future research., Competing Interests: Declarations. Conflict of interest: The authors declare no conflict of interests., (© 2024. The Author(s), under exclusive licence to Springer Nature B.V.)

Nut kernel color is a crucial quality indicator affecting the consumers first impression of the product. While growing evidence suggests that plant phenolics and their derivatives are linked to nut kernel color, the compounds (biomarkers) responsible for kernel color stability during storage remain elusive. Here, pathway-based metabolomics with machine learning algorithms were employed to identify key metabolites of postharvest pecan color stability. Metabolites in phenylpropanoid, flavonoid, and anthocyanin biosynthetic pathways were analyzed in the testa of nine pecan cultivars using liquid chromatography-mass spectrometry. With color measurements, different machine learning models were compared to find relevant biomarkers of pecan color phenotypes. Results revealed potential marker compounds that included flavonoid precursors and anthocyanidins as well as anthocyanins (e.g., peonidin, delphinidin-3-O-glucoside). Our findings provide a foundation for future research in the area, and will help select genes/proteins for the breeding of pecans with stable and desirable kernel color., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Phenol-pyranoanthocyanins, a structurally modified type of anthocyanin, has higher stability than anthocyanins. However, their conversion occurs slowly. Therefore, it is crucial to improve the conversion efficiency and production of pyranoanthocyanins. In this study, cranberry anthocyanin (CRAN) was fermented using two Lactobacillus strains along with caffeic acid to form cranberry-derived pyranoanthocyanins (PY-CRAN). PY-CRAN was characterized and identified. The physicochemical properties, antioxidant activity, and tyrosinase inhibitory capacity of PY-CRAN were assessed. The results showed that phenol-pyranoanthocyanins can be rapidly produced through fermentative transformation using Lactiplantibacillus plantarum and Lacticaseibacillus paracasei. Lacticaseibacillus paracasei exhibits a higher propensity for producing phenol-pyranoanthocyanins. PY-CRAN exhibits high stability under light and various pH conditions. Moreover, they possess excellent antioxidant properties and the ability to inhibit tyrosinase. These results suggest that fermentative biotransformation conducted by Lactobacillus is an ideal method for producing cranberry pyranoanthocyanins. The resulting anthocyanins have potential as antioxidant and whitening agents, making them promising bioactive ingredients., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

The color variation of the leaves in autumn is a significant ornamental feature of Acer truncatum Bunge, especially when the leaves gradually become redder. Many studies focused on leaf color changes; however, less research has been conducted on the mechanism by which A. truncatum 's autumn leaves turn red. Red, middle and green leaves of Acer truncatum were used as the study materials to evaluate their flavonoid-related metabolites and infer gene and metabolite expression patterns in conjunction with transcriptome expression. For a start, phenotypic and leaf color parameters analyses showed that red leaves had the highest color redness and greenness (a*). In addition, a total of 23 flavonoid-related metabolites were identified through the metabolome, including five anthocyanins. Of them, cyanidin 3-O-β-D-sambubioside, cyanidin 3-O rutinoside, pelargonidin 3-O-3″,6″-O-dimalonylglucoside, delphinidin 3,7-di-O-β-D-glucoside and 3-O-β-D-sambubioside would help the leaves turn red in A. truncatum . Similarly, combined transcriptomics and metabolomics analyses showed that most genes in the flavonoid and anthocyanin biosynthetic pathways were differentially expressed in both types of leaves. Chalcone synthase ( CHS ), dihydroflavonol 4-reductase ( DFR ) and anthocyanin synthase ( ANS ) could affect flavonoid synthesis during leaf color change. This study could provide data for the genetic improvement of maple plants by exploring valuable metabolites and genes in flavonoid synthesis, and enhance the understanding of different developmental stages.

Methyl jasmonate (MeJA) is a growth regulator that is involved in plant defense and development. Studies have demonstrated its role in pigmentation, particularly in synergy with UV-B in plant tissues. However, its role in pigment metabolism remains incompletely understood. To explore the metabolic synergistic effects, we evaluated pigments and gene expression in mature green apples. MeJA alone had no effect on pigment accumulation, while UV-B increased flavonols up to S2 stage and anthocyanins up to S3 stage. With UV-B > 2 W m -2 , MeJA doubled anthocyanin accumulation compared to UV-B alone but had no synergistic effect on flavonols. MeJA selectively upregulated structural genes and transcription factors involved in the late biosynthetic pathway, transport, and the MYB-bHLH-WD40 complex, inducing anthocyanin hyperaccumulation without affecting early flavonoid biosynthetic genes or MdFLS , which controls flavonol biosynthesis. These results suggest that MeJA relates to downstream gene expression in flavonoid biosynthesis with alternative regulation of the anthocyanin pathway in UV-B-irradiated apples.

Anthocyanins, a group of flavonoids, play diverse roles in plant growth and environmental adaptation. The biosynthesis and accumulation of anthocyanin are regulated by environmental cues, such as high light. However, the precise mechanism underlying anthocyanin biosynthesis under high light conditions remains largely unclear. Here, we report that the R3-MYB repressor MYB-LIKE 2 (MYBL2) negatively regulates high light-induced anthocyanin biosynthesis in Arabidopsis by repressing two R2R3-MYB activators, PRODUCTION OF ANTHOCYANIN PIGMENT 1 (PAP1) and PAP2, which are core components of the MYB-bHLH-WD40 (MBW) complex. We found that MYBL2 interacts with PAP1/2 and reduces their transcriptional activation activities, thus disrupting the expression of key genes involved in anthocyanin biosynthesis, such as DIHYDROFLAVONOL 4-REDUCTASE (DFR) and TRANSPARENT TESTA 19 (TT19). Additionally, MYBL2 attenuates the transcriptional activation of PAP1 and its own expression, but not that of PAP2. Conversely, PAP1 collaborates with TRANSPARENT TESTA 8 (TT8), a bHLH member of the MBW complex, to activate MYBL2 transcription when excessive anthocyanins are accumulated. Taken together, our findings reveal a negative feedback regulatory module composed of MYBL2 and PAP1 that fine-tunes high light-induced anthocyanin biosynthesis through modulating MBW complex assembly., (© The Author(s) 2024. Published by Oxford University Press on behalf of the Society for Experimental Biology. All rights reserved. For commercial re-use, please contact reprints@oup.com for reprints and translation rights for reprints. All other permissions can be obtained through our RightsLink service via the Permissions link on the article page on our site—for further information please contact journals.permissions@oup.com.)

Background: The anthocyanin synthase (ANS) gene is pivotal in plant anthocyanin biosynthesis, significantly influencing fruit coloration. Despite extensive on various studies, there remains a notable absence of research on the regulatory functions of the ANS gene in the 'Lingwuchangzao' variety under elevated temperature and drought stress., Results: In this study, 107 ZjANS genes were identified. Analyses of their physicochemical properties and gene structures revealed that most are localized in the cytosol and nucleus, with only seven ZjANS genes exhibiting basic characteristics. All genes contained the conserved domains 2OG-FeII_Oxy and DIOX-N. Chromosomal localization and Ka/Ks analysis indicated that these genes are distributed across 12 chromosomes, with 22 tandem gene pairs, suggesting purifying selection. Phylogenetic and synteny analysis demonstrated that ZjANS genes are evolutionarily conserved, forming collinear gene pairs with four species. Cis-acting element analysis identified elements related to anthocyanin synthesis and environmental stress, such as LTR and MBS. Predictions of protein interaction indicated that certain ZjANS proteins interact with MYB, CHI, and GA proteins, potentially involved in the anthocyanin synthesis pathway. Transcriptome and qRT-PCR analysis revealed that thirteen ZjANS genes positively respond to coloring in jujube under elevated temperature and drought stress, with ZjANS17 identified as particularly significant., Conclusion: This study identified 107 ZjANS genes and systematically analyzed their evolutionary conservation. Thirteen ZjANS genes were found to positively influence fruit coloring in 'Lingwuchangzao' under elevated temperature and drought stress. These findings provide a foundation for further investigations into ZjANS genes and the breeding new jujube varieties., Competing Interests: Declarations. Ethics approval and consent to participate: The plant material of this study protocol comply with relevant institutional, national, and international guidelines and legislation. All experimental protocols were approved by the Ningxia University. Consent for publication: Not applicable. Competing interests: The authors declare no competing interests., (© 2024. The Author(s).)

Anthocyanins are water-soluble natural pigments found broadly in plants. As members of the flavonoid family, they are widely distributed in various tissues and organs, including roots, leaves, and flowers, responsible for purple, red, blue, and orange colors. Beyond pigmentation, anthocyanins play a role in plant propagation, stress response, defense mechanisms, and human health benefits. Anthocyanin biosynthesis involves a series of conserved enzymes encoded by structural genes regulated by various transcription factors. In rice, anthocyanin-mediated pigmentation serves as an important morphological marker for varietal identification and purification, a critical nutrient source, and a key trait in studying rice domestication. Anthocyanin biosynthesis in rice is regulated by a ternary conserved MBW transcriptional complexes comprising MYB transcription factors (TFs), basic-helix-loop-helix (bHLH) TFs, and WD40 repeat protein, which activate the expression of structure genes. Wild rice (Oryza rufipogon) commonly has purple hull, purple stigma, purple apiculus, purple leaf, and red pericarp due to the accumulations of anthocyanin or proanthocyanin. However, most cultivated rice (Oryza sativa) varieties lose the anthocyanin phenotypes due to the function variations of some regulators including OsC1, OsRb, and Rc and the structure gene OsDFR. Over the past decades, significant progress has been made in understanding the molecular and genetic mechanisms of anthocyanin biosynthesis. This review summarizes research progress in rice anthocyanin biosynthetic pathways, genes involvements, distribution regulations, and domestication processes. Furthermore, it discusses future prospects for anthocyanin biosynthesis research in rice, aiming to provide a theoretical foundation for future investigations and applications, and to assist in breeding new rice varieties with organ-targeted anthocyanin deposition., Competing Interests: Declaration of Competing Interest The authors declare no conflict of interest., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Cold stress has seriously inhibited the growth and development of strawberry during production. CBF/DREB1 is a key central transcription factor regulating plant cold tolerance, but its regulatory mechanisms are varied in different plants. Especially in strawberry, the molecular mechanism of CBF/DREB1 regulating cold tolerance is still unclear. In this study, we found that FveDREB1B was most significantly induced by cold stress in CBF/DREB1 family of diploid woodland strawberry. FveDREB1B was localized to the nucleus, and DREB1B sequences were highly conserved in diploid and octoploid strawberry, and even similar in Rosaceae. And FveDREB1B overexpressed strawberry plants showed delayed flowering and increased cold tolerance, while FveDREB1B silenced plants showed early flowering and decreased cold tolerance. Under cold stress, FveDREB1B activated FveSCL23 expression by directly binding to its promoter. Meanwhile, FveDREB1B and FveSCL23 interacted with FveDELLA, respectively. In addition, we also found that FveDREB1B promoted anthocyanin accumulation in strawberry leaves by directly activating FveCHS expression after cold treatment and recovery to 25°C. DREB1B genes were also detected to be highly expressed in cold-tolerant strawberry resources 'Fragaria mandschurica' and 'Fragaria nipponica'. In conclusion, our study reveals the molecular mechanism of FveDREB1B-FveSCL23-FveDELLA module and FveDREB1B-FveCHS module to enhance the cold tolerance of woodland strawberry. It provides a new idea for improving the cold tolerance of cultivated strawberry and evaluating the cold tolerance of strawberry germplasm resources., (© 2024 John Wiley & Sons Ltd.)

Peach varieties that differ in red coloration due to varied anthocyanin accumulation result from transcriptional regulation by PpMYB10s, a group of specific R2R3 MYBs. Here we investigated the mechanisms driving a lack of anthocyanin in yellow-skinned 'Jinxiu' peach peel, as well as accumulation induced by UV irradiance. It was found that PpMYB10.1, PpMYB10.2 and PpMYB10.3 were positive regulators of anthocyanin accumulation, but the stimulation by PpMYB10.2 was weak. Low expression of PpMYB10.1 causes natural anthocyanin deficiency in 'Jinxiu' peel. However, the promoter sequences of PpMYB10.1 were identical in 'Jinxiu' and a naturally red-coloured peach 'Hujingmilu'. Therefore, potential negative regulator(s) upstream of PpMYB10.1 were explored. A novel R2R3-MYB repressor termed PpMYB80 was identified through comparative transcriptomic analysis and then functionally confirmed via transiently overexpressing and silencing in peach fruit, as well as transformation in tobacco. PpMYB80 directly binds to the promoter of PpMYB10.1 and inhibits its expression, but does not affect PpMYB10.3. In UV-exposed 'Jinxiu' fruit, expression of PpMYB10.3 was upregulated, while PpMYB10.1 remained low and PpMYB80 enhanced, which results in accumulation of anthocyanin in peel. This study revealed a transcriptional cascade involving PpMYB activators and repressors in regulating basal and UV-induced anthocyanin accumulation in peach peel., (© 2024 John Wiley & Sons Ltd.)

The grape hyacinth is renowned for its profuse blue flowers, which confer substantial scientific and ornamental significance as well as considerable potential for industrial applications. The serine carboxypeptidase-like acyltransferases (SCPL-ATs) family is crucial for the blue flower coloration. To elucidate SCPL-ATs involved in anthocyanin modification in grape hyacinth, we performed a transcriptomic analysis of grape hyacinth SCPL-ATs. Through gene expression profiling, we identified a promising candidate gene, MaSCPL1, whose expression patterns corresponded with variations in anthocyanin content throughout petal coloration. Subsequently, the functional role of the MaSCPL1 gene was validated using the native petal regeneration system, and the silencing of MaSCPL1 led to a decreased total anthocyanin content and Dp3MG content in grape hyacinth petals. Furthermore, we employed yeast one-hybrid (Y1H), electrophoretic mobility shift assay (EMSA), and dual-luciferase assays to explore the regulatory interactions between the anthocyanin biosynthesis transcription factor MaMybA and the MaSCPL1 promoter. Our findings indicate that MaMybA can bind to the MaSCPL1 promoter and significantly activate its expression. Furthermore, the MaMybA-RNAi resulted in a substantial multifold reduction in the expression of MaSCPL1, implying that the regulation of MaSCPL1 expression is mediated by MaMybA. This study revealed the MaSCPL1 gene has been associated with anthocyanin acylated modification in grape hyacinth and elucidated the important role of the MaMybA-MaSCPL1 module in colouration grape hyacinth., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

The basic helix-loop-helix (bHLH) family is one of the three major transcription factor families that play important transcriptional regulatory roles in plant growth and development. One of the most crucial elements in defining Lagerstroemia indica's decorative qualities is flower color. However, the function of the bHLH transcription factor family in L. indica anthocyanin glycoside synthesis has not been clarified. Using the transcriptome data of flower color, 79 LibHLH genes were found in this study. Phylogenetic analysis showed that the LibHLH genes can be divided into 16 subfamilies, and most of the genes in the same subfamily have similar conserved motifs. The total anthocyanin glycoside content of L. indica 'Zihua Guifei' petals was determined during three developmental stages of the petals' growth. The results showed that the total anthocyanin glycoside content grew gradually with growth and development, and that it accumulated most during the full bloom stage. By using gene expression analysis, protein interaction network analysis, and bioinformatics, it was possible to determine which member of the III f family, LibHLH29, is important for the synthesis of anthocyanin glycosides in L. indica. Its expression was confirmed by qRT-PCR, and the results were essentially compatible with the transcriptome data. It was more prominent in the light-colored bloom stage the color-transition stage of L. indica 'Zihua Guifei'. It can be further investigated as a major candidate gene for regulating anthocyanin glycoside synthesis in L. indica, thus laying the foundation for an in-depth study of the interactions among transcription factors., (© 2024. The Author(s), under exclusive licence to Springer Nature Switzerland AG.)

The R2R3-MYB transcription factors (TFs) play a crucial role in regulating plant secondary metabolism and abiotic stress responses, yet they are still poorly understood in common buckwheat (Fagopyrum esculentum), a valuable minor grain crop resource. In this study, a candidate gene, FeR2R3-MYB, was cloned from the anthocyanin-rich common buckwheat variety 'QZZTQ'. FeR2R3-MYB was found to contain two MYB DNA-binding domains and be located at the nucleus with transcriptional activation activity. Molecular analysis indicated that FeR2R3-MYB is predominantly expressed in flowering tissue and is highly responsive to environmental factors such as light, drought, and cold. In addition, the promoter of FeR2R3-MYB showed a positive correlation with fragment length. Further functional analysis suggested that FeR2R3-MYB not only participates in the anthocyanin biosynthetic pathway by interacting with leucoanthocyanidin reductase (FeLAR), but also enhances drought tolerance in common buckwheat. To sum up, FeR2R3-MYB exhibits positive effects on both pigment production (e.g., anthocyanin) and abiotic stress resistance, providing valuable insights for future research in buckwheat molecular breeding and resource development., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Masson SAS. All rights reserved.)

Potato (Solanum tuberosum L.) is nowadays an important component of diversified cropping systems due to its adaptability, yielding capacity, and nutrition contribution. Breeding programs aiming at raising potato's nutritional value have mainly focused on the accumulation in potato tubers of health-promoting phytochemicals such as anthocyanins. In different plant species, increased amounts of anthocyanins in vegetative tissues have been associated with enhanced tolerance to abiotic and biotic stresses that challenge agrifood systems in the current context of global climate change. In the present study, we aimed at gaining insight into the effect of anthocyanin accumulation on the potato plants response to drought stress using three different potato genotypes with differential canopy and tuber pigmentation: the purple fleshed commercial variety Bleuet; the red fleshed breeding clone DAR170; and the non-pigmented commercial variety Monalisa. The varieties Bleuet and DAR170 exhibiting higher anthocyanin content in vegetative tissues than the Monalisa variety showed a remarkable inhibition of stem growth development under drought stress treatment suggestive of an anthocyanin-mediated physiological shift from growth to resilience as a mechanism of stress tolerance. The results of the expression analysis of stu-miR156a and its target StSPL9 gene in the potato plants with different anthocyanin content, as well as their change in response to drought stress support the participation of the conserved miR156-SPL9 regulatory module in coordinating potato plants development and plant responses to drought stress, involving precise fine-tuning of anthocyanin biosynthesis., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

Veggie chips have gained popularity in the European market. These are considered healthier than potato chips by consumers. However, few works evaluate their nutritional and digestibility. The current work aimed to evaluate the effect of four pre-frying treatments (soaking, blanching, pulsed electric field (PEF) and PEF + blanching combination (PEFB)) on the chemical composition, anthocyanins, acrylamide, and digestive behavior (starch hydrolysis and anthocyanins bioaccessibility) of purple sweet potato deep-fried chips. In total 15 independent batches were made, three for each studied treatment (also a control without pretreatment was developed). The studied pretreatments impacted on fat and starch content, especially blanching and PEFB, which caused an increase in fat absorption and break starch, generating maltodextrins. Nineteen anthocyanins were detected, mainly cyanidin and peonidin derivatives, but a drastic loss was observed in blanched, PEF-treated and PEF-B-Treated chips. Acrylamide values ranged from 504.11 to 6350.0- μg/kg, with the highest values reported by untreated chips and the lowest by PEF-B-treated chips (p < 0.05). The anthocyanin's bioaccessibility ranged between 66.57 and 92.88%, with soaked chips that showed the highest values., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Anthocyanins are naturally water-soluble pigments of plants, which can be pink, orange, red, purple, or blue. Anthocyanins belong to a subcategory of flavonoids known as polyphenols and are consumed in plant-based foods. The antioxidant properties of anthocyanins benefit human health. However, there has been no comprehensive review of the classification, distribution, and biosynthesis of anthocyanins and their regulation in plants, along with their potential health benefits. In this review, we provide a systematic synthesis of recent progress in anthocyanin research, specifically focusing on the classification, biosynthetic pathways, regulatory mechanisms, bioactivity, and health benefits. We bridge the gaps in understanding anthocyanin biological significance and potential applications. Furthermore, we discuss future directions for anthocyanin research, such as biotechnology, bioavailability, and the integration of artificial intelligence. We highlight pivotal research questions that warrant further exploration in the field of anthocyanin research., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Masson SAS. All rights reserved.)

The blood oranges accumulate anthocyanins in their peel and pulp, which give them their characteristic red colour. To investigate whether there is a specific volatile compounds profile for blood oranges, we compared the volatile composition of the blood varieties 'Moro', 'Tarocco Rosso' and 'Sanguinelli' to that of the blond ones 'Cadenera', 'Salustina' and 'Hamlim' using a non-targeted approach based on HS-SPME-GC-MS. Moreover, fruits from each blood orange variety were divided into slightly and intensely pigmented fruits, and individual sugars and anthocyanins were determined by liquid chromatography to investigate whether their profiles depend on the degree of pulp pigmentation. A total of 101 volatile compounds were identified in this study. Hierarchical Cluster Analysis, including all compounds, revealed that blood oranges have no unique volatiles profile that makes them more similar to one another than to the blond varieties. For blood orange varieties, our results corroborated that greater anthocyanin accumulation in the most intensely pigmented fruit is associated with higher sugar content (an increase of 0.8-2.3 % depending on the variety). Moreover, we reported for the first time that anthocyanin accumulation is also associated with changes in the volatiles profile. A Multifactor Analysis including data on volatiles, sugars and anthocyanin showed that sucrose is the sugar more strongly linked to anthocyanin accumulation. A group of volatile compounds, mainly esters (ethyl acetate, ethyl-2-methyl butanoate and ethyl-2-butenoate) along with ethanol, are present at increased levels, sometimes even doubling in the most intensely pigmented samples compared to the slightly pigmented ones. These results open the door to further investigate the possible metabolic link between sugars and anthocyanins accumulation and changes in volatile compounds., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Fruit colour is a critical determinant for the appearance quality and commercial value of apple fruits. Viroid-induced dapple symptom severely affects the fruit coloration, however, the underlying mechanism remains unknown. In this study, we identified an apple dimple fruit viroid (ADFVd)-derived small interfering RNA, named vsiR693, which targeted the mRNA coding for a bHLH transcription factor MdPIF1 (PHYTOCHROME-INTERACTING FACTOR 1) to regulate anthocyanin biosynthesis in apple. 5' RLM-RACE and artificial microRNA transient expression system proved that vsiR693 directly targeted the mRNA of MdPIF1 for cleavage. MdPIF1 positively regulated anthocyanin biosynthesis in both apple calli and fruits, and it directly bound to G-box element in the promoter of MdPAL and MdF3H, two anthocyanin biosynthetic genes, to promote their transcription. Expression of vsiR693 negatively regulated anthocyanin biosynthesis in both apple calli and fruits. Furthermore, co-expression of vsiR693 and MdPIF1 suppressed MdPIF1-promoted anthocyanin biosynthesis in apple fruits. Infiltration of ADFVd infectious clone suppressed coloration surrounding the injection sites in apple fruits, while a mutated version of ADFVd, in which the vsiR693 producing region was mutated, failed to repress fruit coloration around the injection sites. These data provide evidence that a viroid-derived small interfering RNA targets host transcription factor to regulate anthocyanin biosynthesis in apple., (© 2024 John Wiley & Sons Ltd.)