Phenomics and transcriptomic profiling of fruit development in distinct apple varieties.

Apple is one of the most economically important and popular temperate fruit trees. The domestication of apple has resulted in substantial phenotypic differences, particularly between wild and cultivated varieties. However, the relationship between gene expression and phenotypic variations in apple remains poorly understood. Here, we present a comprehensive dataset featuring five distinct apple varieties, including two wild varieties and three representative cultivated varieties. The dataset comprises of both phenomics data, encompassing twelve fruit quality-related traits continuously measured over two years, and transcriptomic data obtained at different developmental stages with three biological replicates. We performed basic quality control process, gene expression normalization and differential gene expression analysis to demonstrate the utility and reliability of the dataset. Our findings indicate that gene expression strongly related with phenotypic variations in apple. This dataset serves as a valuable resource, encompassing phenomics and transcriptomic data in multiple formats, thereby facilitating further exploration of the relationships between gene expression and phenotypic traits in apple.

Epigenomic mechanism regulating the quality and ripeness of apple fruit with differing harvest maturity.

Harvest maturity significantly affects the quality of apple fruit in post-harvest storage process. Although the regulatory mechanisms underlying fruit ripening have been studied, the associated epigenetic modifications remain unclear. Thus, we compared the DNA methylation changes and the transcriptional responses of mature fruit (MF) and immature fruit (NF). There were significant correlations between DNA methylation and gene expression. Moreover, the sugar contents (sucrose, glucose, and fructose) were higher in MF than in NF, whereas the opposite pattern was detected for the starch content. The expression-level differences were due to DNA methylations and ultimately resulted in diverse fruit textures and ripeness. Furthermore, the higher ethylene, auxin, and abscisic acid levels in MF than in NF, which influenced the fruit texture and ripening, were associated with multiple differentially expressed genes in hormone synthesis, signaling, and response pathways (ACS, ACO, ZEP, NCED, and ABA2) that were regulated by DNA methylations. Multiple transcription factor genes involved in regulating fruit ripening and quality via changes in DNA methylation were identified, including MIKCC-type MADS-box genes and fruit ripening-related genes (NAP, SPL, WRKY, and NAC genes). These findings reflect the diversity in the epigenetic regulation of gene expression and may be relevant for elucidating the epigenetic regulatory mechanism underlying the ripening and quality of apple fruit with differing harvest maturity.

Pigmentation and Flavonoid Metabolite Diversity in Immature 'Fuji' Apple Fruits in Response to Lights and Methyl Jasmonate.

Artificial pigmentation of apple fruits has been intensely evaluated to generate less pigmented red apples, which are profitable because of the changes in fruit quality. In this study, we analyzed the diversity of flavonoids and the patterns of flavonoid metabolic gene expression under light irradiation with or without methyl jasmonate (MeJA) treatment in immature (S1) and color-turning (S2) staged 'Fuji' apples. Further, we assessed the metabolic regulation at the gene level between anthocyanin and flavonol in light-responsive apple skins. UV-B exposure within 3 days was found to significantly stimulate anthocyanin accumulation in apple skin compared to other light exposure. S1 skin was more sensitive to UV-B and MeJA treatment, in the aspect of indaein accumulation. The enhancement of apple pigmentation following treatment with adequate levels of UV-B and MeJA was maximized at approximately 72 h. Red (range from 4.25 to 17.96 µg·g-1 DW), blue (range from 4.59 to 9.17 µg·g-1 DW) and UV-A (range from 3.98 to 19.12 µg·g-1 DW) lights contributed to the induction of idaein content. Most genes related to the flavonoid pathways increased their expression under UV-B exposure, including the gene expression of the transcription factor, MdMYB10, a well-known upstream factor of flavonoid biosynthesis in apples. The boosted upregulation of MdMYB10, MdCHS, MdF3H MdLDOX, and MdUFGT genes due to MeJA in UV-B was found and may contribute the increase of idaein. UV-A and UV-B caused higher quercetin glycoside content in both S1 and S2 apple skins than longer wavelengths, resulting in significant increases in quercetin-3-O-galactoside and quercetin-3-O-glucoside. These results suggest that the application of adequate UV-B with MeJA in less-pigmented postharvest apples will improve apple color quality within a short period.

Functional Characterization of MdTAC1a Gene Related to Branch Angle in Apple (Malus x domestica Borkh.).

The Tiller Angle Control 1 (TAC1) gene belongs to the IGT family, which mainly controls plant branch angle, thereby affecting plant form. Two members of MdTAC1 are identified in apple; the regulation of apple branch angle by MdTAC1 is still unclear. In this study, a subcellular localization analysis detected MdTAC1a in the nucleus and cell membrane, but MdTAC1b was detected in the cell membrane. Transgenic tobacco by overexpression of MdTAC1a or MdTAC1b showed enlarged leaf angles, the upregulation of several genes, such as GA 2-oxidase (GA2ox), and a sensitive response to light and gravity. According to a qRT-PCR analysis, MdTAC1a and MdTAC1b were strongly expressed in shoot tips and vegetative buds of weeping cultivars but were weakly expressed in columnar cultivars. In the MdTAC1a promoter, there were losses of 2 bp in spur cultivars and 6 bp in weeping cultivar compared with standard and columnar cultivars. An InDel marker specific to the MdTAC1a promoter was developed to distinguish apple cultivars and F1 progeny. We identified a protein, MdSRC2, that interacts with MdTAC1a, whose encoding gene which was highly expressed in trees with large branch angles. Our results indicate that differences in the MdTAC1a promoter are major contributors to branch-angle variation in apple, and the MdTAC1a interacts with MdSRC2 to affect this trait.

Overexpression of MdZAT5, an C2H2-Type Zinc Finger Protein, Regulates Anthocyanin Accumulation and Salt Stress Response in Apple Calli and Arabidopsis.

Zinc finger proteins are widely involved and play an important role in plant growth and abiotic stress. In this research, MdZAT5, a gene encoding C2H2-type zinc finger protein, was cloned and investigated. The MdZAT5 was highly expressed in flower tissues by qRT-PCR analyses and GUS staining. Promoter analysis showed that MdZAT5 contained multiple response elements, and the expression levels of MdZAT5 were induced by various abiotic stress treatments. Overexpression of MdZAT5 in apple calli positively regulated anthocyanin accumulation by activating the expressions of anthocyanin biosynthesis-related genes. Overexpression of MdZAT5 in Arabidopsis also enhanced the accumulation of anthocyanin. In addition, MdZAT5 increased the sensitivity to salt stress in apple calli. Ectopic expression of MdZAT5 in Arabidopsis reduced the expression of salt-stress-related genes (AtNHX1 and AtABI1) and improved the sensitivity to salt stress. In conclusion, these results suggest that MdZAT5 plays a positive regulatory role in anthocyanin accumulation and negatively regulates salt resistance.

Cyclanilide Induces Lateral Bud Outgrowth by Modulating Cytokinin Biosynthesis and Signalling Pathways in Apple Identified via Transcriptome Analysis.

Cyclanilide (CYC), a plant growth regulator, is a potent shoot branching agent in apple. However, its mechanism remains unclear. The current study revealed that CYC treatment resulted in massive reprogramming of the axillary bud transcriptome, implicating several hormones in the response. We observed a marked increase (approximately 2-fold) in the level of zeatin riboside and a significant decrease (approximately 2-fold) in the level of abscisic acid (ABA). Zeatin metabolism gene cytokinin (CTK) oxidase 1 (CKX 1) was down-regulated at 168 h after CYC treatment compared with the control. Weighted gene co-expression network analysis of differentially expressed genes demonstrated the turquoise module clusters exhibited the highest positive correlation with zeatin riboside (r = 0.92) and the highest negative correlation with ABA (r = -0.8). A total of 37 genes were significantly enriched in the plant hormone signal transduction pathway in the turquoise module. Among them, the expressions of CTK receptor genes WOODEN LEG and the CTK type-A response regulators genes ARR3 and ARR9 were up-regulated. ABA signal response genes protein phosphatase 2C genes ABI2 and ABI5 were down-regulated in lateral buds after CYC treatment at 168 h. In addition, exogenous application of 6-benzylaminopurine (6-BA, a synthetic type of CTK) and CYC enhanced the inducing effect of CYC, whereas exogenous application of lovastatin (a synthetic type of inhibitor of CTK biosynthesis) or ABA and CYC weakened the promoting effect of CYC. These results collectively revealed that the stimulation of bud growth by CYC might involve CTK biosynthesis and signalling, including genes CKX1 and ARR3/9, which provided a direction for further study of the branching promoting mechanism of CYC.

Transcriptome Analysis Reveals Multiple Genes and Complex Hormonal-Mediated Interactions with PEG during Adventitious Root Formation in Apple.

Adventitious root (AR) formation is a bottleneck for the mass propagation of apple rootstocks, and water stress severely restricts it. Different hormones and sugar signaling pathways in apple clones determine AR formation under water stress, but these are not entirely understood. To identify them, GL-3 stem cuttings were cultured on polyethylene glycol (PEG) treatment. The AR formation was dramatically decreased compared with the PEG-free control (CK) cuttings by increasing the endogenous contents of abscisic acid (ABA), zeatin riboside (ZR), and methyl jasmonate (JA-me) and reducing the indole-3-acetic acid (IAA) and gibberellic acid 3 (GA3) contents. We performed a transcriptomic analysis to identify the responses behind the phenotype. A total of 3204 differentially expressed genes (DEGs) were identified between CK and PEG, with 1702 upregulated and 1502 downregulated genes. Investigation revealed that approximately 312 DEGs were strongly enriched in hormone signaling, sugar metabolism, root development, and cell cycle-related pathways. Thus, they were selected for their possible involvement in adventitious rooting. However, the higher accumulation of ABA, ZR, and JA-me contents and the upregulation of their related genes, as well as the downregulation of sugar metabolism-related genes, lead to the inhibition of ARs. These results indicate that AR formation is a complicated biological process chiefly influenced by multiple hormonal signaling pathways and sugar metabolism. This is the first study to demonstrate how PEG inhibits AR formation in apple plants.

Intrinsic and extrinsic attributes related to the influence of growing altitude on consumer acceptability and sensory perception of fresh apple.

BACKGROUND: Growing altitude might have an effect on the quality of fresh apple and therefore on the perception of sensory attributes and on acceptance by consumers. On the other hand, extrinsic information provided to the consumer might affect consumer acceptability and sensory perception of the fruit's main quality traits. The main objective of this work was to study the effect of the growing altitude on the physico-chemical attributes (soluble solid content (SSC), flesh firmness, fruit weight (g), and titratable acidity (TA)), consumer acceptability, and perception of the main sensory attributes (sweetness, acidity and texture) of two reference apple cultivars ('Golden D.' and 'Reineta') by using a panel of 195 consumers. A secondary objective was to study whether extrinsic information about cultivar and growing place have an effect on the consumer perception and acceptance of fresh apple. RESULTS: Significant effects on physico-chemical and sensory attributes were found for 'Golden D.' and 'Reineta' apple cultivars due to the growing altitude, and they were perceived by consumers. Moreover, extrinsic information about the cultivar and the growing site influenced consumers' sensory perceptions significantly. CONCLUSIONS: This work demonstrated that differences between apple fruit grown at different altitudes can be perceived by consumers. We demonstrated the importance of providing consumers with information about the cultivar and the growing place to increase their acceptance and support local produce. © 2021 The Authors. Journal of The Science of Food and Agriculture published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Effects of Indole-3-Butyric Acid (IBA) and rooting media on rooting and survival of air layered wax apple (Syzygium samarangense) CV Jambu Madu / Efeitos do Ácido Indol-3-Butírico (AIB) e meio de enraizamento no enraizamento e sobrevivência de maçã de cera em camada de ar (Syzygium samarangense) CV Jambu Madu

The wax apple or jambu madu, is a non-climacteric tropical fruit from Myrtaceae family and widely cultivated in South East Asia. The limited availability of good quality seedlings of wax apple is the main problem to development of flourish it's market share in the current fruit industry. Therefore, in order to produce good quality planting materials, a study aimed at optimizing propagation and adventitious rooting technique and survivability of wax apple air layer was conducted. In this study, four different levels of Indole-3-Butyric Acid (IBA) concentration (0, 1000, 1500 and 2000 mg L-1) and three rooting media (sphagnum moss, vermicompost and garden soil) were applied after removal of bark (phloem) on the shoot to determine the effect on rooting and survivability of the wax apple air layer under field conditions. The results showed that the wax apple shoots treated with 2000 mg L-1 IBA produced the significantly higher number of roots, increased length of root, diameter of branch, length of branch, number of leaf and leaf area of air layers. In addition, the highest chlorophyll content and stomatal aperture were recorded in 2000 mg L-1 IBA treatment compared to other treatments including control. Vermicompost medium was better than garden soil and sphagnum moss in respect of rooting and survivability of air layers. The results showed that the combination of 2000 mg L-1 IBA and vermicompost as rooting media give the best combination to root initiation, root number, root length and survival rate (100%) of wax apple air layers. From this study, it can be concluded that 2000 mg L-1 IBA and vermicompost treatment enhance the root initiation, early establishment and survivability of wax apple air layered under field conditions.

A maçã de cera, ou jambu madu, é uma fruta tropical não climatérica da família Myrtaceae e amplamente cultivada no Sudeste Asiático. A disponibilidade limitada de mudas de macieira de boa qualidade é o principal problema para o desenvolvimento de sua participação de mercado na fruticultura atual. Portanto, com o objetivo de produzir materiais de plantio de boa qualidade, foi realizado um estudo visando otimizar a técnica de propagação e enraizamento adventício e a sobrevivência da camada aérea da cera de macieira. Neste estudo, quatro diferentes níveis de concentração de ácido indol-3-butírico (AIB) (0, 1000, 1500 e 2000 mg L-1) e três meios de enraizamento (musgo esfagno, vermicomposto e solo de jardim) foram aplicados após a remoção da casca (floema) na parte aérea para determinar o efeito no enraizamento e capacidade de sobrevivência da camada de ar da macieira em condições de campo. Os resultados mostraram que os brotos de macieira tratados com 2000 mg L-1 de AIB produziram significativamente maior número de raízes, maior comprimento de raiz, diâmetro de galho, comprimento de galho, número de folhas e área foliar das camadas aéreas. Além disso, o maior teor de clorofila e abertura estomática foram registrados no tratamento 2000 mg L-1 IBA em comparação com outros tratamentos, incluindo o controle. O meio de vermicomposto foi melhor do que o solo de jardim e o musgo esfagno em relação ao enraizamento e capacidade de sobrevivência das camadas aéreas. Os resultados mostraram que a combinação de 2000 mg L-1 de AIB e vermicomposto como meio de enraizamento proporciona a melhor combinação para iniciação radicular, número de raízes, comprimento radicular e taxa de sobrevivência (100%) das camadas aéreas de macieira. A partir deste estudo, pode-se concluir que 2.000 mg L-1 de AIB e tratamento com vermicomposto melhoram a iniciação radicular, o estabelecimento precoce e a capacidade de sobrevivência de macieiras em camadas de ar em condições de campo.

Horticultural performance of 'Fuji Suprema' and 'Maxi Gala' apple trees trained at Tall Spindle and Bi-Axis systems grafted on different rootstocks in Southern Brazil / Comportamento agronômico de macieiras 'Fuji Suprema' e 'Maxi Gala' conduzidas em Tall Spindle e Bi-Axis enxertadas em diferentes porta-enxertos no Sul do Brasil

The use of new rootstocks combined with different training systems have been studied to increase productivity, fruit quality, and reduce costs. Another important factor is the validation of studies under replanting conditions, due to the growing limitation of new areas for planting commercial apple orchards in Brazil. The present study aimed to evaluate the behavior of 'Fuji Suprema' and 'Maxi Gala' apple trees in two training systems grafted on two rootstocks. The study occurred during the seasons 2016/17, 2017/18 and 2018/19, in an orchard located in southern Brazil in an area previously cultivated with apple trees. A randomized block design consisting of a 2x2 factorial was used, with Tall Spindle and Bi-Axis as the training system and 'M.9' and 'G.213' as rootstocks of Fuji Suprema and Maxi Gala cultivars. Variables were evaluated to determine tree vigor, as well as yield and fruit quality characteristics. It was observed that 'G.213' promoted higher vigor for both cultivars as well as the Bi-Axis training system in the Maxi Gala cultivar. The highest yield was observed in Tall Spindle as well as the rootstock 'G.213' in both cultivars. The most significant changes on fruit quality occurred in the Bi-Axis training system, presenting higher intensity of red color as well as 'Maxi Gala' grafted on 'G.213' rootstock.

A utilização de novos porta-enxertos e diferentes sistemas de condução tem sido estudado para aumentar a produtividade, qualidade dos frutos e reduzir custos. Outro fator importante é a validação de estudos em condições de replantio, devido à crescente limitação de novas áreas para o plantio comercial de pomares de macieiras no Brasil. O presente estudo teve como objetivo avaliar o comportamento de macieiras 'Fuji Suprema' e 'Maxi Gala' em dois sistemas de condução enxertados sobre dois porta-enxertos. O estudo ocorreu durante as safras 2016/17, 2017/18 e 2018/19, em um pomar localizado no sul do Brasil, em área previamente cultivada com macieiras. Utilizou-se delineamento de blocos casualizados, consistindo em um fatorial 2x2, sendo Tall Spindle e Bi-Axis como sistemas de condução e 'M.9' e 'G.213' como porta-enxertos das cultivares Fuji Suprema e Maxi Gala. Foram avaliadas variáveis para determinar o comportamento vegetativo, bem como as características de produtividade e qualidade dos frutos. Observou-se que 'G.213' promoveu maior vigor para ambas cultivares, assim como o sistema de condução Bi-Axis na cultivar Maxi Gala. O maior rendimento foi observado nas macieiras conduzidas em Tall Spindle e enxertadas sobre 'G.213' em ambas cultivares. As mudanças mais significativas na qualidade dos frutos ocorreram nas macieiras conduzidas em Bi-Axis, apresentando maior intensidade de coloração vermelha, assim como na macieira 'Maxi Gala' enxertada sobre 'G.213'.

Morphological, sensory and biochemical characteristics of summer apple genotypes / Características morfológicas, sensoriais e bioquímicas de genótipos de maçã cultivada no verão

Summer apples are one of the most important plant community in Artvin province located Northeastern part of Turkey. In present study 22 local apple genotypes were characterized by phenological, morphological, biochemical and sensory properties. Harvest date was the main phenological data. Morphological measurements included fruit weight, fruit shape, fruit ground color, fruit over color, fruit over color coverage and fruit firmness, respectively. Sensory measurements were as juiciness and aroma and biochemical characteristics included organic acids, SSC (Soluble Solid Content), vitamin C, total phenolic content and antioxidant capacity. Genotypes exhibited variable harvest dates ranging from 11 July to 13 August and cv. Summered harvested 30 July 2017. The majority of genotypes were harvested before cv. Summered. Fruit weight were also quite variable among genotypes which found to be between 89 g and 132 g, and most of the genotypes had bigger fruits than cv. Summered. Pink, red, yellow and green fruit skin color was evident and main fruit shape were determined as round, conic and oblate among genotypes. ART08-9, ART08-4, ART08-21 and ART08-22 had distinct bigger fruits and ART08-1, ART08-2, ART08-5, ART08-12 and ART08-17 had higher total phenolic content and antioxidant capacity. The results of the study showed significant differences for most of the phenological, morphological, sensory and biochemical characteristics. Thus, the phonological, morphological, sensory and biochemical characteristics of summer apple genotypes were distinguishable and these results suggest that phonological, morphological, sensory and biochemical differences of the summer apple genotypes can be attributed to differences in genetic background of genotypes which placed different groups by PCoA analysis.

As maçãs cultivadas no verão são uma das culturas vegetais mais importantes da província de Artvin, localizada no nordeste da Turquia. No presente estudo, 22 genótipos locais de maçã foram caracterizados quanto às suas propriedades morfológicas, bioquímicas e sensoriais. As características analisadas foram peso do fruto, data da colheita, forma do fruto, coloração da casca, firmeza do fruto, suculência, aroma, teor de ácidos orgânicos e de sólidos solúveis, vitamina C, teor fenólico total e capacidade antioxidante. Os genótipos exibiram datas de colheita que variaram de 11 de julho a 13 de agosto, e a cultivar de verão foi colhida em 30 de julho 2017. A maioria dos genótipos foi colhida antes da cultivar de verão. O peso dos frutos também foi bastante variável entre os genótipos, apresentando entre 89 e 132 g, e a maioria dos genótipos apresentou frutos maiores que a cultivar de verão. As colorações rosa, vermelho, amarelo e verde da casca dos frutos foram as mais evidentes, e a forma principal dos frutos foi determinada como redonda, cônica e oblata entre os genótipos. ART08-9, ART08-4, ART08-21 e ART08-22apresentaram frutos maiores distintos, e ART08-1, ART08-2, ART08-5, ART08-12 e ART08-17 apresentaram maior conteúdo fenólico total e capacidade antioxidante. Os resultados do estudo mostraram diferenças significativas para a maioria das características morfológicas, sensoriais e bioquímicas. Assim, essas características dos genótipos da maçã cultivadas no verão foram distintos, e esses resultados podem ser atribuídos a diferenças no contexto genético dos genótipos.

Activation of the ABA Signal Pathway Mediated by GABA Improves the Drought Resistance of Apple Seedlings.

Drought seriously affects the yield and quality of apples. γ-aminobutyric acid (GABA) plays an important role in the responses of plants to various stresses. However, the role and possible mechanism of GABA in the drought response of apple seedlings remain unknown. To explore the effect of GABA on apple seedlings under drought stress, seedlings of Malus hupehensis were treated with seven concentrations of GABA, and the response of seedlings under 15-day drought stress was observed. The results showed that 0.5 mM GABA was the most effective at relieving drought stress. Treatment with GABA reduced the relative electrical conductivity and MDA content of leaves induced by drought stress and significantly increased the relative water content of leaves. Exogenous GABA significantly decreased the stomatal conductance and intercellular carbon dioxide concentration and transpiration rate, and it significantly increased the photosynthetic rate under drought. GABA also reduced the accumulation of superoxide anions and hydrogen peroxide in leaf tissues under drought and increased the activities of POD, SOD, and CAT and the content of GABA. Exogenous treatment with GABA acted through the accumulation of abscisic acid (ABA) in the leaves to significantly decrease stomatal conductance and increase the stomatal closure rate, and the levels of expression of ABA-related genes PYL4, ABI1, ABI2, HAB1, ABF3, and OST1 changed in response to drought. Taken together, exogenous GABA can enhance the drought tolerance of apple seedlings.

Comprehensive Genome-Wide Identification and Transcript Profiling of GABA Pathway Gene Family in Apple (Malus domestica).

γ-Aminobutyric Acid (GABA), a four-carbon non-protein amino acid, is a significant component of the free amino acid pool in most prokaryotic and eukaryotic organisms. GABA is involved in pH regulation, maintaining C/N balance, plant development and defence, as well as a compatible osmolyte and an alternative pathway for glutamate utilization via anion flux. Glutamate decarboxylase (GAD, EC 4.1.1.15) and GABA transaminase (GABA-T, EC 2.6.1.19) are two key enzymes involved in the synthesis and metabolism of GABA. Recently, GABA transporters (GATs), protein and aluminium-activated malate transporter (ALMT) proteins which function as GABA receptors, have been shown to be involved in GABA regulation. However, there is no report on the characterization of apple GABA pathway genes. In this study, we performed a genome-wide analysis and expression profiling of the GABA pathway gene family in the apple genome. A total of 24 genes were identified including five GAD genes (namely MdGAD 1-5), two GABA-T genes (namely MdGABA-T 1,2), 10 GAT genes (namely GAT 1-10) and seven ALMT genes (namely MdALMT1-7). These genes were randomly distributed on 12 chromosomes. Phylogenetic analyses grouped GABA shunt genes into three clusters-cluster I, cluster II, and cluster III-which had three, four, and five genes, respectively. The expression profile analysis revealed significant MdGAD4 expression levels in both fruit and flower organs, except pollen. However, there were no significant differences in the expression of other GABA shunt genes in different tissues. This work provides the first characterization of the GABA shunt gene family in apple and suggests their importance in apple response to abiotic stress. These results can serve as a guide for future studies on the understanding and functional characterization of these gene families.

Heterologous Expression of the Melatonin-Related Gene HIOMT Improves Salt Tolerance in Malus domestica.

Melatonin, a widely known indoleamine molecule that mediates various animal and plant physiological processes, is formed from N-acetyl serotonin via N-acetylserotonin methyltransferase (ASMT). ASMT is an enzyme that catalyzes melatonin synthesis in plants in the rate-determining step and is homologous to hydroxyindole-O-methyltransferase (HIOMT) melatonin synthase in animals. To date, little is known about the effect of HIOMT on salinity in apple plants. Here, we explored the melatonin physiological function in the salinity condition response by heterologous expressing the homologous human HIOMT gene in apple plants. We discovered that the expression of melatonin-related gene (MdASMT) in apple plants was induced by salinity. Most notably, compared with the wild type, three transgenic lines indicated higher melatonin levels, and the heterologous expression of HIOMT enhanced the expression of melatonin synthesis genes. The transgenic lines showed reduced salt damage symptoms, lower relative electrolyte leakage, and less total chlorophyll loss from leaves under salt stress. Meanwhile, through enhanced activity of antioxidant enzymes, transgenic lines decreased the reactive oxygen species accumulation, downregulated the expression of the abscisic acid synthesis gene (MdNCED3), accordingly reducing the accumulation of abscisic acid under salt stress. Both mechanisms regulated morphological changes in the stomata synergistically, thereby mitigating damage to the plants' photosynthetic ability. In addition, transgenic plants also effectively stabilized their ion balance, raised the expression of salt stress-related genes, as well as alleviated osmotic stress through changes in amino acid metabolism. In summary, heterologous expression of HIOMT improved the adaptation of apple leaves to salt stress, primarily by increasing melatonin concentration, maintaining a high photosynthetic capacity, reducing reactive oxygen species accumulation, and maintaining normal ion homeostasis.

Overexpression of the tyrosine decarboxylase gene MdTyDC in apple enhances long-term moderate drought tolerance and WUE.

Drought stress affects the apple yield and quality. Tyrosine decarboxylase (TyDC) plays a fundamental role in many secondary metabolite reactions in higher plants (including those involving dopamine). Our aims of this study are: 1) to identify the role of TyDC in dopamine derivative biosynthesis and its function in long-term moderate drought conditions; and 2) to explore the role of MdTyDC in plant growth and development as well as the drought stress response. Wild type and three independently apple plants overexpression of MdTyDC were treated for long-term moderate drought stress. The growth and physiological parameters of apple plant, photosynthetic capacity, antioxidant enzymes activity, water use efficiency (WUE), stomatal behavior, amino acid content and dopamine content were detected under long-term moderate drought stress. Overexpression of MdTyDC (OE) in apple showed better growth performance, higher photosynthetic capacity and higher capacity for photochemical reactions than wild type lines (WT). Under long-term moderate drought stress, OE lines showed higher WUE, increased ABA content, decreased stomatal aperture, higher antioxidant activity, lower accumulation of ROS and increases in amino acids, such as proline, phenylalanine and lysine. In addition, qRT-PCR revealed higher gene expression of MdTyDC and dopamine content in OE compared with WT lines under long-term moderate drought stress. These results indicate that MdTyDC confers long-term moderate drought tolerance by improving photosynthetic capacity, WUE, antioxidant activity, dopamine content and changing the contents of amino acids (such as proline accumulation).

MxRop1-MxrbohD1 interaction mediates ROS signaling in response to iron deficiency in the woody plant Malus xiaojinensis.

Iron (Fe) deficiency affects crop production and quality. Rho of plants (ROPs) involves in multiple physiological processes in plants. While it has not been well characterized under Fe deficiency, especially in perennial woody plants. In our study, we cloned ROP homologous gene MxRop1 from Malus xiaojinenesis, then overexpressed it in Arabidopsis, showing enhanced plant tolerance to Fe deficiency, which demonstrated its gene function during this stress. Overexpression of MxRop1 also increased reactive oxygen species (ROS) levels. Moreover, active state of MxRop1 (CA-MxRop1) interacted with N-terminal region of MxrbohD1, one ROS synthesis gene. When MxrbohD1 was overexpressed in apple calli, it showed significantly increased H2O2 content, fresh weight and FCR activity, while ROS inhibitor application dramatically inhibited FCR activity, demonstrating ROS produced by MxrbohD1 regulated Fe deficiency responses. Furthermore, using Agrobacterium rhizogenes transformation, MxrbohD1 was overexpressed in apple roots, with increased expression of Fe deficiency-induced genes and increased root FCR activity. Under Fe deficiency, it exhibited slight leaf yellowing phenotype. Co-expression of CA-MxRop1 and MxrbohD1 significantly induced ROS generation. Finally, we proposed that MxRop1 interacted with MxrbohD1 to modulate ROS mediated Fe deficiency adaptive responses in Malus xiaojinensis, which will provide a guidance of cultivation of Fe-deficiency tolerant apple plant.

Analyses of the photosynthetic characteristics, chloroplast ultrastructure, and transcriptome of apple (Malus domestica) grown under red and blue lights.

BACKGROUND: Light quality significantly affects plant growth and development, photosynthesis, and carbon and nitrogen metabolism. Apple (Malus domestica Borkh.) is a widely cultivated and economically important fruit crop worldwide. However, there are still few studies on the effects of different light qualities on the growth and development of apple seedlings. RESULTS: In this study, we explored the effects of blue and red light treatments on the growth and development, photosynthetic characteristics, leaf chloroplast ultrastructure, and carbon and nitrogen metabolism of apple seedlings. Blue light significantly inhibited apple plant growth and leaf extension, but it promoted the development of leaf tissue structures and chloroplasts and positively affected leaf stomatal conductance, the transpiration rate, and photosynthetic efficiency. The red light treatment promoted apple plant growth and root development, but it resulted in loosely organized leaf palisade tissues and low chlorophyll contents. The blue and red light treatments enhanced the accumulation of ammonium nitrogen in apple seedlings. Moreover, the blue light treatment significantly promoted nitrogen metabolism. Additionally, an RNA-seq analysis revealed that both blue light and red light can significantly up-regulate the expression of genes related to carbon and nitrogen metabolism. Blue light can also promote amino acid synthesis and flavonoid metabolism, whereas red light can induce plant hormone signal transduction. The expression of a gene encoding a bHLH transcription factor (MYC2-like) was significantly up-regulated in response to blue light, implying it may be important for blue light-mediated plant development. CONCLUSIONS: Considered together, blue and red light have important effects on apple growth, carbon and nitrogen metabolism. These findings may be useful for determining the ideal light conditions for apple cultivation to maximize fruit yield and quality.

Transcriptomics of Improved Fruit Retention by Hexanal in 'Honeycrisp' Reveals Hormonal Crosstalk and Reduced Cell Wall Degradation in the Fruit Abscission Zone.

Apples (Malus domestica Borkh) are prone to preharvest fruit drop, which is more pronounced in 'Honeycrisp'. Hexanal is known to improve fruit retention in several economically important crops. The effects of hexanal on the fruit retention of 'Honeycrisp' apples were assessed using physiological, biochemical, and transcriptomic approaches. Fruit retention and fruit firmness were significantly improved by hexanal, while sugars and fresh weight did not show a significant change in response to hexanal treatment. At commercial maturity, abscisic acid and melatonin levels were significantly lower in the treated fruit abscission zone (FAZ) compared to control. At this stage, a total of 726 differentially expressed genes (DEGs) were identified between treated and control FAZ. Functional classification of the DEGs showed that hexanal downregulated ethylene biosynthesis genes, such as S-adenosylmethionine synthase (SAM2) and 1-aminocyclopropane-1-carboxylic acid oxidases (ACO3, ACO4, and ACO4-like), while it upregulated the receptor genes ETR2 and ERS1. Genes related to ABA biosynthesis (FDPS and CLE25) were also downregulated. On the contrary, key genes involved in gibberellic acid biosynthesis (GA20OX-like and KO) were upregulated. Further, hexanal downregulated the expression of genes related to cell wall degrading enzymes, such as polygalacturonase (PG1), glucanases (endo-ß-1,4-glucanase), and expansins (EXPA1-like, EXPA6, EXPA8, EXPA10-like, EXPA16-like). Our findings reveal that hexanal reduced the sensitivity of FAZ cells to ethylene and ABA. Simultaneously, hexanal maintained the cell wall integrity of FAZ cells by regulating genes involved in cell wall modifications. Thus, delayed fruit abscission by hexanal is most likely achieved by minimizing ABA through an ethylene-dependent mechanism.

An HD-ZIP transcription factor, MxHB13, integrates auxin-regulated and juvenility-determined control of adventitious rooting in Malus xiaojinensis.

Adventitious root (AR) formation is a critical factor in the vegetative propagation of forestry and horticultural plants. Competence for AR formation declines in many species during the miR156/SPL-mediated vegetative phase change. Auxin also plays a regulatory role in AR formation. In apple rootstock, both high miR156 expression and exogenous auxin application are prerequisites for AR formation. However, the mechanism by which the miR156/SPL module interacts with auxin in controlling AR formation is unclear. In this paper, leafy cuttings of juvenile (Mx-J) and adult (Mx-A) phase Malus xiaojinensis were used in an RNA-sequencing experiment. The results revealed that numerous genes involved in phytohormone signaling, carbohydrate metabolism, cell dedifferentiation, and reactivation were downregulated in Mx-A cuttings in response to indole butyric acid treatment. Among the differentially expressed genes, an HD-ZIP transcription factor gene, MxHB13, was found to be under negative regulation of MdSPL26 by directly binding to MxHB13 promoter. MxTIFY9 interacts with MxSPL26 and may play a role in co-repressing the expression of MxHB13. The expression of MxTIFY9 was induced by exogenous indole butyric acid. MxHB13 binds to the promoter of MxABCB19-2 and positively affects the expression. A model is proposed in which MxHB13 links juvenility-limited and auxin-limited AR recalcitrance mechanisms in Mx-A.

Molecular mechanism of MdWUS2-MdTCP12 interaction in mediating cytokinin signaling to control axillary bud outgrowth.

Shoot branching is an important factor that influences the architecture of apple trees and cytokinin is known to promote axillary bud outgrowth. The cultivar 'Fuji', which is grown on ~75% of the apple-producing area in China, exhibits poor natural branching. The TEOSINTE BRANCHED1/CYCLOIDEA/PCF (TCP) family genes BRANCHED1/2 (BRC1/2) are involved in integrating diverse factors that function locally to inhibit shoot branching; however, the molecular mechanism underlying the cytokinin-mediated promotion of branching that involves the repression of BRC1/2 remains unclear. In this study, we found that apple WUSCHEL2 (MdWUS2), which interacts with the co-repressor TOPLESS-RELATED9 (MdTPR9), is activated by cytokinin and regulates branching by inhibiting the activity of MdTCP12 (a BRC2 homolog). Overexpressing MdWUS2 in Arabidopsis or Nicotiana benthamiana resulted in enhanced branching. Overexpression of MdTCP12 inhibited axillary bud outgrowth in Arabidopsis, indicating that it contributes to the regulation of branching. In addition, we found that MdWUS2 interacted with MdTCP12 in vivo and in vitro and suppressed the ability of MdTCP12 to activate the transcription of its target gene, HOMEOBOX PROTEIN 53b (MdHB53b). Our results therefore suggest that MdWUS2 is involved in the cytokinin-mediated inhibition of MdTCP12 that controls bud outgrowth, and hence provide new insights into the regulation of shoot branching by cytokinin.