MdWER interacts with MdERF109 and MdJAZ2 to mediate methyl jasmonate- and light-induced anthocyanin biosynthesis in apple fruit.

Anthocyanin generation in apples (Malus domestica) and the pigmentation that results from it may be caused by irradiation and through administration of methyl jasmonate (MeJA). However, their regulatory interrelationships associated with fruit coloration are not well defined. To determine whether MdERF109, a transcription factor (TF) involved in light-mediated coloration and anthocyanin biosynthesis, has synergistic effects with other proteins, we performed a yeast two-hybrid assessment and identified another TF, MdWER. MdWER was induced by MeJA treatment, and although overexpression of MdWER alone did not promote anthocyanin accumulation co-overexpression with MdERF109 resulted in significantly increase in anthocyanin biosynthesis. MdWER may form a protein complex with MdERF109 to promote anthocyanin accumulation by enhancing combinations between the proteins and their corresponding genes. In addition, MdWER, as a MeJA responsive protein, interacts with the anthocyanin repressor MdJAZ2. Transient co-expression in apple fruit and protein interaction assays allowed us to conclude that MdERF109 and MdJAZ2 interact with MdWER and take part in the production of anthocyanins upon MeJA treatment and irradiation. Our findings validate a role for the MdERF109-MdWER-MdJAZ2 module in anthocyanin biosynthesis and uncover a novel mechanism for how light and MeJA signals are coordinated anthocyanin biosynthesis in apple fruit.

Systematic identification of long noncoding RNAs expressed during light-induced anthocyanin accumulation in apple fruit.

Anthocyanin pigments contribute to the red color of apple (Malus × domestica) fruit and have a major influence on their ornamental, dietary and market value. In this study, we investigated the potential role of long noncoding RNAs (lncRNAs) in anthocyanin biosynthesis. RNA-seq analysis of apple peels from the 'Red Fuji' cultivar during light-induced rapid anthocyanin accumulation revealed 5297 putative lncRNAs. Differential expression analysis further showed that lncRNAs were induced during light treatment and were involved in photosynthesis. Using the miRNA-lncRNA-mRNA network and endogenous target mimic (eTM) analysis, we predicted that two differentially expressed lncRNAs, MLNC3.2 and MLNC4.6, were potential eTMs for miRNA156a and promoted the expression of the SPL2-like and SPL33 transcription factors. Transient expression in apple fruit and stable transformation of apple callus showed that overexpression of the eTMs and SPLs promoted anthocyanin accumulation, with the opposite results in eTM and SPL-silenced fruit. Silencing or overexpressing of miR156a also affected the expression of the identified eTMs and SPLs. These results indicated that MLNC3.2 and MLNC4.6 function as eTMs for miR156a and prevent cleavage of SPL2-like and SPL33 by miR156a during light-induced anthocyanin biosynthesis. Our study provides fundamental insights into lncRNA involvement in the anthocyanin biosynthetic pathway in apple fruit.

Application of a Krypton-Chlorine Excilamp To Control Alicyclobacillus acidoterrestris Spores in Apple Juice and Identification of Its Sporicidal Mechanism.

The aim of this study was to investigate the sporicidal effect of a krypton-chlorine (KrCl) excilamp against Alicyclobacillus acidoterrestris spores and to compare its inactivation mechanism to that of a conventional UV lamp containing mercury (Hg). The inactivation effect of the KrCl excilamp was not significantly different from that of the Hg UV lamp for A. acidoterrestris spores in apple juice despite the 222-nm wavelength of the KrCl excilamp having a higher absorption coefficient in apple juice than the 254-nm wavelength of the Hg UV lamp; this is because KrCl excilamps have a fundamentally greater inactivation effect than Hg UV lamps, which is confirmed under ideal conditions (phosphate-buffered saline). The inactivation mechanism analysis revealed that the DNA damage induced by the KrCl excilamp was not significantly different (P > 0.05) from that induced by the Hg UV lamp, while the KrCl excilamp caused significantly higher (P < 0.05) lipid peroxidation incidence and permeability change in the inner membrane of A. acidoterrestris spores than did the Hg UV lamp. Meanwhile, the KrCl excilamp did not generate significant (P > 0.05) intracellular reactive oxygen species, indicating that the KrCl excilamp causes damage only through the direct absorption of UV light. In addition, after KrCl excilamp treatment with a dose of 2,011 mJ/cm2 to reduce A. acidoterrestris spores in apple juice by 5 logs, there were no significant (P > 0.05) changes in quality parameters such as color (L*, a*, and b*), total phenolic compounds, and DPPH (2,2-diphenyl-1-picrylhydrazyl) free radical scavenging activity.IMPORTANCEAlicyclobacillus acidoterrestris spores, which have high resistance to thermal treatment and can germinate even at low pH, are very troublesome in the juice industry. UV technology, a nonthermal treatment, can be an excellent means to control heat-resistant A. acidoterrestris spores in place of thermal treatment. However, the traditionally applied UV sources are lamps that contain mercury (Hg), which is harmful to humans and the environment; thus, there is a need to apply novel UV technology without the use of Hg. In response to this issue, excilamps, an Hg-free UV source, have been actively studied. However, no studies have been conducted applying this technique to control A. acidoterrestris spores. Therefore, the results of this study, which applied a KrCl excilamp for the control of A. acidoterrestris spores and elucidated the inactivation principle, are expected to be utilized as important basic data for application to actual industry or conducting further studies.

Smartphone Application-Enabled Apple Fruit Surface Temperature Monitoring Tool for In-Field and Real-Time Sunburn Susceptibility Prediction.

Heat stress and resulting sunburn is a major abiotic stress in perineal specialty crops. For example, such stress to the maturing fruits on apple tree canopies can cause several physiological disorders that result in considerable crop losses and reduced marketability of the produce. Thus, there is a critical technological need to effectively monitor the abiotic stress under field conditions for timely actuation of remedial measures. Fruit surface temperature (FST) is one of the stress indicators that can reliably be used to predict apple fruit sunburn susceptibility. This study was therefore focused on development and in-field testing of a mobile FST monitoring tool that can be used for real-time crop stress monitoring. The tool integrates a smartphone connected thermal-Red-Green-Blue (RGB) imaging sensor and a custom developed application ('AppSense 1.0') for apple fruit sunburn prediction. This tool is configured to acquire and analyze imagery data onboard the smartphone to estimate FST. The tool also utilizes geolocation-specific weather data to estimate weather-based FST using an energy balance modeling approach. The 'AppSense 1.0' application, developed to work in the Android operating system, allows visual display, annotation and real-time sharing of the imagery, weather data and pertinent FST estimates. The developed tool was evaluated in orchard conditions during the 2019 crop production season on the Gala, Fuji, Red delicious and Honeycrisp apple cultivars. Overall, results showed no significant difference (t110 = 0.51, p = 0.6) between the mobile FST monitoring tool outputs, and ground truth FST data collected using a thermal probe which had accuracy of ±0.4 °C. Upon further refinements, such tool could aid growers in real-time apple fruit sunburn susceptibility prediction and assist in more effective actuation of apple fruit sunburn preventative measures. This tool also has the potential to be customized for in-field monitoring of the heat stressors in some of the sun-exposed perennial and annual specialty crops at produce maturation.

Transcription profiles reveal the regulatory mechanisms of spur bud changes and flower induction in response to shoot bending in apple (Malus domestica Borkh.).

KEY MESSAGE: Shoot bending, as an effective agronomic measure, has been widely used to promote flowering in 'Fuji' apple trees. Here, we examined the transcriptional responses of genes in 'Fuji' apple buds at different flowering stages under a shoot-bending treatment using RNA sequencing. A complex genetic crosstalk-regulated network, involving abscisic acid-related genes, starch metabolism and circadian rhythm-related genes, as well as stress response-related genes, was up-regulated by shoot bending, in which were contrbuted to apple flower bud formation in response to shoot-bending conditions. Flower induction plays an important role in the apple tree life cycle, but young trees produce fewer and inferior flower buds. Shoot bending, as an effective agronomic measure, has been widely used to promote flowering in 'Fuji' apple trees. However, little is known about the gene expression network patterns and molecular regulatory mechanisms caused by shoot bending during the induced flowering. Here, we examined the transcriptional responses of genes in 'Fuji' apple buds at different flowering stages under a shoot-bending treatment using RNA sequencing. A steady up-regulation of carbon metabolism-related genes led to relatively high levels of sucrose in early induced flowering stages and starch accumulation during shoot bending. Additionally, global gene expression profiling determined that cytokinin, indole-3-acetic acid, gibberellin synthesis and signalling-related genes were significantly regulated by shoot bending, contributing to cell division and differentiation, bud growth and flower induction. A complex genetic crosstalk-regulated network, involving abscisic acid-related genes, starch metabolism- and circadian rhythm-related genes, as well as stress response-related genes, was up-regulated by shoot bending. Additionally, some transcription factor family genes that were involved in sugar, abscisic acid and stress response signalling were significantly induced by shoot bending. These important flowering genes, which were mainly involved in photoperiod, age and autonomous pathways, were up-regulated by shoot bending. Thus, a complex genetic network of regulatory mechanisms involved in sugar, hormone and stress response signalling pathways may mediate the induction of apple tree flowering in response to shoot-bending conditions.

MdCOL4 Interaction Mediates Crosstalk Between UV-B and High Temperature to Control Fruit Coloration in Apple.

In many plants, anthocyanin biosynthesis is affected by environmental conditions. Ultraviolet-B (UV-B) radiation promotes anthocyanin accumulation and fruit coloration in apple skin, whereas high temperature suppresses these processes. In this study, we characterized a B-box transcription factor, MdCOL4, from 'Fuji' apple, and identified its role in anthocyanin biosynthesis by overexpressing its encoding gene in apple red callus. The expression of MdCOL4 was reduced by UV-B, but promoted by high temperature. We explored the regulatory relationship between heat shock transcription factors (HSFs) and MdCOL4, and found that MdHSF3b and MdHSF4a directly bound to the heat shock element cis-element of the MdCOL4 promoter. MdCOL4 interacted with MdHY5 to synergistically inhibit the expression of MdMYB1, and MdCOL4 directly bound to the promoters of MdANS and MdUFGT, which encode genes in the anthocyanin biosynthetic pathway, to suppress their expression. Our findings shed light on the molecular mechanism by which MdCOL4 suppresses anthocyanin accumulation in apple skin under UV-B and high temperature.

A feedback loop involving MdMYB108L and MdHY5 controls apple cold tolerance.

The MYB transcription factors are important for many aspects of plant stress responses. In this study, we isolated and identified an apple MYB gene, MdMYB108L, whose expression is induced by light and cold stresses. An analysis of MdMYB108L-overexpressing transgenic apple calli revealed that MdMYB108L enhances cold tolerance in apple by upregulating MdCBF3 expression. Interestingly, the expression of MdHY5, which encodes an integrator of light and cold signals, was significantly downregulated in transgenic calli. Yeast one-hybrid and electrophoretic mobility shift assays indicated that MdMYB108L positively regulates cold tolerance by binding to the MdCBF3 promoter. Additionally, MdHY5 functions upstream of MdMYB108L, and the resulting increase in MdMYB108L abundance downregulates MdHY5 transcription. The results of this study elucidate a new pathway for the regulation of apple cold tolerance via a feedback mechanism involving MdMYB108L and MdHY5.

The B-box zinc finger protein MdBBX20 integrates anthocyanin accumulation in response to ultraviolet radiation and low temperature.

Ultraviolet-B (UV-B) radiation and low temperature promote the accumulation of anthocyanins, which give apple skins their red colour. Although many transcription regulators have been characterized in the UV-B and low-temperature pathways, their interregulation and synergistic effects are not well understood. Here, a B-box transcription factor gene, MdBBX20, was characterized in apple and identified to promote anthocyanin biosynthesis under UV-B conditions in field experiments and when overexpressed in transgenic apple calli. The transcript level of MdBBX20 was significantly induced by UV-B. Specific G-box elements in the promoters of target genes were identified as interaction sites for MdBBX20. Further experimental interrogation of the UV-B signalling pathways showed that MdBBX20 could interact with MdHY5 in vitro and in vivo and that this interaction was required to significantly enhance the promoter activity of MdMYB1. MdBBX20 also responded to low temperature (14°C) with the participation of MdbHLH3, which directly bound a low temperature-response cis elements in the MdBBX20 promoter. These findings demonstrate the molecular mechanism by which MdBBX20 integrates low-temperature- and UV-B-induced anthocyanin accumulation in apple skin.

Analysis of apple beverages treated with high-power ultrasound: a quality function deployment approach.

BACKGROUND: The objective of this paper was to demonstrate application of quality function deployment in analysing effects of high power ultrasound on quality properties of apple juices and nectars. In order to develop a quality function deployment model, joint with instrumental analysis of treated samples, a field survey was performed to identify consumer preferences towards quality characteristics of juices/nectar. RESULTS: Based on field research, the three most important characteristics were 'taste' and 'aroma' with 28.5% of relative absolute weight importance, followed by 'odour' (16.9%). The quality function deployment model showed that the top three 'quality scores' for apple juice were treatments with amplitude 90 µm, 9 min treatment time and sample temperature 40 °C; 60 µm, 9 min, 60 °C; and 90 µm, 6 min, 40 °C. For nectars, the top three were treatments 120 µm, 9 min, 20 °C; 60 µm, 9 min, 60 °C; and A2.16 60 µm, 9 min, 20 °C. CONCLUSION: This type of quality model enables a more complex measure of large scale of different quality parameters. Its simplicity should be understood as its practical advantage and, as such, this tool can be a part of design quality when using novel preservation technologies. © 2017 Society of Chemical Industry.

A robust impact assessment that informs actionable climate change adaptation: future sunburn browning risk in apple.

Climate change impact assessments are predominantly undertaken for the purpose of informing future adaptation decisions. Often, the complexity of the methodology hinders the actionable outcomes. The approach used here illustrates the importance of considering uncertainty in future climate projections, at the same time providing robust and simple to interpret information for decision-makers. By quantifying current and future exposure of Royal Gala apple to damaging temperature extremes across ten important pome fruit-growing locations in Australia, differences in impact to ripening fruit are highlighted, with, by the end of the twenty-first century, some locations maintaining no sunburn browning risk, while others potentially experiencing the risk for the majority of the January ripening period. Installation of over-tree netting can reduce the impact of sunburn browning. The benefits from employing this management option varied across the ten study locations. The two approaches explored to assist decision-makers assess this information (a) using sunburn browning risk analogues and (b) through identifying hypothetical sunburn browning risk thresholds, resulted in varying recommendations for introducing over-tree netting. These recommendations were location and future time period dependent with some sites showing no benefit for sunburn protection from nets even by the end of the twenty-first century and others already deriving benefits from employing this adaptation option. Potential best and worst cases of sunburn browning risk and its potential reduction through introduction of over-tree nets were explored. The range of results presented highlights the importance of addressing uncertainty in climate projections that result from different global climate models and possible future emission pathways.

Epigenetic regulation of MdMYB1 is associated with paper bagging-induced red pigmentation of apples.

MAIN CONCLUSION: Paper-bagging treatment can transform non-transcribed MdMYB1 - 2 and MdMYB1 - 3 alleles into transcribed alleles through epigenetic regulations, resulting in the red pigmentation of a normally non-red apple cultivar 'Mutsu.' Anthocyanin biosynthesis in apples is regulated by MdMYB1/A/10, an R2R3-Type MYB gene. 'Mutsu,' a triploid apple cultivar harboring non-transcribed MdMYB1-2 and MdMYB1-3 alleles, retains green skin color under field conditions. However, it can show red/pink pigmentation under natural or artificial ultraviolet-B (UV-B) light exposure after paper-bagging and bag removal treatment. In the present study, we found that in 'Mutsu,' paper bagging-induced red pigmentation was due to the activation of non-transcribed MdMYB1-2/-3 alleles, which triggered the expression of downstream anthocyanin biosynthesis genes in a UV-B-dependent manner. By monitoring the epigenetic changes during UV-B-induced pigmentation, no significant differences in DNA methylation and histone modifications in the 5' upstream region of MdMYB1-2/-3 were recorded between the UV-B-treated fruit skin (red) and the fruit skin treated only by white light (green). In contrast, bag treatment lowered the DNA methylation in this region of MdMYB1-2/-3 alleles. Similarly, higher levels of histone H3 acetylation and trimethylation of H3 tail at lysine 4, and lower level of trimethylation of H3 tail at lysine 27 were observed in the 5' upstream region of MdMYB1-2/-3 in the skin of the fruit immediately after bag removal. These results suggest that bagging treatment can induce epigenetic changes, facilitating the binding of trans factor(s) to MdMYB1-2/-3 alleles, resulting in the activation of these MYBs after bag removal.

Photoprotection mechanism in the 'Fuji' apple peel at different levels of photooxidative sunburn.

The xanthophyll cycle, flavonoid metabolism, the antioxidant system and the production of active oxygen species were analyzed in the peel of 'Fuji' apples re-exposed to sunlight after extended periods of fruit bagging treatment, resulting in different levels of photooxidative sunburn. After re-exposing bagged fruits to sunlight, the production of active oxygen species and the photoprotective capacity in apple peels were both significantly enhanced. As sunburn severity increased, the concentration of hydrogen peroxide increased, while xanthophyll cycle pool size decreased. For the key genes involved in flavonoid synthesis, expressions of MdMYB10 and MdPAL were upregulated, whereas the expressions of MdCHS, MdANS, MdFLS and MdUFGT were downregulated in sunburnt fruit peel. Correspondingly, concentrations of both quercetin-3-glycoside and cyanidin-3-galactoside decreased. Total ascorbate concentrations decreased as sunburn severity increased, with the decrease being faster for oxidized than for reduced ascorbate. Transcription levels of MdGMP, MdGME, MdGGP, MdGPP, MdGalDH and MdGalLDH, the genes involved in ascorbate synthesis, were similar in non-sunburnt and sunburnt fruit peels, whereas activities of l-galactose dehydrogenase and l-galactono-1,4-lactone dehydrogenase decreased in severely sunburnt peel. Although activities of superoxide dismutase and ascorbate peroxidase increased, the activities of monodehydroascorbate reductase, dehydroascorbate reductase and glutathione reductase decreased as sunburn severity increased. In summary, the occurrence of photooxidative sunburn in 'Fuji' apple peel is closely associated with a relatively lower xanthophyll cycle pool size, reduced levels of ascorbate reduction and synthesis and reduced flavonoid synthesis. Our data are consistent with the idea that ascorbate plays a key role in protecting apple fruit from photooxidative sunburn.

Inactivation and potential reactivation of pathogenic Escherichia coli O157:H7 in apple juice following ultraviolet light exposure at three monochromatic wavelengths.

Ultraviolet (UV) light irradiation at 254 nm is considered as a novel non-thermal method for decontamination of foodborne pathogenic bacteria. However, lower penetration depth of UV light at 254 nm in apple juice resulted in higher UV dose consumption during apple juice decontamination. In addition, no studies are available on the reactivation of pathogens following exposure to UV light in drinks and beverages. Two novel monochromatic UV light sources (λ = 222 and 282 nm) have been developed for bacterial disinfection. However, the inactivation of pathogenic Escherichia coli O157:H7 following exposure to these UV wavelengths is still unclear. Therefore, the present study was conducted to determine the inactivation and reactivation potential of pathogenic E. coli O157:H7 in apple juice following exposure to UV light at three monochromatic wavelengths: Far UV (λ = 222 nm), Far UV+ (λ = 282 nm) and UVC light (λ = 254 nm). The results showed that E. coli O157:H7 is acid-resistant, and up to 99.50% of cells survived in apple juice when incubated at 20 °C for 24 h. Inactivation of E. coli O157:H7 following exposure to Far UV light (2.81 Log reduction) was higher (P < 0.05) than the inactivation caused by UVC light (1.95 Log reduction) and Far UV+ light (1.83 Log reduction) at the similar levels of UV fluence of 75 mJ/cm(2). No any reactivation potential was observed for E. coli O157:H7 in dark incubation phases after exposure to UV light as determined by the regular plating method. In addition, the exposure to Far UV light at 222 nm followed by incubating at 37 °C significantly decreased (P < 0.05) the survival of E. coli O157:H7 during dark incubation phase compared to that of UVC and Far UV+ light.

Influence of the variation of geometrical and topological traits on light interception efficiency of apple trees: sensitivity analysis and metamodelling for ideotype definition.

BACKGROUND AND AIMS: The impact of a fruit tree's architecture on its performance is still under debate, especially with regard to the definition of varietal ideotypes and the selection of architectural traits in breeding programmes. This study aimed at providing proof that a modelling approach can contribute to this debate, by using in silico exploration of different combinations of traits and their consequences on light interception, here considered as one of the key parameters to optimize fruit tree production. METHODS: The variability of organ geometrical traits, previously described in a bi-parental population, was used to simulate 1- to 5-year-old apple trees (Malus × domestica). Branching sequences along trunks observed during the first year of growth of the same hybrid trees were used to initiate the simulations, and hidden semi-Markov chains previously parameterized were used in subsequent years. Tree total leaf area (TLA) and silhouette to total area ratio (STAR) values were estimated, and a sensitivity analysis was performed, based on a metamodelling approach and a generalized additive model (GAM), to analyse the relative impact of organ geometry and lateral shoot types on STAR. KEY RESULTS: A larger increase over years in TLA mean and variance was generated by varying branching along trunks than by varying organ geometry, whereas the inverse was observed for STAR, where mean values stabilized from year 3 to year 5. The internode length and leaf area had the highest impact on STAR, whereas long sylleptic shoots had a more significant effect than proleptic shoots. Although the GAM did not account for interactions, the additive effects of the geometrical factors explained >90% of STAR variation, but much less in the case of branching factors. CONCLUSIONS: This study demonstrates that the proposed modelling approach could contribute to screening architectural traits and their relative impact on tree performance, here viewed through light interception. Even though trait combinations and antagonism will need further investigation, the approach opens up new perspectives for breeding and genetic selection to be assisted by varietal ideotype definition.

MdCOP1 ubiquitin E3 ligases interact with MdMYB1 to regulate light-induced anthocyanin biosynthesis and red fruit coloration in apple.

MdMYB1 is a crucial regulator of light-induced anthocyanin biosynthesis and fruit coloration in apple (Malus domestica). In this study, it was found that MdMYB1 protein accumulated in the light but degraded via a ubiquitin-dependent pathway in the dark. Subsequently, the MdCOP1-1 and MdCOP1-2 genes were isolated from apple fruit peel and were functionally characterized in the Arabidopsis (Arabidopsis thaliana) cop1-4 mutant. Yeast (Saccharomyces cerevisiae) two-hybrid, bimolecular fluorescence complementation, and coimmunoprecipitation assays showed that MdMYB1 interacts with the MdCOP1 proteins. Furthermore, in vitro and in vivo experiments indicated that MdCOP1s are necessary for the ubiquitination and degradation of MdMYB1 protein in the dark and are therefore involved in the light-controlled stability of the MdMYB1 protein. Finally, a viral vector-based transformation approach demonstrated that MdCOP1s negatively regulate the peel coloration of apple fruits by modulating the degradation of the MdMYB1 protein. Our findings provide new insight into the mechanism by which light controls anthocyanin accumulation and red fruit coloration in apple and even other plant species.

The role of anthocyanin in photoprotection and its relationship with the xanthophyll cycle and the antioxidant system in apple peel depends on the light conditions.

The synthesis of anthocyanin, the xanthophyll cycle, the antioxidant system and the production of active oxygen species (AOS) were compared between red and non-red apple cultivars, in response to either long-term sunlight exposure (high light intensity) during fruit development, or to exposure of bagged fruits to lower light intensity late in fruit development. During fruit development of red and non-red apples, the xanthophyll cycle pool size decreased much more in red apple peel late in development. With accumulation of AOS induced by long-term sunlight exposure, enhancement of the antioxidant system was found. However, this change became significantly lower in red apple than non-red apple as fruit developed, which might serve to accelerate the anthocyanin synthesis in red apple peel. When, late in fruit development, bagged fruits were exposed to sunlight, the accumulation of AOS was lower in red apple peel than in non-red peel. This could be due to the higher anthocyanin concentration in the red peels. Meanwhile, compared with that in non-red cultivar, the xanthophyll cycle and the antioxidant system in red apple peel were protected first but then down-regulated by its higher anthocyanin concentration during sunlight exposure. In conclusions, red and non-red apples peel possess different photoprotective mechanisms under high light conditions. The relationship between anthocyanin synthesis and the xanthophyll cycle, and the antioxidant system, depends on the light conditions that fruit undergoes.

Screening of UV-B-induced genes from apple peels by SSH: possible involvement of MdCOP1-mediated signaling cascade genes in anthocyanin accumulation.

Suppression subtractive hybridization (SSH) was employed to identify candidate genes involved in red coloration in apple peel with the ultraviolet (UV)-B-treated 'Mutsu'. After reverse Northern blotting verification, nearly 80 clones were successfully sequenced. Large portions of the expressed sequence tags (ESTs) are well characterized anthocyanin biosynthesis-related genes, such as chalcone synthase (11A5), flavonol synthase (12F3), anthocyanidin synthase (11H5) and UDP-glycosyl transferase (14A12) whose presence proved the success of SSH. Eight ESTs were selected for quantitative real-time polymerase chain reaction analysis and their expressions were all elevated in 'Induction', further confirming the reliability of the SSH library. One EST, 11F4 (CONSTITUTIVE PHOTOMORPHOGENIC 1: COP1) with putative function in light signal relay was further analyzed in 'Mutsu' and 'Tsugaru', along with MdHY5 (ELONGATED HYPOCOTYL 5: the downstream target of COP1), MdMYB22 (a possible flavonol-specific activator under the regulation of HY5, belonging to the SG7/PRODUCTION OF FLAVONOL GLYCOSIDES family) and MdMYBA. Results showed that MdCOP1, MdHY5, MdMYB22 and MdMYBA were all UV-B inducible genes and anthocyanin accumulation occurred after their increased expressions. Moreover, their expressions and anthocyanin content were enhanced under UV-B plus 17°C treatment. The presence of G box, a known consensus binding site of HY5, in the MdMYBA promoter region implicated that it could be regulated by MdHY5, which was verified by the result of the yeast one-hybrid analysis. Our data suggested that UV-B irradiation would induce the utmost upstream light signaling factor, MdCOP1, which activates MdHY5 signaling by binding to the promoter regions of MdMYBs, and finally leads to the red coloration of apple peels.

Transcriptomics of shading-induced and NAA-induced abscission in apple (Malus domestica) reveals a shared pathway involving reduced photosynthesis, alterations in carbohydrate transport and signaling and hormone crosstalk.

BACKGROUND: Naphthaleneacetic acid (NAA), a synthetic auxin analogue, is widely used as an effective thinner in apple orchards. When applied shortly after fruit set, some fruit abscise leading to improved fruit size and quality. However, the thinning results of NAA are inconsistent and difficult to predict, sometimes leading to excess fruit drop or insufficient thinning which are costly to growers. This unpredictability reflects our incomplete understanding of the mode of action of NAA in promoting fruit abscission. RESULTS: Here we compared NAA-induced fruit drop with that caused by shading via gene expression profiling performed on the fruit abscission zone (FAZ), sampled 1, 3, and 5 d after treatment. More than 700 genes with significant changes in transcript abundance were identified from NAA-treated FAZ. Combining results from both treatments, we found that genes associated with photosynthesis, cell cycle and membrane/cellular trafficking were downregulated. On the other hand, there was up-regulation of genes related to ABA, ethylene biosynthesis and signaling, cell wall degradation and programmed cell death. While the differentially expressed gene sets for NAA and shading treatments shared only 25% identity, NAA and shading showed substantial similarity with respect to the classes of genes identified. Specifically, photosynthesis, carbon utilization, ABA and ethylene pathways were affected in both NAA- and shading-induced young fruit abscission. Moreover, we found that NAA, similar to shading, directly interfered with leaf photosynthesis by repressing photosystem II (PSII) efficiency within 10 minutes of treatment, suggesting that NAA and shading induced some of the same early responses due to reduced photosynthesis, which concurred with changes in hormone signaling pathways and triggered fruit abscission. CONCLUSIONS: This study provides an extensive transcriptome study and a good platform for further investigation of possible regulatory genes involved in the induction of young fruit abscission in apple, which will enable us to better understand the mechanism of fruit thinning and facilitate the selection of potential chemicals for the thinning programs in apple.

Reduction of patulin in apple cider by UV radiation.

The presence of the mycotoxin patulin in processed apple juice and cider presents a continual challenge to the food industry as both consumer health and product quality issues. Although several methods for control and/or elimination of patulin have been proposed, no unifying method has been commercially successful for reducing patulin burdens while maintaining product quality. In the present study, exposure to germicidal UV radiation was evaluated as a possible commercially viable alternative for the reduction and possible elimination of the patulin mycotoxin in fresh apple cider. UV exposure of 14.2 to 99.4 mJ/cm(2) resulted in a significant and nearly linear decrease in patulin levels while producing no quantifiable changes in the chemical composition (i.e., pH, Brix, and total acids) or organoleptic properties of the cider. For the range of UV doses tested, patulin levels decreased by 9.4 to 43.4%; the greatest reduction was achieved after less than 15 s of UV exposure. The method of UV radiation (the CiderSure 3500 system) is an easily implemented, high-throughput, and cost-effective method that offers simultaneous UV pasteurization of cider and juice products and reduction and/or elimination of patulin without unwanted alterations in the final product.