FLOWERING LOCUS T1 and TERMINAL FLOWER1 regulatory networks mediate flowering initiation in apple.

Flower bud formation is a critical process that directly determines yield and fruit quality in fruit crops. Floral induction is modulated by the balance between 2 flowering-related proteins, FLOWERING LOCUS T (FT) and TERMINAL FLOWER1 (TFL1); however, the mechanisms underlying the establishment and maintenance of this dynamic balance remain largely elusive. Here, we showed that in apple (Malus × domestica Borkh.), MdFT1 is predominantly expressed in spur buds and exhibits an increase in expression coinciding with flower induction; in contrast, MdTFL1 exhibited downregulation in apices during flower induction, suggesting that MdTFL1 has a role in floral repression. Interestingly, both the MdFT1 and MdTFL1 transcripts are directly regulated by transcription factor basic HELIX-LOOP-HELIX48 (MdbHLH48), and overexpression of MdbHLH48 in Arabidopsis (Arabidopsis thaliana) and tomato (Solanum lycopersicum) results in accelerated flowering. Binding and activation analyses revealed that MdbHLH48 functions as a positive regulator of MdFT1 and a negative regulator of MdTFL1. Further studies established that both MdFT1 and MdTFL1 interact competitively with MdWRKY6 protein to facilitate and inhibit, respectively, MdWRKY6-mediated transcriptional activation of target gene APPLE FLORICAULA/LFY (AFL1, an apple LEAFY-like gene), ultimately regulating apple flower bud formation. These findings illustrate the fine-tuned regulation of flowering by the MdbHLH48-MdFT1/MdTFL1-MdWRKY6 module and provide insights into flower bud formation in apples.

Age-dependent changes in leaf size in apple are governed by a cytokinin-integrated module.

Plants undergo various age-dependent changes in leaf morphology during juvenile to adult vegetative stage. However, the precise molecular mechanisms governing these changes in apple (Malus domestica) remain unknown. Here, we showed that CYTOKININ OXIDASE/DEHYDROGENASE5 (MdCKX5), an age-dependent gene, encodes a functional CKX enzyme and serves as the common downstream target of SQUAMOSA PROMOTER BINDING PROTEIN-LIKE (SPL) transcription factor MdSPL14 and WRKY transcription factor MdWRKY24 to control the degradation of cytokinin (CK). As the target of mdm-microRNA156a, MdSPL14 interacts with MdWRKY24 to coordinately repress the transcription of MdCKX5 by forming the age-mediated mdm-miR156a-MdSPL14-MdWRKY24 module, which regulates age-dependent changes in CK during the juvenile-to-adult phase transition. We further demonstrated that MdARR6, a type-A ARABIDOPSIS RESPONSE REGULATOR (ARR), is a negative feedback regulator in the CK signaling pathway. Silencing of MdARR6 in apple resulted in large leaves with smaller epidermal cells and a greater number of epidermal cells. Biochemical analysis showed that the mdm-miR156a-MdSPL14-MdWRKY24 module acts as a transcriptional repressor to directly regulate MdARR6 expression, thus controlling the age-dependent changes in leaf size by reducing CK responses. These findings established a link between the age pathway and CK signaling and revealed the molecular mechanism underlying age-dependent changes during the juvenile-to-adult phase transition; our results also provide targets for the genetic improvement of the vegetative phase transition in apple.

Plant growth promotion and biocontrol properties of a synthetic community in the control of apple disease.

BACKGROUND: Apple Replant Disease (ARD) is common in major apple-growing regions worldwide, but the role of rhizosphere microbiota in conferring ARD resistance and promoting plant growth remains unclear. RESULTS: In this study, a synthetic microbial community (SynCom) was developed to enhance apple plant growth and combat apple pathogens. Eight unique bacteria selected via microbial culture were used to construct the antagonistic synthetic community, which was then inoculated into apple seedlings in greenhouse experiments. Changes in the rhizomicroflora and the growth of aboveground plants were monitored. The eight strains, belonging to the genera Bacillus and Streptomyces, have the ability to antagonize pathogens such as Fusarium oxysporum, Rhizoctonia solani, Botryosphaeria ribis, and Physalospora piricola. Additionally, these eight strains can stably colonize in apple rhizosphere and some of them can produce siderophores, ACC deaminase, and IAA. Greenhouse experiments with Malus hupehensis Rehd indicated that SynCom promotes plant growth (5.23%) and increases the nutrient content of the soil, including soil organic matter (9.25%) and available K (1.99%), P (7.89%), and N (0.19%), and increases bacterial richness and the relative abundance of potentially beneficial bacteria. SynCom also increased the stability of the rhizosphere microbial community, the assembly of which was dominated by deterministic processes (|ß NTI| > 2). CONCLUSIONS: Our results provide insights into the contribution of the microbiome to pathogen inhibition and host growth. The formulation and manipulation of similar SynComs may be a beneficial strategy for promoting plant growth and controlling soil-borne disease.

Spatiotemporal patterns of leaf nutrients of wild apples in a wild fruit forest plot in the Ili Valley, China.

Malus sieversii, commonly known as wild apples, represents a Tertiary relict plant species and serves as the progenitor of globally cultivated apple varieties. Unfortunately, wild apple populations are facing significant degradation in localized areas due to a myriad of factors. To gain a comprehensive understanding of the nutrient status and spatiotemporal variations of M. sieversii, green leaves were collected in May and July, and the fallen leaves were collected in October. The concentrations of leaf nitrogen (N), phosphorus (P), and potassium (K) were measured, and the stoichiometric ratios as well as nutrient resorption efficiencies were calculated. The study also explored the relative contributions of soil, topographic, and biotic factors to the variation in nutrient traits. The results indicate that as the growing period progressed, the concentrations of N and P in the leaves significantly decreased (P < 0.05), and the concentration of K in October was significantly lower than in May and July. Throughout plant growth, leaf N-P and N-K exhibited hyperallometric relationships, while P-K showed an isometric relationship. Resorption efficiency followed the order of N < P < K (P < 0.05), with all three ratios being less than 1; this indicates that the order of nutrient limitation is K > P > N. The resorption efficiencies were mainly regulated by nutrient concentrations in fallen leaves. A robust spatial dependence was observed in leaf nutrient concentrations during all periods (70.1-97.9% for structural variation), highlighting that structural variation, rather than random factors, dominated the spatial variation. Nutrient resorption efficiencies (NRE, PRE, and KRE) displayed moderate structural variation (30.2-66.8%). The spatial patterns of nutrient traits varied across growth periods, indicating they are influenced by multifactorial elements (in which, soil property showed the highest influence). In conclusion, wild apples manifested differentiated spatiotemporal variability and influencing factors across various leaf nutrient traits. These results provide crucial insights into the spatiotemporal patterns and influencing factors of leaf nutrient traits of M. sieversii at the permanent plot scale for the first time. This work is of great significance for the ecosystem restoration and sustainable management of degrading wild fruit forests.

Self-(in)compatibility in Tunisian apple accessions [Malus domestica. Borkh]: S-genotypes identification and pollen tube growth analysis.

MAIN CONCLUSION: Self-incompatibility studies have revealed a potential use of Tunisian apple resources for crop improvement and modern breeding programs and a likely correlation between the pollen tube growth and flowering period. Apples [Malus domestica. Borkh] exhibit an S-RNase-based gametophytic self-incompatibility (GSI) system. Four primer combinations were used to S-genotype eighteen Tunisian local apple accessions and twelve introduced accessions that served as references. Within the Tunisian local accessions, S2, S3, S7, and S28 S-alleles were the most frequent and were assigned to 14 S-genotypes; among them, S7S28, S3S7, S2S5, and S2S3 were the most abundant. PCA plot showed that population structuring was affected by the S-alleles frequencies and revealed a modern origin of the Tunisian varieties rather than being ancient ones. Nonetheless, the results obtained with 17 SSR markers showed a separate grouping of local Tunisian accessions that calls into question the hypothesis discussed. Pollination experiments showed that the pollen started to germinate within 24 h of pollination but 48 h after pollination in the "El Fessi" accession. The first pollen tubes arrived in the styles within 36 h of pollination in two early flowering accessions known as "Arbi" and "Bokri", and after 72 h of pollination in late flowering "El Fessi" and 48 h after pollination in remaining accessions. The first pollen tube arrests were observed in accessions "Arbi" and "Bokri" within 84 h of pollination, within 108 h of pollination in "El Fessi" and within 108 h of pollination in remaining accessions. In the apple accession called "Boutabgaya," the pollen tubes reached the base of the style within 120 h of pollination without being aborted. Nevertheless, the self-compatible nature of "Boutabgaya" needs more studies to be confirmed. However, our results revealed the malfunction of the female component of the GSI in this accession. To conclude, this work paved the path for further studies to enhance the insight (i) into the relation between the flowering period and the pollen tube growth, (ii) self-compatible nature of "Boutabgaya", and (iii) the origin of the Tunisian 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.

Apple (Malus × domestica Borkh.) production and quality in response to anti-hail nets.

Anti-hail nets are the best way to mitigate the effects of hailstorms in the orchards. Apple trees covered with nets may exhibit a variety of vegetative and reproductive responses, inclusive of changes in tree vigour, cropping, sugar contents, and fruit colour. The present study was conducted to assess the effect of timing of installation and colour of anti-hail net on cropping and fruit quality in high-density apple orchard for two consecutive seasons (2021 and 2022). White and blue colour nets of size (9 m × 30 m) 80 GSM (square mesh with non-sliding threaded, leno weave, and < 30% shading factor) were installed at three different time intervals (15 days before estimated full bloom, at full bloom, and 15 days after full bloom) on apple cultivar 'Jeromine'. The installation at different time and colour of anti-hail nets significantly exhibit variability in cropping, fruit quality, and bio-chemical metrics. The significant highest cropping (fruit yield, productivity, and yield efficiency) and fruit biochemical parameters (total soluble solids) were recorded in T3C2 (15 days after full bloom + white colour anti-hail net) followed by T2C2 (installed at full bloom + white colour anti-hail net). Hence, white colour anti-hail nets installed 15 days after full bloom increased fruit production and improved quality in comparison to blue colour anti-hail net in apple under high-density plantations.

MdABI5 works with its interaction partners to regulate abscisic acid-mediated leaf senescence in apple.

Abscisic acid (ABA) induces chlorophyll degradation and leaf senescence; however, the molecular mechanism remains poorly understood, especially in woody plants. In this study, we found that MdABI5 plays an essential role in the regulation of ABA-triggered leaf senescence in Malus domestica (apple). Through yeast screening, three transcription factors, MdBBX22, MdWRKY40 and MdbZIP44, were found to interact directly with MdABI5 in vitro and in vivo. Physiological and biochemical assays showed that MdBBX22 delayed leaf senescence in two pathways. First, MdBBX22 interacted with MdABI5 to inhibit the transcriptional activity of MdABI5 on the chlorophyll catabolic genes MdNYE1 and MdNYC1, thus negatively regulating chlorophyll degradation and leaf senescence. Second, MdBBX22 interacted with MdHY5 to interfere with the transcriptional activation of MdHY5 on MdABI5, thereby inhibiting the expression of MdABI5, which also contributed to the delay of leaf senescence. MdWRKY40 and MdbZIP44 were identified as positive regulators of leaf senescence. They accelerated MdABI5-promoted leaf senescence through the same regulatory pathways, i.e., interacting with MdABI5 to enhance the transcriptional activity of MdABI5 on MdNYE1 and MdNYC1. Taken together, our results suggest that MdABI5 works with its positive or negative interaction partners to regulate ABA-mediated leaf senescence in apple, in which it acts as a core regulator. The antagonistic regulation pathways ensure that plants respond to external stresses flexibly and efficiently. Our results provide a concept for further study on the regulation mechanisms of leaf senescence.

The apple gene responsible for columnar tree shape reduces the abundance of biologically active gibberellin.

Ectopic expression of the apple 2-oxoglutarate-dependent dioxygenase (DOX, 2ODD) gene, designated MdDOX-Co, is thought to cause the columnar shape of apple trees. However, the mechanism underlying the formation of such a unique tree shape remains unclear. To solve this problem, we demonstrated that Arabidopsis thaliana overexpressing MdDOX-Co contained reduced levels of biologically active gibberellin (GA) compared with wild type. In summary: (i) with biochemical approaches, the gene product MdDOX-Co was shown to metabolize active GA A4 (GA4 ) to GA58 (12-OH-GA4 ) in vitro. MdDOX-Co also metabolized its precursors GA12 and GA9 to GA111 (12-OH-GA12 ) and GA70 (12-OH-GA9 ), respectively; (ii) Of the three 12-OH-GAs, GA58 was still active physiologically, but not GA70 or GA111 ; (iii) Arabidopsis MdDOX-Co OE transformants converted exogenously applied deuterium-labeled (d2 )-GA12 to d2 -GA111 but not to d2 -GA58 , whereas transformants converted applied d2 -GA9 to d2 -GA58 ; (iv) GA111 is converted poorly to GA70 by GA 20-oxidases in vitro when GA12 is efficiently metabolized to GA9 ; (v) no GA58 was detected endogenously in MdDOX-Co OE transformants. Overall, we conclude that 12-hydroxylation of GA12 by MdDOX-Co prevents the biosynthesis of biologically active GAs in planta, resulting in columnar phenotypes.

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.

Interaction of BTB-TAZ protein MdBT2 and DELLA protein MdRGL3a regulates nitrate-mediated plant growth.

Nitrate acts as a vital signal molecule in the modulation of plant growth and development. The phytohormones gibberellin (GA) is also involved in this process. However, the exact molecular mechanism of how nitrate and GA signaling pathway work together in regulating plant growth remains poorly understood. In this study, we found that a nitrate-responsive BTB/TAZ protein MdBT2 participates in regulating nitrate-induced plant growth in apple (Malus × domestica). Yeast two-hybridization, protein pull-down, and bimolecular fluorescence complementation (BiFC) assays showed that MdBT2 interacts with a DELLA protein MdRGL3a, which is required for the ubiquitination and degradation of MdRGL3a proteins via a 26S proteasome-dependent pathway. Furthermore, heterologous expression of MdBT2 partially rescued growth inhibition caused by overexpression of MdRGL3a in Arabidopsis. Taken together, our findings indicate that MdBT2 promotes nitrate-induced plant growth partially through reducing the abundance of the DELLA protein MdRGL3a.

A multifaceted module of BRI1 ETHYLMETHANE SULFONATE SUPRESSOR1 (BES1)-MYB88 in growth and stress tolerance of apple.

The R2R3 transcription factor MdMYB88 has previously been reported to function in biotic and abiotic stress responses. Here, we identify BRI1 ETHYLMETHANE SULFONATE SUPRESSOR1 (MdBES1), a vital component of brassinosteroid (BR) signaling in apple (Malus × domestica) that directly binds to the MdMYB88 promoter, regulating the expression of MdMYB88 in a dynamic and multifaceted mode. MdBES1 positively regulated expression of MdMYB88 under cold stress and pathogen attack, but negatively regulated its expression under control and drought conditions. Consistently, MdBES1 was a positive regulator for cold tolerance and disease resistance in apple, but a negative regulator for drought tolerance. In addition, MdMYB88 participated in BR biosynthesis by directly regulating the BR biosynthetic genes DE ETIOLATED 2 (MdDET2), DWARF 4 (MdDWF4), and BRASSINOSTEROID 6 OXIDASE 2 (MdBR6OX2). Applying exogenous BR partially rescued the erect leaf and dwarf phenotypes, as well as defects in stress tolerance in MdMYB88/124 RNAi plants. Moreover, knockdown of MdMYB88 in MdBES1 overexpression (OE) plants decreased resistance to a pathogen and C-REPEAT BINDING FACTOR1 expression, whereas overexpressing MdMYB88 in MdBES1 OE plants increased expression of SQUAMOSA PROMOTER BINDING PROTEIN-LIKE 3 （MdSPL3） and BR biosynthetic genes, suggesting that MdMYB88 contributes to MdBES1 function during BR biosynthesis and the stress response. Taken together, our results reveal multifaceted regulation of MdBES1 on MdMYB88 in BR biosynthesis and stress tolerance.

RNA-Seq profiling reveals the plant hormones and molecular mechanisms stimulating the early ripening in apple.

Fruit development and ripening are essential components of human and animal diets. Fruit ripening is also a vital plant trait for plant shelf life at the commercial level. In the present study, two apple cultivars, Hanfu wild (HC) and Hanfu mutant (HM), were employed for RNA-Sequencing (RNA-Seq) to explore the genes involved in fruit ripening. We retrieved 2642 genes, differentially expressed in HC and HM apple cultivars. Gene ontology (GO) analysis revealed the 569 categories, significantly enriched in biological process, cellular component, and molecular function. KEGG analysis exhibited the plant hormone transduction and flavonoid-anthocyanin biosynthesis pathways, might be involved in the fruit ripening and anthocyanin biosynthesis mechanism. A cluster of 13 and 26 DEGs was retrieved, representing the plant hormones and transcription factors, respectively, that may be important for early ripening in HM genotype. This transcriptome study would be useful for researchers to functionally characterize the DEGs responsible for early ripening.

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.

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.

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.

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.

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.

ERF4 affects fruit firmness through TPL4 by reducing ethylene production.

The firmness of fleshy fruit crops has a significant effect on their quality, consumer preference, shelf life and transportability. In a combined quantitative trait locus and genome-wide association studies study of apple fruit texture, we identified a mutation (C-G) in the ethylene response factor-associated amphiphilic repression (EAR) motif in the coding region of the apple ETHYLENE RESPONSE FACTOR4 (ERF4) gene. Chromatin immunoprecipitation sequencing showed that ERF4 binds to the promoter of ERF3, which is involved in regulation of ethylene biosynthesis. The EAR mutation in ERF4 results in reduced repression of ERF3 expression, which is turn promotes ethylene production and loss of fruit firmness. ERF4 acts as a transcriptional repressor whose activity is modulated by a TOPLESS co-repressor 4 (TPL4)-binding EAR repression motif. Biolayer interferometry analysis showed that the mutation in the EAR motif causes a reduction in the interaction with TPL4. Suppression of ERF4 or TPL4 promoted fruit ripening and ethylene production. Taken together, our results provide insights into how ERF4 allelic variation underlies an important fruit quality trait.