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Glomerella leaf spot (GLS), a fungal disease caused by Colletotrichum fructicola, severely affects apple (Malus domestica) quality and yield. In this study, we found that the transcription factor MdWRKY71 was significantly induced by C. fructicola infection in the GLS‐susceptible apple cultivar Royal Gala. The overexpression of MdWRKY71 in apple leaves resulted in increased susceptibility to C. fructicola, whereas RNA interference of MdWRKY71 in leaves showed the opposite phenotypes. These findings suggest that MdWRKY71 functions as a susceptibility factor for the apple—C. fructicola interaction. Furthermore, MdWRKY71 directly bound to the promoter of the salicylic acid (SA) degradation gene Downy Mildew Resistant 6 (DMR6)‐Like Oxygenase 1 (DLO1) and promoted its expression, resulting in a reduced SA level. The sensitivity of 35S:MdWRKY71 leaves to C. fructicola can be effectively alleviated by knocking down MdDLO1 expression, confirming the critical role of MdWRKY71‐mediated SA degradation via regulating MdDLO1 expression in GLS susceptibility. In summary, we identified a GLS susceptibility factor, MdWRKY71, that targets the apple SA degradation pathway to promote fungal infection. [ABSTRACT FROM AUTHOR]

Macroautophagy/autophagy is a conserved degradation pathway in eukaryotes that is required for recycling unwanted intracellular components, maintaining homeostasis, and coping with biotic and abiotic stresses. Pathogens have evolved to subvert autophagic machinery by secreting host cell-entering effector proteins. Here, we provided evidence that an apple autophagy-related gene MdATG8i, activated autophagy and contributed to resistance against Valsa canker caused by Valsa Mali (Vm) when being overexpressed in apple. MdATG8i interacted with a plastid elongation factor Tu (MdEF-Tu) which became insoluble and aggregated during Vm infection and was degraded through the autophagy pathway. Intriguingly, we identified a highly-induced effector secreted from Vm, Vm1G-1794, which competitively interacted with MdATG8i, suppressed autophagy, and depleted MdEF-Tu out of MdATG8i complexes. The formation of stable MdEF-Tu aggregates caused by Vm1G-1794 promoted the susceptibility of apple to Vm. Overall, our study demonstrated that MdATG8i contributed to Vm resistance by targeting and degrading MdEF-Tu, and Vm1G-1794 competed with MdEF-Tu to target MdATG8i and prevent MdEF-Tu degradation, thus favoring infection. Abbreviations: 35S: cauliflower mosaic virus 35S promoter; AIM: ATG8-interacting motif; ATG8–PE: ATG8 conjugated with phosphatidylethanolamine; BiFC: biomolecular fluorescence complementation; Con A: concanamycin A; Co-IP: co-immunoprecipitation; DEPs: differentially expressed proteins; DMSO: dimethyl sulfoxide; GFP: green fluorescent protein; hpt: hours post-treatment; LCI: luciferase complementation imaging; MdATG8i: autophagy-related protein 8i in Malus domestica; MDC: monodansylcadaverine; MdEF-Tu: elongation factor Tu in Malus domestica; MdNBR1: neighbor of BRCA1 in Malus domestica; N. benthamiana: Nicotiana benthamiana; OE: overexpression; PAMP: pathogen-associated molecular pattern; PTI: pattern-triggered immunity; qRT-PCR: quantitative reverse transcription PCR; RFP: red fluorescent protein; RNAi: RNA interference; ROS: reactive oxygen species; Ub: ubiquitin; V. Mali: Valsa Mali; WT: wild-type plant; YFP: yellow fluorescent protein [ABSTRACT FROM AUTHOR]

Apples are a rich source of polyphenols in the human diet. However, the distribution of polyphenols in different apple varieties and tissues is still largely unclear. In this study, a new liquid chromatography–tandem mass spectrometry (LC-MS/MS) strategy was developed to reveal the spatial distribution of polyphenols in different apple tissues and varieties. A method based on multiple reaction monitoring (MRM)-enhanced product ion (EPI) was established in the information-dependent acquisition (IDA) mode for pseudo-target screening of major apple polyphenols. A total of 39 apple polyphenolic metabolites were finally identified. Qualitative and quantitative results showed that the variety and content of polyphenols in apple peels were higher than those of other tissues. In apple roots, stems, and leaves, the highest polyphenol variety and content were found in wild species, followed by cultivars and elite varieties. Dihydrochalcone substances, one kind of major apple polyphenols, were more abundant in apple roots, stems, and leaves. This strategy can be applied as a model for other agricultural products, in addition to revealing the distribution of polyphenols in different tissues of apples, which provides a theoretical basis for the utilization of polyphenol resources and variety selection. [ABSTRACT FROM AUTHOR]

The photoprotective role of anthocyanin remains controversial. In this study, we explored the effects of anthocyanin on photosynthesis and photoprotection using transgenic 'Galaxy Gala' apple plants overexpressing MdMYB10 under high light stress. The overexpression of MdMYB10 dramatically enhanced leaf anthocyanin accumulation, allowing more visible light to be absorbed, particularly in the green region. However, through post-transcriptional regulation, anthocyanin accumulation lowered leaf photosynthesis in both photochemical reaction and CO2 fixation capacities. Anthocyanin accumulation also led to a decreased de-epoxidation state of the xanthophyll cycle and antioxidant capacities, but this is most likely a response to the light-shielding effect of anthocyanin, as indicated by a higher chlorophyll concentration and lower chlorophyll a/b ratio. Under laboratory conditions when detached leaves lost carbon fixation capacity due to the limitation of CO2 supply, the photoinhibition of detached transgenic red leaves was less severe under strong white, green, or blue light, but it became more severe in response to strong red light compared with that of the wild type. In field conditions when photosynthesis was performed normally in both green and transgenic red leaves, the degree of photoinhibition was comparable between transgenic red leaves and wild type leaves, but it was less severe in transgenic young shoot bark compared with the wild type. Taken together, these data show that anthocyanin protects plants from high light stress by absorbing excessive visible light despite reducing photosynthesis. [ABSTRACT FROM AUTHOR]

Drought and nitrogen (N) deficiency are common factors that limit apple production in the Loess Plateau region of China. Different apple cultivars respond to drought and low N differently; however, the mechanism that underlies the difference in nitrogen-use efficiency (NUE) under drought conditions is not well understood. In this study, by comparing the physiological responses of two apple (Malus domestica Borkh.) cultivars with contrasting NUE, 'Qinguan' (higher NUE) and 'Honeycrisp' (lower NUE), under low N and drought conditions, we discovered that, 'Qinguan' had larger stomatal apertures, higher chlorophyll fluorescence levels, more active N metabolism and antioxidant enzymes, higher abscisic acid and auxin concentrations, larger root size and more efficient N uptake mediated by higher expression of MdNRT2.4 in rootstock than that of 'Honeycrisp'. Additionally, we experimentally confirmed that MdNRT2.4 enhanced low N and osmotic stress tolerance in Arabidopsis when being overexpressed. Taken together, our findings shed light on the mechanism that underlies the difference in NUE of apple under drought and N-deficient conditionss and provide MdNRT2.4 as a candidate gene for future genetic engineering. [ABSTRACT FROM AUTHOR]

Abstract: Effects of light irradiation on the photosynthetic electron transport chain between P680 and P700 in apple tree leaves was probed with chlorophyll a fluorescence transient and 820 nm transmission measurements during dehydration under different light intensities. The results showed that light accelerated the leaf water-loss rate during dehydration. Leaf dehydration lowered the maximum quantum yield of PSII and the far-red light induced maximal transmission change at 820 nm, but increased the relative variable fluorescence intensity at J-step, especially under increasing irradiation conditions. During leaf dehydration, irradiation lowered the relative variable fluorescence intensity at I-step. At the beginning of leaf dehydration, moderate light accelerated the leaf water-loss rate and then lowered the maximal light-trapping efficiency of P680. Upon further dehydration under moderate light or dehydration under high light, light accelerated the water-loss rate and also directly decreased the maximal light-trapping efficiency of P680. The more significant decrease in the exchange capacity of plastoquinones at the QB site was mainly attributed to the faster water-loss rate under moderate light than in the dark. Under high light, irradiation also directly lowered the capacity. The reoxidation of PQH2 in the dehydrated leaves was enhanced by the light irradiation. The rapidly decreased contents of P700 + plastocyanin were mainly attributed to the faster water-loss rate under light conditions in contrast with that in the dark. The different effects of light irradiations on the photosynthetic electron transport chain might be involved in the acclimation of apple tree leaves to dehydration. [Copyright &y& Elsevier]

SUMMARY Flavonols are widely synthesized throughout the plant kingdom, playing essential roles in plant physiology and providing unique health benefits for humans. Their glycosylation plays significant role in improving their stability and solubility, thus their accumulation and function. However, the genes encoding the enzymes catalyze this glycosylation remain largely unknown in apple. This study utilized a combination of methods to identify genes encoding such enzymes. Initially, candidate genes were selected based on their potential to encode UDP‐dependent glycosyltransferases (UGTs) and their expression patterns in response to light induction. Subsequently, through testing the in vitro enzyme activity of the proteins produced in Escherichia coli cells, four candidates were confirmed to encode a flavonol 3‐O‐galactosyltransferase (UGT78T6), flavonol 3‐O‐glucosyltransferase (UGT78S1), flavonol 3‐O‐xylosyltransferase/arabinosyltransferase (UGT78T5), and flavonol 3‐O‐rhamnosyltransferase (UGT76AE22), respectively. Further validation of these genes' functions was conducted by modulating their expression levels in stably transformed apple plants. As anticipated, a positive correlation was observed between the expression levels of these genes and the content of specific flavonol glycosides corresponding to each gene. Moreover, overexpression of a flavonol synthase gene, MdFLS, resulted in increased flavonol glycoside content in apple roots and leaves. These findings provide valuable insights for breeding programs aimed at enriching apple flesh with flavonols and for identifying flavonol 3‐O‐glycosyltransferases of other plant species. [ABSTRACT FROM AUTHOR]

Sunlight radiation is a main environmental factor which affects anthocyanin synthesis. To clarify the regulatory mechanism of sunlight on the synthesis of anthocyanin in apple peel, bagged apples were exposed to diverse intensities of sunlight through different shading treatments. Under an increased solar ultraviolet-B (UV-B) light intensity, the concentration of anthocyanin in apple peels was consistent with the Michaelis–Menten equation. Under lower sunlight intensities, diphenyleneiodonium chloride (DPI, an inhibitor of plasma membrane NAD(P)H oxidase) treatment increased both the concentration of cyanidin-3-glycoside and the activity of dihydroflavonol 4-reductase (DFR). However, under higher sunlight intensities, DPI treatment decreased the concentrations of cyanidin-3-glycoside and quercetin-3-glycoside, as well as the activities of DFR and UDP-glycose: flavonoid 3-O-glycosyltransferase (UFGT). These results indicate that, under low sunlight intensity, anthocyanin synthesis in apple peel was limited by the supply of the substrate cyanidin, which was regulated by the DFR activity. Nevertheless, after exposure to high sunlight intensity, the anthocyanin produced in the apple peel was dependent on UFGT activity. [ABSTRACT FROM AUTHOR]

The NITRATE TRANSPORTER 1/PEPTIDE TRANSPORTER family (NPF) proteins play important roles in moving substrates such as nitrate, peptides, amino acids, dicarboxylates, malate, glucosinolates, indole acetic acid (IAA), abscisic acid (ABA), and jasmonic acid. Although a unified nomenclature of NPF members in plants has been reported, this gene family has not been studied as thoroughly in apple (Malus × domestica Borkh.) as it has in other species. Our objective was to provide general information about apple MdNPFs and analyze the transcriptional responses of some members to different levels of nitrate supplies. We identified 73 of these genes from the apple genome and used phylogenetic analysis to organize them into eight major groups. These apple NPFs are structurally conserved, based on alignment of amino acid sequences and analyses of phylogenetics and conserved domains. Examination of their genomic structures indicated that these genes are highly conserved among other species. We monitored 14 cloned MdNPFs that showed varied expression patterns under different nitrate concentrations and in different tissues. Among them, NPF6.5 was significantly induced by both low and high levels of nitrate. When compared with the wild type, 35S:MdNPF6.5 transgenic apple calli were more tolerant to low-N stress, which demonstrated that this gene confers greater capacity for nitrogen uptake under those conditions. We also analyzed the expression patterns of those 73 genes in various tissues. Our findings benefit future research on this family of genes. [ABSTRACT FROM AUTHOR]

Abstract: Fractions of absorbed light energy consumed via photochemistry and different thermal dissipation processes was quantified and compared between the sun-exposed peel and the shaded peel of apple fruits at different developmental stages. During fruit development, the fraction of absorbed light consumed via photochemistry was no more than 7% in the sun-exposed peel and no more than 5% in the shaded peel under high light conditions. Under high light, the fraction of absorbed light energy consumed via light dependent thermal dissipation was higher whereas that via constitutive thermal dissipation was lower in the sun-exposed peel. The light dependent thermal dissipation in the sun-exposed peel mainly depended on the xanthophyll cycle, and the xanthophyll cycle pool size was significantly larger in the sun-exposed peel than in the shaded peel. The light dependent thermal dissipation in the shaded peel was dependent on both the xanthophyll cycle and the presence of inactivated reaction centers. Under high light conditions, the densities of both QA-reducing reaction centers and QB-reducing reaction centers decreased faster in the shaded peel than in the sun-exposed peel. The thermal dissipation related to photoinhibition increased and then kept unchanged in the sun-exposed peel but decreased in the shaded peel during fruit development. We conclude that under high light intensities, fruit peel looses the excess energy in order of predominance: first by the xanthophyll cycle, then the thermal dissipation related to photoinhibition, next through inactivated reaction centers, and finally by constitutive thermal dissipation. [Copyright &y& Elsevier]

To investigate the effects of relative air humidity (AH) on ‘Fuji’ apple ( Malus domestica Borkh.) peel phenolic compounds and pigment coloration, the bagged fruits were bags moved and then exposed to high AH and low AH conditions, respectively. Cyanidin-3-galactoside, the main anthocyanin compound in apple, and 17 other types of phenolic compounds were detected and their concentrations were also compared between high and low AH treated apple peels. We observed that the removal of bags and subsequent re-exposure to sunlight could significantly decrease the concentrations of syringic acid, caffeic acid, ρ-coumaric acid, and quercetin-3-rhamnoside, but increase the concentrations of ferulic acid, quercetin-3-galactoside, quercetin-3-glucoside, quercetin-3-rutinoside, and cyanidin-3-galactoside. Furthermore, high AH samples had higher concentrations of chlorogenic acid, ferulic acid, epicatechin, quercetin-3-rutinoside, and cyanidin-3-galactoside than low AH samples as they were in the similar temperature conditions. These results suggested that high AH conditions could significantly improve coloration and the eating quality of fruits. [ABSTRACT FROM AUTHOR]