Control of ovule development in Vitis vinifera by VvMADS28 and interacting genes.

Seedless grapes are increasingly popular throughout the world, and the development of seedless varieties is a major breeding goal. In this study, we demonstrate an essential role for the grapevine MADS-box gene VvMADS28 in morphogenesis of the ovule. We found that VvMADS28 mRNA accumulated in the ovules of a seeded cultivar, 'Red Globe', throughout the course of ovule and seed development, especially within the integument/seed coat. In contrast, in the seedless cultivar 'Thompson Seedless', VvMADS28 was expressed only weakly in ovules, and this was associated with increased levels of histone H3 lysine 27 trimethylation (H3K27me3) within the VvMADS28 promoter region. RNAi-mediated transient suppression of VvMADS28 expression in 'Red Globe' led to reduced seed size associated with inhibition of episperm and endosperm cell development. Heterologous overexpression of VvMADS28 in transgenic tomatoes interfered with sepal development and resulted in smaller fruit but did not obviously affect seed size. Assays in yeast cells showed that VvMADS28 is subject to regulation by the transcription factor VvERF98, and that VvMADS28 could interact with the Type I/ Mß MADS-domain protein VvMADS5. Moreover, through DNA-affinity purification-sequencing (DAP-seq), we found that VvMADS28 protein specifically binds to the promoter of the grapevine WUSCHEL (VvWUS) gene, suggesting that maintenance of the VvMADS28-VvMADS5 dimer and VvWUS expression homeostasis influences seed development. Taken together, our results provide insight into regulatory mechanisms of ovule and seed development associated with VvMADS28.

Conservation and divergence of expression of GA2-oxidase homeologs in apple (Malus x domestica Borkh.).

In domesticated apple (Malus x domestica Borkh.) and other woody perennials, floral initiation can be repressed by gibberellins (GAs). The associated mechanism is a major unanswered question in plant physiology, and understanding organismal aspects of GA signaling in apple has important commercial applications. In plants, the major mechanism for elimination of GAs and resetting of GA signaling is through catabolism by GA2-oxidases (GA2ox). We found that the GA2ox gene family in apple comprises 16 genes representing eight, clearly defined homeologous pairs, which were named as MdGA2ox1A/1B to MdGA2ox8A/8B. Expression of the genes was analyzed in the various structures of the spur, where flowers are initiated, as well as in various structures of seedlings over one diurnal cycle and in response to water-deficit and salt stress. Among the results, we found that MdGA2ox2A/2B dominated expression in the shoot apex and were strongly upregulated in the apex after treatment with exogenous GA3, suggesting potential involvement in repression of flowering. Several MdGA2ox genes also showed preferential expression in the leaf petiole, fruit pedicel, and the seed coat of developing seeds, potentially representing mechanisms to limit diffusion of GAs across these structures. In all contexts studied, we documented both concerted and distinct expression of individual homeologs. This work introduces an accessible woody plant model for studies of GA signaling, GA2ox gene regulation, and conservation/divergence of expression of homeologous genes, and should find application in development of new cultivars of apple and other tree fruits.

The putative ABCG transporter VviABCG20 from grapevine (Vitis vinifera) is strongly expressed in the seed coat of developing seeds and may participate in suberin biosynthesis.

Half-size ATP binding cassette G (ABCG) transporters participate in many biological processes by transporting specific substrates. Our previous study showed that VviABCG20 was strongly expressed in the seeds of seeded grape and the silencing of VviABCG20 homolog gene in tomato led to a reduction in seed number. To reveal the molecular mechanism of VviABCG20 gene involved in grape seed development/abortion, the gene expression and functional analysis of VviABCG20 were further carried out in the grapevine. It was shown that the gene expression of VviABCG20 was higher in seeds of seeded grapes compared with seedless. Further the expression of VviABCG20 in the seed coat was significantly higher than in ovules (young seeds) and endosperm. VviABCG20 was also induced by exogenous hormones (especially MeJA) in grape leaves. Subcellular localization analysis showed that VviABCG20 is a membrane protein. In overexpressed VviABCG20 transgenic callus of Thompson seedless, expression of genes GPAT5, FAR1 and FAR5 was increased significantly. After treatment with suberin precursors, the transgenic callus reduced the sensitivity to three cinnamic acid derivatives (cis-ferulic acid, caffeic acid, coumaric acid), succinic acid, and glycerol. In suspension cells, expression of VviABCG20 was increased significantly after treatment with suberin precursors. Our research suggested that VviABCG20 may function in seed development in grapevine, at least in part by participating in suberin biosynthesis in the seed coat.

Role of grapevine SEPALLATA-related MADS-box gene VvMADS39 in flower and ovule development.

Seedlessness is one of the most important breeding goals for table grapes; thus, understanding the molecular genetic regulation of seed development and abortion is critical for the development of seedless cultivars. In the present study, we characterized VvMADS39, a class E MADS-box gene of grapevine (Vitis vinifera) orthologous to Arabidopsis SEP2. Heterologous overexpression of VvMADS39 in tomato reduced the fruit and seed size and seed number. Targeted mutagenesis of the homologous SlMADS39 in tomato induced various floral and fruit defects. It could reasonable to suppose that active VvMADS39 expression in "Thompson Seedless" may restrict cellular expansion, resulting in the development of smaller fruits and seeds, VvMADS39 may play a role in the regulation of ovule development in grapevine and contributes to seedless fruit formation. In contrast, VvMADS39 suppression in "Red Globe" was associated with enhanced histone H3 lysine 27 trimethylation in the promoter region of VvMADS39, allowing normal ovule and fruit development; Meanwhile, VvMADS39 interacts with VvAGAMOUS, and the activity of the VvMADS39-VvAGAMOUS dimer to induce integument development requires the activation and maintenance of VvINO expression. The synergistic cooperation between VvMADS39 and related proteins plays an important role in maintaining floral meristem characteristics, and fruit and ovule development.

Fine-tuning of SUMOylation modulates drought tolerance of apple.

SUMOylation is involved in various aspects of plant biology, including drought stress. However, the relationship between SUMOylation and drought stress tolerance is complex; whether SUMOylation has a crosstalk with ubiquitination in response to drought stress remains largely unclear. In this study, we found that both increased and decreased SUMOylation led to increased survival of apple (Malus × domestica) under drought stress: both transgenic MdSUMO2A overexpressing (OE) plants and MdSUMO2 RNAi plants exhibited enhanced drought tolerance. We further confirmed that MdDREB2A is one of the MdSUMO2 targets. Both transgenic MdDREB2A OE and MdDREB2AK192R OE plants (which lacked the key site of SUMOylation by MdSUMO2A) were more drought tolerant than wild-type plants. However, MdDREB2AK192R OE plants had a much higher survival rate than MdDREB2A OE plants. We further showed SUMOylated MdDREB2A was conjugated with ubiquitin by MdRNF4 under drought stress, thereby triggering its protein degradation. In addition, MdRNF4 RNAi plants were more tolerant to drought stress. These results revealed the molecular mechanisms that underlie the relationship of SUMOylation with drought tolerance and provided evidence for the tight control of MdDREB2A accumulation under drought stress mediated by SUMOylation and ubiquitination.

Overexpression of the Apple (Malus × domestica) MdERF100 in Arabidopsis Increases Resistance to Powdery Mildew.

APETALA2/ETHYLENE RESPONSIVE FACTOR (AP2/ERF) transcription factors play important roles in plant development and stress response. Although AP2/ERF genes have been extensively investigated in model plants such as Arabidopsis thaliana, little is known about their role in biotic stress response in perennial fruit tree crops such as apple (Malus × domestica). Here, we investigated the role of MdERF100 in powdery mildew resistance in apple. MdERF100 localized to the nucleus but showed no transcriptional activation activity. The heterologous expression of MdERF100 in Arabidopsis not only enhanced powdery mildew resistance but also increased reactive oxygen species (ROS) accumulation and cell death. Furthermore, MdERF100-overexpressing Arabidopsis plants exhibited differential expressions of genes involved in jasmonic acid (JA) and salicylic acid (SA) signaling when infected with the powdery mildew pathogen. Additionally, yeast two-hybrid and bimolecular fluorescence complementation assays confirmed that MdERF100 physically interacts with the basic helix-loop-helix (bHLH) protein MdbHLH92. These results suggest that MdERF100 mediates powdery mildew resistance by regulating the JA and SA signaling pathways, and MdbHLH92 is involved in plant defense against powdery mildew. Overall, this study enhances our understanding of the role of MdERF genes in disease resistance, and provides novel insights into the molecular mechanisms of powdery mildew resistance in apple.

Genetic mechanisms associated with floral initiation and the repressive effect of fruit on flowering in apple (Malus x domestica Borkh).

Many apple cultivars are subject to biennial fluctuations in flowering and fruiting. It is believed that this phenomenon is caused by a repressive effect of developing fruit on the initiation of flowers in the apex of proximal bourse shoots. However, the genetic pathways of floral initiation are incompletely described in apple, and the biological nature of floral repression by fruit is currently unknown. In this study, we characterized the transcriptional landscape of bourse shoot apices in the biennial cultivar, 'Honeycrisp', during the period of floral initiation, in trees bearing a high fruit load and in trees without fruit. Trees with high fruit load produced almost exclusively vegetative growth in the subsequent year, whereas the trees without fruit produced flowers on the majority of the potential flowering nodes. Using RNA-based sequence data, we documented gene expression at high resolution, identifying >11,000 transcripts that had not been previously annotated, and characterized expression profiles associated with vegetative growth and flowering. We also conducted a census of genes related to known flowering genes, organized the phylogenetic and syntenic relationships of these genes, and compared expression among homeologs. Several genes closely related to AP1, FT, FUL, LFY, and SPLs were more strongly expressed in apices from non-bearing, floral-determined trees, consistent with their presumed floral-promotive roles. In contrast, a homolog of TFL1 exhibited strong and persistent up-regulation only in apices from bearing, vegetative-determined trees, suggesting a role in floral repression. Additionally, we identified four GIBBERELLIC ACID (GA) 2 OXIDASE genes that were expressed to relatively high levels in apices from bearing trees. These results define the flowering-related transcriptional landscape in apple, and strongly support previous studies implicating both gibberellins and TFL1 as key components in repression of flowering by fruit.

Physiological and transcriptome analyses of the effects of exogenous dopamine on drought tolerance in apple.

Water shortage is one of the main limiting factors in apple (Malus domestica Borkh.) production. Although dopamine is produced in plants and has been linked with response to abiotic stress, the underlying mechanism remains unknown. In this study, physiological analyses revealed that pretreatment with 100 µM dopamine alleviated drought stress in apple seedlings. Dopamine inhibited the degradation of photosynthetic pigments and increased net photosynthetic rate under drought stress. Dopamine also reduced H2O2 content, possibly through direct scavenging and by mediating the antioxidant enzyme activity. Seedlings pretreated with dopamine had higher sucrose and malic acid contents but lower starch accumulation in their leaves. RNA-Seq analysis identified 1052 differentially expressed genes (DEGs) between non-treated and dopamine-pretreated plants under drought. An in-depth analysis of these DEGs revealed that dopamine regulated the expression of genes related to metabolism of nitrogen, secondary compounds, and amino acids under drought stress. In addition, dopamine may improve apple drought tolerance by activating Ca2+ signaling pathways through increased expression of CNGC and CAM/CML family genes. Moreover, analysis of transcription factor expression suggested that dopamine affected drought tolerance mainly through the regulation of WRKY, ERF, and NAC transcription factors.

Overexpression of MdIAA9 confers high tolerance to osmotic stress in transgenic tobacco.

Auxin is a plant hormone that takes part in a series of developmental and physiological processes. There are three major gene families that play a role in the early response of auxin and auxin/indole-3-acetic acid (Aux/IAA) is one of these. Although the genomic organization and function of Aux/IAA genes have been recognized in reference plants there have only been a few focused studies conducted with non-model crop plants, especially in the woody perennial species. We conducted a genomic census and expression analysis of Aux/IAA genes in the cultivated apple (Malus × domestica Borkh.). The Aux/IAA gene family of the apple genome was identified and analyzed in this study. Phylogenetic analysis showed that MdIAAs could be categorized into nine subfamilies and that these MdIAA proteins contained four whole or partially conserved domains of the MdIAA family. The spatio-specific expression profiles showed that most of the MdIAAs were preferentially expressed in specific tissues. Some of these genes were significantly induced by treatments with one or more abiotic stresses. The overexpression of MdIAA9 in tobacco (Nicotiana tabacum L.) plants significantly increased their tolerance to osmotic stresses. Our cumulative data supports the interactions between abiotic stresses and plant hormones and provides a theoretical basis for the mechanism of Aux/IAA and drought resistance in apples.

The grapevine homeobox gene VvHB58 influences seed and fruit development through multiple hormonal signaling pathways.

BACKGROUND: The homeobox transcription factor has a diversity of functions during plant growth and development process. Previous transcriptome analyses of seed development in grape hybrids suggested that specific homeodomain transcription factors are involved in seed development in seedless cultivars. However, the molecular mechanism of homeobox gene regulating seed development in grape is rarely reported. RESULTS: Here, we report that the grapevine VvHB58 gene, encoding a homeodomain-leucine zipper (HD-Zip I) transcription factor, participates in regulating fruit size and seed number. The VvHB58 gene was differentially expressed during seed development between seedless and seeded cultivars. Subcellular localization assays revealed that the VvHB58 protein was located in the nucleus. Transgenic expression of VvHB58 in tomato led to loss of apical dominance, a reduction in fruit pericarp expansion, reduced fruit size and seed number, and larger endosperm cells. Analysis of the cytosine methylation levels within the VvHB58 promoter indicated that the differential expression during seed development between seedless and seeded grapes may be caused by different transcriptional regulatory mechanisms rather than promoter DNA methylation. Measurements of five classic endogenous hormones and expression analysis of hormone-related genes between VvHB58 transgenic and nontransgenic control plants showed that expression of VvHB58 resulted in significant changes in auxin, gibberellin and ethylene signaling pathways. Additionally, several DNA methylation-related genes were expressed differentially during seed development stages in seedless and seeded grapes, suggesting changes in methylation levels during seed development may be associated with seed abortion. CONCLUSION: VvHB58 has a potential function in regulating fruit and seed development by impacting multiple hormonal pathways. These results expand understanding of homeodomain transcription factors and potential regulatory mechanism of seed development in grapevine, and provided insights into molecular breeding for grapes.

Genome-Wide Analysis of the YABBY Gene Family in Grapevine and Functional Characterization of VvYABBY4.

Genes of the plant-specific YABBY transcription factor family have various roles, including lateral organ development, establishment of dorsoventral polarity, and response to abiotic stress. In this study, we carried out a genomic census of YABBY genes in grapevine (Vitis vinifera) and characterized their expression pattern during ovule development. We identified seven YABBY genes and classified them into five subfamilies, based on peptide sequence, similarity of exon-intron structure and composition of peptide sequence motifs. Analysis of YABBY gene expression in various grapevine structures and organs suggested that these genes function in diverse aspects of development and physiology. Analysis of expression during ovule development in four cultivars showed that one gene, VvYABBY4, was preferentially expressed during the period of ovule abortion in seedless cultivars. Transgenic expression of VvYABBY4 in tomato conferred reduced plant stature, dark green leaves, elongated pistil, and reduced size of fruit and seeds. Reduced seed size was associated with smaller endosperm cells. Expression of VvYABBY4 also affected expression of numerous tomato genes with presumed roles in seed development. These data suggest the potential for VvYABBY4 to influence seed development in grapevine, which may impact seedless grape breeding.

Genetic mechanisms in the repression of flowering by gibberellins in apple (Malus x domestica Borkh.).

BACKGROUND: Gibberellins (GAs) can have profound effects on growth and development in higher plants. In contrast to their flowering-promotive role in many well-studied plants, GAs can repress flowering in woody perennial plants such as apple (Malus x domestica Borkh.). Although this effect of GA on flowering is intriguing and has commercial importance, the genetic mechanisms linking GA perception with flowering have not been well described. RESULTS: Application of a mixture of bioactive GAs repressed flower formation without significant effect on node number or shoot elongation. Using Illumina-based transcriptional sequence data and a newly available, high-quality apple genome sequence, we generated transcript models for genes expressed in the shoot apex, and estimated their transcriptional response to GA. GA treatment resulted in downregulation of a diversity of genes participating in GA biosynthesis, and strong upregulation of the GA catabolic GA2 OXIDASE genes, consistent with GA feedback and feedforward regulation, respectively. We also observed strong downregulation of numerous genes encoding potential GA transporters and receptors. Additional GA-responsive genes included potential components of cytokinin (CK), abscisic acid (ABA), brassinosteroid, and auxin signaling pathways. Finally, we observed rapid and strong upregulation of both of two copies of a gene previously observed to inhibit flowering in apple, MdTFL1 (TERMINAL FLOWER 1). CONCLUSION: The rapid and robust upregulation of genes associated with GA catabolism in response to exogenous GA, combined with the decreased expression of GA biosynthetic genes, highlights GA feedforward and feedback regulation in the apple shoot apex. The finding that genes with potential roles in GA metabolism, transport and signaling are responsive to GA suggests GA homeostasis may be mediated at multiple levels in these tissues. The observation that TFL1-like genes are induced quickly in response to GA suggests they may be directly targeted by GA-responsive transcription factors, and offers a potential explanation for the flowering-inhibitory effects of GA in apple. These results provide a context for investigating factors that may transduce the GA signal in apple, and contribute to a preliminary genetic framework for the repression of flowering by GAs in a woody perennial plant.

The jasmonate-ZIM domain gene VqJAZ4 from the Chinese wild grape Vitis quinquangularis improves resistance to powdery mildew in Arabidopsis thaliana.

Grape (Vitis vinifera L.) is one of the most widely cultivated and economically important fruits. Most cultivated varieties of grape are highly susceptible to fungal diseases, and one of the most pervasive is powdery mildew, caused by Uncinula necator. The jasmonate-ZIM domain (JAZ) family proteins are critical for plant responses to environmental stresses. Here, we report the characterization of VqJAZ4, a jasmonate-ZIM domain gene isolated from Vitis quinquangularis, a Chinese wild Vitis species that exhibits high tolerance to several kinds of fungi. Subcellular localization assay indicated that the VqJAZ4 protein is targeted to the nucleus. The VqJAZ4 gene was strongly induced by U. necator inoculation, as well as by the defense-related hormones methyl jasmonate (MeJA) and salicylic acid (SA). The upregulation of VqJAZ4 after inoculation was dependent on its promoter sequences. Expression of VqJAZ4 in Arabidopsis thaliana improved resistance to powdery mildew. Histochemical staining assays indicated that plants expressing VqJAZ4 displayed a larger number of dead cells and stronger reactive oxygen species (ROS) burst than non-transgenic control (NTC) plants. Expression analysis of several disease-related genes suggested that VqJAZ4 expression enhanced defense responses though SA and/or JA signaling pathways. We also found that VqJAZ4-expressing Arabidopsis showed increased susceptibility to Botrytis cinerea. Taken together, these results provide evidence that VqJAZ4 may play an important role in response to fungal pathogens in grape, and may represent a candidate for future grape molecular breeding for disease resistance.

Apple whole genome sequences: recent advances and new prospects.

In 2010, a major scientific milestone was achieved for tree fruit crops: publication of the first draft whole genome sequence (WGS) for apple (Malus domestica). This WGS, v1.0, was valuable as the initial reference for sequence information, fine mapping, gene discovery, variant discovery, and tool development. A new, high quality apple WGS, GDDH13 v1.1, was released in 2017 and now serves as the reference genome for apple. Over the past decade, these apple WGSs have had an enormous impact on our understanding of apple biological functioning, trait physiology and inheritance, leading to practical applications for improving this highly valued crop. Causal gene identities for phenotypes of fundamental and practical interest can today be discovered much more rapidly. Genome-wide polymorphisms at high genetic resolution are screened efficiently over hundreds to thousands of individuals with new insights into genetic relationships and pedigrees. High-density genetic maps are constructed efficiently and quantitative trait loci for valuable traits are readily associated with positional candidate genes and/or converted into diagnostic tests for breeders. We understand the species, geographical, and genomic origins of domesticated apple more precisely, as well as its relationship to wild relatives. The WGS has turbo-charged application of these classical research steps to crop improvement and drives innovative methods to achieve more durable, environmentally sound, productive, and consumer-desirable apple production. This review includes examples of basic and practical breakthroughs and challenges in using the apple WGSs. Recommendations for "what's next" focus on necessary upgrades to the genome sequence data pool, as well as for use of the data, to reach new frontiers in genomics-based scientific understanding of apple.

Overexpression of a protein kinase gene MpSnRK2.10 from Malus prunifolia confers tolerance to drought stress in transgenic Arabidopsis thaliana and apple.

Members of the sucrose non-fermenting-1-related protein kinase 2 (SnRK2) family play central roles in the abscisic acid (ABA) signaling pathway and in mediating osmotic stress signaling and tolerance in plants. Previously, 12 full-length coding sequences that belong to SnRK2 gene family were identified and cloned in wild apple species Malus prunifolia. In this study, one of the members, MpSnRK2.10, was overexpressed in Arabidopsis and apple to investigate its potential function in response to drought stress. The results showed that overexpression of this gene did not affect plant growth under normal conditions. However, the transgenic plants showed enhanced tolerance to drought, as indicated by the amelioration in phenotype appearance and physiological indices related to drought stress damage. Additionally, transgenic apple plants overexpressing MpSnRK2.10 exhibited greater sensitivity to ABA compared to wild-type (WT) plants. Moreover, expressions of three stress response genes, MdRAB18, MdRD22, and MdRD29B, were more strongly induced in transgenic apple plants than in the WT when subjected to ABA and mannitol treatments. Taken as a whole, our study suggests that MpSnRK2.10 has a role in the enhancement of ABA signal transduction in response to stress, and that the manipulation of MpSnRK2.10 expression could be a feasible approach for improving abiotic stress tolerance in apple and other important crops.

Comprehensive genomic analysis of the TYROSINE AMINOTRANSFERASE (TAT) genes in apple (Malus domestica) allows the identification of MdTAT2 conferring tolerance to drought and osmotic stresses in plants.

Tyrosine aminotransferase (TAT, EC 2.6.1.5) is the first key enzyme that catalyzes the reversible interconversion of tyrosine and 4-hydroxyphenylpyruvate in the tyrosine-derived pathway for syntheses of important secondary metabolites and compounds. Although plant TAT genes have been proposed to be important in response to abiotic stress, there is little information about TAT genes in woody perennial tree species, especially in economic fruit trees. Based on TAT domain searching, sequence homology screening and phylogenetic analysis, we identified four TATs in apple genome. Then, we carried out a detailed phylogenetic analysis of TAT genes from multi-species, focusing on apple (Malus domestica). The result showed that the TAT family comprises three major classes corresponding to genes from angiosperms, mammals, and bacteria. Angiosperm TAT genes could be further divided into six subclasses. Analysis of intron-exon structure revealed that the typical TAT gene contains six introns and seven exons, with exons of similar size at each exon location. Promoter analysis showed that the 5'-flanking region of apple MdTATs contain multiple cis-acting elements including those implicated in light, biotic stress, abiotic stress, and hormone response. MdTATs were expressed to various levels in all apple structures and organs evaluated, and showed distinct expression patterns under water deficit stress. Ectopic expression of MdTAT2 in Arabidopsis or over-expression of MdTAT2 in apple callus tissue conferred enhanced tolerance to drought and osmotic stress. Collectively, these results suggest a role for TAT genes in drought and osmotic stresses and provide valuable information for further research of TAT genes and their function in plants.

Inferences on specificity recognition at the Malus×domestica gametophytic self-incompatibility system.

In Malus × domestica (Rosaceae) the product of each SFBB gene (the pollen component of the gametophytic self-incompatibility (GSI) system) of a S-haplotype (the combination of pistil and pollen genes that are linked) interacts with a sub-set of non-self S-RNases (the pistil component), but not with the self S-RNase. To understand how the Malus GSI system works, we identified 24 SFBB genes expressed in anthers, and determined their gene sequence in nine M. domestica cultivars. Expression of these SFBBs was not detected in the petal, sepal, filament, receptacle, style, stigma, ovary or young leaf. For all SFBBs (except SFBB15), identical sequences were obtained only in cultivars having the same S-RNase. Linkage with a particular S-RNase was further established using the progeny of three crosses. Such data is needed to understand how other genes not involved in GSI are affected by the S-locus region. To classify SFBBs specificity, the amino acids under positive selection obtained when performing intra-haplotypic analyses were used. Using this information and the previously identified S-RNase positively selected amino acid sites, inferences are made on the S-RNase amino acid properties (hydrophobicity, aromatic, aliphatic, polarity, and size), at these positions, that are critical features for GSI specificity determination.

Single-base methylome analysis reveals dynamic epigenomic differences associated with water deficit in apple.

Cytosine methylation is an essential feature of epigenetic regulation and is involved in various biological processes. Although cytosine methylation has been analysed at the genomic scale for several plant species, there is a general lack of understanding of the dynamics of global and genic DNA methylation in plants growing in environments challenged with biotic and abiotic stresses. In this study, we mapped cytosine methylation at single-base resolution in the genome of commercial apple (Malus x domestica), and analysed changes in methylation patterns associated with water deficit in representative drought-sensitive and drought-tolerant cultivars. We found that the apple genome exhibits ~54%, ~38% and ~8.5% methylation at CG, CHG and CHH sequence contexts, respectively. We additionally documented changes in gene expression associated with water deficit in an attempt to link methylation and gene expression changes. Global methylation and transcription analysis revealed that promoter-unmethylated genes showed higher expression levels than promoter-methylated genes. Gene body methylation appears to be positively correlated with gene expression. Water deficit stress was associated with changes in methylation at a multitude of genes, including those encoding transcription factors (TFs) and transposable elements (TEs). These results present a methylome map of the apple genome and reveal widespread DNA methylation alterations in response to water deficit stress. These data will be helpful for understanding potential linkages between DNA methylation and gene expression in plants growing in natural environments and challenged with abiotic and biotic stresses.