Multi-omics analysis reveals the biosynthesis of flavonoids during the browning process of Malus sieversii explants.

Malus sieversii is a precious apple germplasm resource. Browning of explants is one of the most important factors limiting the survival rate of plant tissue culture. In order to explore the molecular mechanism of the browning degree of different strains of Malus sieversii, we compared the dynamic changes of Malus sieversii and Malus robusta Rehd. during the whole browning process using a multi-group method. A total of 44 048 differentially expressed genes (DEGs) were identified by transcriptome analysis on the DNBSEQ-T7 sequencing platform. KEGG enrichment analysis showed that the DEGs were significantly enriched in the flavonoid biosynthesis pathway. In addition, metabonomic analysis showed that (-)-epicatechin, astragalin, chrysin, irigenin, isoquercitrin, naringenin, neobavaisoflavone and prunin exhibited different degrees of free radical scavenging ability in the tissue culture browning process, and their accumulation in different varieties led to differences in the browning degree among varieties. Comprehensive transcriptome and metabonomics analysis of the data related to flavonoid biosynthesis showed that PAL, 4CL, F3H, CYP73A, CHS, CHI, ANS, DFR and PGT1 were the key genes for flavonoid accumulation during browning. In addition, WGCNA analysis revealed a strong correlation between the known flavonoid structure genes and the selected transcriptional genes. Protein interaction predictions demonstrated that 19 transcription factors (7 MYBs and 12 bHLHs) and 8 flavonoid structural genes had targeted relationships. The results show that the interspecific differential expression of flavonoid genes is the key influencing factor of the difference in browning degree between Malus sieversii and Malus robusta Rehd., providing a theoretical basis for further study on the regulation of flavonoid biosynthesis.

Complete chloroplast genome of the Malus baccata var. gracilis provides insights into the evolution and phylogeny of Malus species.

Malus baccata (L.) var. gracilis (Rehd.) has high ornamental value and breeding significance, and comparative chloroplast genome analysis was applied to facilitate genetic breeding for desired traits and resistance and provide insight into the phylogeny of this genus. Using data from whole-genome sequencing, a tetrameric chloroplast genome with a length of 159,992 bp and a total GC content of 36.56% was constructed. The M. baccata var. gracilis chloroplast genome consists of a large single-copy sequence (88,100 bp), a short single-copy region (19,186 bp), and two inverted repeat regions, IRa (26,353 bp) and IRb (26,353 bp). This chloroplast genome contains 112 annotated genes, including 79 protein-coding genes (nine multicopy), 29 tRNA genes (eight multicopy), and four rRNA genes (all multicopy). Calculating the relative synonymous codon usage revealed a total of 32 high-frequency codons, and the codons exhibited a biased usage pattern towards A/U as the ending nucleotide. Interspecific sequence comparison and boundary analysis revealed significant sequence variation in the vast single-copy region, as well as generally similar expansion and contraction of the SSC and IR regions for 10 analyzed Malus species. M. baccata var. gracilis and Malus hupehensis were grouped together into one branch based on phylogenetic analysis of chloroplast genome sequences. The chloroplast genome of Malus species provides an important foundation for species identification, genetic diversity analysis, and Malus chloroplast genetic engineering. Additionally, the results can facilitate the use of pendant traits to improve apple tree shape.

The regulatory role of MdNAC14-Like in anthocyanin synthesis and proanthocyanidin accumulation in red-fleshed apples.

Flavonoids, such as anthocyanins and proanthocyanidins (PAs), play essential roles in plant growth, development, and stress response. Red-fleshed apples represent a valuable germplasm resource with high flavonoid content. Understanding and enriching the regulatory network controlling flavonoid synthesis in red-fleshed apples holds significant importance for cultivating high-quality fruits. In this study, we successfully isolated an NAC transcription factor, MdNAC14-Like, which exhibited a significant negative correlation with the content of anthocyanin. Transient injection of apple fruit and stable expression of callus confirmed that MdNAC14-Like acts as an inhibitor of anthocyanin synthesis. Through yeast monohybrid, electrophoretic mobility shift, and luciferase reporter assays, we demonstrated the ability of MdNAC14-Like to bind to the promoters of MdMYB9, MdMYB10, and MdUFGT, thus inhibiting their transcriptional activity and subsequently suppressing anthocyanin synthesis. Furthermore, our investigation revealed that MdNAC14-Like interacts with MdMYB12, enhancing the transcriptional activation of MdMYB12 on the downstream structural gene MdLAR, thereby promoting PA synthesis. This comprehensive functional characterization of MdNAC14-Like provides valuable insights into the intricate regulatory network governing anthocyanin and PA synthesis in apple.

Transcription factors MdMYC2 and MdMYB85 interact with ester aroma synthesis gene MdAAT1 in apple.

Volatile esters in apple (Malus domestica) fruit are the critical aroma components determining apple flavor quality. While the exact molecular regulatory mechanism remains unknown, jasmonic acid (JA) plays a crucial role in stimulating the synthesis of ester aromas in apples. In our study, we investigated the effects of methyl jasmonate (MeJA) on the production of ester aroma in apples. MeJA treatment significantly increased ester aroma synthesis, accompanied by the upregulation of several genes involved in the jasmonate pathway transduction. Specifically, expression of the gene MdMYC2, which encodes a transcription factor associated with the jasmonate pathway, and the R2R3-MYB transcription factor gene MdMYB85 increased upon MeJA treatment. Furthermore, the essential gene ALCOHOL ACYLTRANSFERASE 1 (MdAAT1), encoding an enzyme responsible for ester aroma synthesis, showed increased expression levels as well. Our investigation revealed that MdMYC2 and MdMYB85 directly interacted with the promoter region of MdAAT1, thereby enhancing its transcriptional activity. In addition, MdMYC2 and MdMYB85 directly bind their promoters and activate transcription. Notably, the interaction between MdMYC2 and MdMYB85 proteins further amplified the regulatory effect of MdMYB85 on MdMYC2 and MdAAT1, as well as that of MdMYC2 on MdMYB85 and MdAAT1. Collectively, our findings elucidate the role of the gene module consisting of MdMYC2, MdMYB85, and MdAAT1 in mediating the effects of JA and promoting ester aroma synthesis in apples.

Remediation of the microecological environment of heavy metal-contaminated soil with fulvic acid, improves the quality and yield of apple.

The excessive application of chemical fertilizers and pesticides in apple orchards is responsible for high levels of manganese and copper in soil, and this poses a serious threat to soil health. We conducted a three-year field experiment to study the remediation effect and mechanism of fulvic acid on soil with excess manganese and copper. The exogenous application of fulvic acid significantly reduced the content of manganese and copper in soil and plants; increased the content of calcium; promoted the growth of apple plants; improved the fruit quality and yield of apple; increased the content of chlorophyll; increased the activity of superoxide dismutase, peroxidase, and catalase; and reduced the content of malondialdehyde. The number of soil culturable microorganisms, soil enzyme activity, soil microbial community diversity, and relative abundance of functional bacteria were increased, and the detoxification of the glutathione metabolism function was enhanced. The results of this study provide new insights that will aid the remediation of soil with excess manganese and copper using fulvic acid.

Transcriptome analysis reveals that PbMYB61 and PbMYB308 are involved in the regulation of lignin biosynthesis in pear fruit stone cells.

Pear fruit stone cells have thick walls and are formed by the secondary deposition of lignin in the primary cell wall of thin-walled cells. Their content and size seriously affect fruit characteristics related to edibility. To reveal the regulatory mechanism underlying stone cell formation during pear fruit development and to identify hub genes, we examined the stone cell and lignin contents of 30 'Shannongsu' pear flesh samples and analyzed the transcriptomes of 15 pear flesh samples collected at five developmental stages. On the basis of the RNA-seq data, 35 874 differentially expressed genes were detected. Additionally, two stone cell-related modules were identified according to a WGCNA. A total of 42 lignin-related structural genes were subsequently obtained. Furthermore, nine hub structural genes were identified in the lignin regulatory network. We also identified PbMYB61 and PbMYB308 as candidate transcriptional regulators of stone cell formation after analyzing co-expression networks and phylogenetic relationships. Finally, we experimentally validated and characterized the candidate transcription factors and revealed that PbMYB61 regulates stone cell lignin formation by binding to the AC element in the PbLAC1 promoter to upregulate expression. However, PbMYB308 negatively regulates stone cell lignin synthesis by binding to PbMYB61 to form a dimer that cannot activate PbLAC1 expression. In this study, we explored the lignin synthesis-related functions of MYB family members. The results presented herein are useful for elucidating the complex mechanisms underlying lignin biosynthesis during pear fruit stone cell development.

The ABA-induced NAC transcription factor MdNAC1 interacts with a bZIP-type transcription factor to promote anthocyanin synthesis in red-fleshed apples.

Anthocyanins are valuable compounds in red-fleshed apples. The MdMYB10 transcription factor is an important regulator of the anthocyanin synthesis pathway. However, other transcription factors are key components of the complex network controlling anthocyanin synthesis and should be more thoroughly characterized. In this study, we used a yeast-based screening technology to identify MdNAC1 as a transcription factor that positively regulates anthocyanin synthesis. The overexpression of MdNAC1 in apple fruits and calli significantly promoted the accumulation of anthocyanins. In binding experiments, we demonstrated that MdNAC1 combines with the bZIP-type transcription factor MdbZIP23 to activate the transcription of MdMYB10 and MdUFGT. Our analyses also indicated that the expression of MdNAC1 is strongly induced by ABA because of the presence of an ABRE cis-acting element in its promoter. Additionally, the accumulation of anthocyanins in apple calli co-transformed with MdNAC1 and MdbZIP23 increased in the presence of ABA. Therefore, we revealed a novel anthocyanin synthesis mechanism involving the ABA-induced transcription factor MdNAC1 in red-fleshed apples.

Effect of Hydrogen Peroxide on the Soil Microbial Community Structure and Growth of Malus hupehensis Rehd. Seedlings under Replant Conditions.

Apple replant disease (ARD) is common in apple production, which seriously affects the growth and development of apples. In this study, hydrogen peroxide with a bactericidal effect was used to treat the replanted soil, and the effects of different concentrations of hydrogen peroxide on replanted seedlings and soil microbiology were investigated in order to seek a green, clean way to control ARD. Five treatments were set up in this study: replanted soil (CK1), replanted soil with methyl bromide fumigation (CK2), replanted soil + 1.5% hydrogen peroxide (H1), replanted soil + 3.0% hydrogen peroxide (H2), and replanted soil + 4.5% hydrogen peroxide (H3). The results showed that hydrogen peroxide treatment improved replanted seedling growth and also inactivated a certain number of Fusarium, while the Bacillus, Mortierella, and Guehomyces also became more abundant in relative terms. The best results were obtained with replanted soil + 4.5% hydrogen peroxide (H3). Consequently, hydrogen peroxide applied to the soil can effectively prevent and control ARD.

Abscisic acid and regulation of the sugar transporter gene MdSWEET9b promote apple sugar accumulation.

Enhancing fruit sugar contents, especially for high-flavonoid apples with a sour taste, is one of the main goals of horticultural crop breeders. This study analyzed sugar accumulation and the underlying mechanisms in the F2 progenies of a hybridization between the high-sugar apple (Malus × domestica) variety "Gala" and high-flavonoid apple germplasm "CSR6R6". We revealed that MdSWEET9b (sugars will eventually be exported transporter) helps mediate sugar accumulation in fruits. Functional characterization of MdSWEET9b in yeast mutants lacking sugar transport as well as in overexpressing and CRISPR/Cas9 knockdown apple calli revealed MdSWEET9b could transport sucrose specifically, ultimately promoting normal yeast growth and accumulation of total sugar contents. Moreover, MdWRKY9 bound to the MdSWEET9b promoter and regulated its activity, which responded to abscisic acid (ABA) signaling. Furthermore, MdWRKY9 interacted with MdbZIP23 (basic leucine zipper) and MdbZIP46, key ABA signal transducers, at the protein and DNA levels to enhance its regulatory effect on MdSWEET9b expression, thereby influencing sugar accumulation. Based on the contents of ABA in lines with differing sugar contents and the effects of ABA treatments on fruits and calli, we revealed ABA as one of the main factors responsible for the diversity in apple fruit sugar content. The results of this study have clarified how MdSWEET9b influences fruit sugar accumulation, while also further elucidating the regulatory effects of the ABA-signaling network on fruit sugar accumulation. This work provides a basis for future explorations of the crosstalk between hormone and sugar metabolism pathways.

MdMYB305-MdbHLH33-MdMYB10 regulates sugar and anthocyanin balance in red-fleshed apple fruits.

Sugar and anthocyanin are important indicators of fruit quality, and understanding the mechanism underlying their accumulation is essential for breeding high-quality fruit. We identified an R2R3-MYB transcription factor MdMYB305 in the red-fleshed apple progeny, which was positively correlated with fruit sugar content but negatively correlated with anthocyanin content. Transient injection, stable expression [overexpressing and clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9)], and heterologous transformation of tomato confirmed that MdMYB305 promotes the accumulation of sugar and inhibits the synthesis of anthocyanin. A series of molecular experiments (such as electrophoretic mobility shift and luciferase assays) confirmed that MdMYB305 combines with sugar-related genes (MdCWI1/MdVGT3/MdTMT2) and anthocyanin-related genes (MdF3H/MdDFR/MdUFGT), promoting and inhibiting their activities, and finally regulating the sugar and anthocyanin content of fruits. In addition, the study also found that MdMYB305 competes with MdMYB10 for the MdbHLH33 binding site to balance sugar and anthocyanin accumulation in the fruits, which provides a reference value for exploring more functions of the MYB-bHLH-MYB complex and the balance relationship between sugar and anthocyanin in the future.

Mdm-miR858 targets MdMYB9 and MdMYBPA1 to participate anthocyanin biosynthesis in red-fleshed apple.

Anthocyanins are important secondary metabolites in plants. They are important for human health because of their antioxidant activities and because their dietary intake reduces the incidence of cardiovascular and cerebrovascular diseases and tumors. The biosynthesis of anthocyanins and its regulation in fruits and vegetables is a global research hotspot. Compared with cultivated apples, the red-fleshed apple is a relatively new and popular commodity in the market. Previous studies on red-fleshed apples have focused on the basis for the high anthocyanin content and the transcriptional regulation of anthocyanin synthesis. In the present study, we focused on the mechanism of microRNA-mediated post-transcriptional regulation of anthocyanin synthesis in red-fleshed apples. We identified a microRNA (miRNA), designated mdm-miR858, that is specifically expressed in the flesh of apple fruit. The expression level of miR858 was significantly lower in red-fleshed apples than in white-fleshed apples. The overexpression of mdm-miR858 significantly inhibited anthocyanin accumulation, whereas the silencing of mdm-miR858 promoted anthocyanin synthesis in STTM858 transgenic apple calli. Further analyses showed that mdm-miR858 targets the transcription factor genes MdMYB9 and MdMYBPA1 to participate anthocyanin accumulation in apple. Our results also show that MdHY5, a transcription factor in the light signaling pathway, can bind to the promoter of mdm-miR858 to inhibit its transcription, thereby regulating anthocyanin synthesis. Based on our results, we describe a novel HY5-miR858-MYB loop involved in the modulation of anthocyanin biosynthesis. These findings provide new information about how plant miRNAs regulate anthocyanin anabolism and provide a basis for breeding new anthocyanin-rich, red-fleshed apple varieties.

The regulatory module MdBZR1-MdCOL6 mediates brassinosteroid- and light-regulated anthocyanin synthesis in apple.

The anthocyanin content is an important indicator of the nutritional value of most fruits, including apple (Malus domestica). Anthocyanin synthesis is coordinately regulated by light and various phytohormones. In this study on apple, we revealed the antagonistic relationship between light and brassinosteroid (BR) signaling pathways, which is mediated by BRASSINAZOLE-RESISTANT 1 (MdBZR1) and the B-box protein MdCOL6. The exogenous application of brassinolide inhibited the high-light-induced anthocyanin accumulation in red-fleshed apple seedlings, whereas increases in the light intensity decreased the endogenous BR content. The overexpression of MdBZR1 inhibited the anthocyanin synthesis in apple plants. An exposure to a high-light intensity induced the degradation of dephosphorylated MdBZR1, resulting in functional impairment. MdBZR1 was identified as an upstream repressor of MdCOL6, which promotes anthocyanin synthesis in apple plants. Furthermore, MdBZR1 interacts with MdCOL6 to attenuate its ability to activate MdUFGT and MdANS transcription. Thus, MdBZR1 negatively regulates MdCOL6-mediated anthocyanin accumulation. Our study findings have clarified the molecular basis of the integration of light and BR signals during the regulation of anthocyanin biosynthesis, which is an important process influencing fruit quality.

Effects of Organic Acid Root Exudates of Malus hupehensis Rehd. Derived from Soil and Root Leaching Liquor from Orchards with Apple Replant Disease.

Organic acids secreted by plants, such as p-hydroxybenzoic acid, ferulic acid, cinnamic acid, and benzoic acid, can inhibit seed germination and root growth. The effects of root and soil leaching liquor from orchards on the growth of M. hupehensis Rehd. seedlings under sand culture are studied; the seedlings are sampled at 15, 30, 45, and 60 d. Changes in the amount of root exudates are determined using HPLC. Low concentrations of root leaching liquor (A1) and soil leaching liquor (B1) significantly promoted plant growth and chlorophyll synthesis; high concentrations of root leaching liquor (A6) and soil leaching liquor (B4-6) inhibited growth. Low concentrations of soil leaching liquor had no significant effect on the POD, SOD, and CAT activities. A5-6 and B5-6 significantly decreased Fv/Fm and qP values, respectively, and increased NPQ values. All root and soil leaching liquor treatments inhibited the secretion of gallic acid, hydroxybenzoic acid, benzoic acid, and phloridzin, and promoted the secretion of caffeic acid. The root leaching liquor treatments inhibited the secretion of catechin and promoted the secretion of phloretin. The soil leaching liquor treatments promoted the secretion of cinnamic acid. The secretion of other phenolic acids is likely associated with the different concentrations of leaching liquor.

Allium fistulosum L. Alleviates Apple Replant Disease by Suppressing Fusarium solani.

Fusarium solani has often been isolated from replanted apple roots, suggesting that it is associated with apple replant disease. The mechanism underlying the ability of the mixed cropping of apple trees with Allium fistulosum L. to alleviate apple replant disease remains unclear. The aim of this study was to determine the pathogenicity of the Fusarium solani isolate HBH 08 isolated from diseased roots and the effect of A. fistulosum L. and its root secretions on Fusarium solani isolate HBH 08 and apple seedings. The field experiment showed that A. fistulosum L. not only significantly reduced the amount of the Fusarium solani isolate HBH 08 in replanted soil but also increased the biomass of the grafted apple seedlings. The GC-MS analysis indicated that dimethyl disulphide and diallyl disulphide were active molecules in the root exudates of A. fistulosum L. They inhibited the growth of the Fusarium solani isolate HBH 08 mycelium and decreased the number of spores germinated. In addition, these compounds reduced the amount of the Fusarium solani isolate HBH 08 under replanted conditions and promoted the growth of grafted apple seedlings. Overall, mixed cropping with A. fistulosum L. might be an effective approach for cultivating apple trees and controlling apple replant disease.

MdERF1B-MdMYC2 module integrates ethylene and jasmonic acid to regulate the biosynthesis of anthocyanin in apple.

Ethylene and jasmonic acid (JA) are crucial hormones that promote anthocyanin synthesis in apple (Malus × domestica). However, the mechanism by which these hormones cooperate to modulate anthocyanin production in apple is unclear. According to our results, MdERF1B expression was strongly induced by ethylene and JA. Physiological phenotypes and the results of molecular biological analyses indicated that MdERF1B encodes a positive regulator of anthocyanin synthesis. Specifically, MdERF1B was capable of combining directly with the MdMYC2 promoter to promote gene expression. Additionally, MdERF1B interacted with two JA signaling pathway inhibitors, namely MdJAZ5 and MdJAZ10. The MdERF1B-MdJAZ5/10 protein complex decreased the ability of MdERF1B to activate the MdMYC2 promoter. Furthermore, MdEIL1, which is a crucial protein for ethylene signal transduction, was observed to bind directly to the MdERF1B promoter, thereby upregulating gene expression. These results suggest that MdERF1B is a core gene responsive to JA and ethylene signals. The encoded protein, together with MdMYC2, MdJAZ5/10, and MdEIL1, modulates anthocyanin synthesis in apple. This study clarifies the synergistic mechanism by which JA and ethylene regulate anthocyanin production in apple.

Study of the grafting compatibility of the apple rootstock 12-2, resistant to apple replant diseases (ARD).

BACKGROUND: Cultivation of resistant rootstocks can effectively prevent apple replant disease (ARD), and grafting tests are an important means of evaluating the compatibility of rootstocks with scions. METHODS: The apple rootstocks 12-2 (self-named) and Malus hupehensis Rehd. (PYTC) were planted in a replanted 20-year-old apple orchard. The two rootstocks were grafted with scions of 13 apple varieties. Multiple aboveground physiological parameters of the grafted combinations were measured and evaluated to verify the grafting affinity of 12-2 with the scions as compared to Malus hupehensis Rehd. (PYTC). RESULTS: The graft survival rate and graft interface healing of 12-2 did not differ significantly from those of PYTC. Mechanical strength tests of the grafted interfaces showed that some mechanical strength indices of Redchief, Jonagold, Starking, Goldspur and Yinv apple varieties were significantly higher when they were grafted onto 12-2 compared to the PYTC control. The height and diameter of shoots and the relative chlorophyll content, photosynthetic and fluorescence parameters, antioxidant enzyme activities and malondialdehyde content of leaves showed that Fuji 2001, Tengmu No.1, RedChief, Gala, USA8, and Shoufu1 grew similarly on the two rootstocks, but Tianhong 2, Lvguang, Jonagold, Starking, Goldspur, Yinv and Luli grew better when grafted onto 12-2 than onto the PYTC control. The rootstock 12-2, therefore, showed good grafting affinity. CONCLUSION: These results provide experimental materials and theoretical guidance for the cultivation of a new grafting compatible rootstock to the 13 studied apple cultivars.

Research Progress on Genetic Basis of Fruit Quality Traits in Apple (Malus × domestica).

Identifying the genetic variation characteristics of phenotypic traits is important for fruit tree breeding. During the long-term evolution of fruit trees, gene recombination and natural mutation have resulted in a high degree of heterozygosity. Apple (Malus × domestica Borkh.) shows strong ecological adaptability and is widely cultivated, and is among the most economically important fruit crops worldwide. However, the high level of heterozygosity and large genome of apple, in combination with its perennial life history and long juvenile phase, complicate investigation of the genetic basis of fruit quality traits. With continuing augmentation in the apple genomic resources available, in recent years important progress has been achieved in research on the genetic variation of fruit quality traits. This review focuses on summarizing recent genetic studies on apple fruit quality traits, including appearance, flavor, nutritional, ripening, and storage qualities. In addition, we discuss the mapping of quantitative trait loci, screening of molecular markers, and mining of major genes associated with fruit quality traits. The overall aim of this review is to provide valuable insights into the mechanisms of genetic variation and molecular breeding of important fruit quality traits in apple.

Chemical fumigants control apple replant disease: Microbial community structure-mediated inhibition of Fusarium and degradation of phenolic acids.

Fusarium and phenolic acids in apple replant soil have deleterious effects on soil, which affects the growth of young replanted apple trees. Here, we studied the effects of different chemical fumigants (metham sodium, dazomet, calcium cyanamide, 1,3-dichloropropene, and methyl bromide) on Fusarium and phenolic acids in soil. The chemical fumigants disturbed the apple replant soil microbial community to different degrees in the order from highest to the lowest as methyl bromide > 1,3-dichloropropene > dazomet > metham sodium > calcium cyanamide. Compared with the control, the total numbers of Operational Taxonomic Unit (OTU) were 104.63 % and 9.38 % lower in the methyl bromide and calcium cyanamide treatments, respectively while the average contents of Fusarium were 88.04 % and 59.18% lower in these treatments, respectively. Higher disturbance degrees resulted in a slower recovery rate of the soil microbial community, which facilitated the transformation of the soil into a disease-suppressing state. During the recovery process, the roots recruited Streptomyces OTU2796 and Bacillus OTU2243, which alleviated Fusarium-induced stress via the synthesis of polyketones and macrolides. The roots also recruited Sphingomonas OTU3488, OTU5572, and OTU8147, which alleviated phenolic acid-induced stress through the degradation of benzoate and polycyclic aromatic hydrocarbons.

Involvement of MdWRKY40 in the defense of mycorrhizal apple against fusarium solani.

BACKGROUND: Apple (Malus domestica Borkh.) is an important economic crop. The pathological effects of Fusarium solani, a species complex of soilborne pathogens, on the root systems of apple plants was unknown. It was unclear how mycorrhizal apple seedlings resist infection by F. solani. The transcriptional profiles of mycorrhizal and non-mycorrhizal plants infected by F. solani were compared using RNA-Seq. RESULTS: Infection with F. solani significantly reduced the dry weight of apple roots, and the roots of mycorrhizal apple plants were less damaged when the plants were infected with F. solani. They also had enhanced activity of antioxidant enzymes and a reduction in the oxidation of membrane lipids. A total of 1839 differentially expressed genes (DEGs) were obtained after mycorrhizal and non-mycorrhizal apple plants were infected with F. solani. A gene ontogeny (GO) analysis showed that most of the DEGs were involved in the binding of ADP and calcium ions. In addition, based on a MapMan analysis, a large number of DEGs were found to be involved in the response of mycorrhizal plants to stress. Among them, the overexpressed transcription factor MdWRKY40 significantly improved the resistance of the apple 'Orin' callus to F. solani and the expression of the resistance gene MdGLU by binding the promoter of MdGLU. CONCLUSION: This paper outlines how the inoculation of apple seedlings roots by arbuscular mycorrhizal fungi responded to infection with F. solani at the transcriptional level. In addition, MdWRKY40 played an important role in the resistance of mycorrhizal apple seedlings to infection with F. solani.

Transcriptome changes associated with apple (Malus domestica) root defense response after Fusarium proliferatum f. sp. malus domestica infection.

BACKGROUND: Apple replant disease is a soilborne disease caused by Fusarium proliferatum f. sp. malus domestica strain MR5 (abbreviated hereafter as Fpmd MR5) in China. This pathogen causes root tissue rot and wilting leaves in apple seedlings, leading to plant death. A comparative transcriptome analysis was conducted using the Illumina Novaseq platform to identify the molecular defense mechanisms of the susceptible M.26 and the resistant M9T337 apple rootstocks to Fpmd MR5 infection. RESULTS: Approximately 518.1 million high-quality reads were generated using RNA sequencing (RNA-seq). Comparative analysis between the mock-inoculated and Fpmd MR5 infected apple rootstocks revealed 28,196 significantly differentially expressed genes (DEGs), including 14,572 up-regulated and 13,624 down-regulated genes. Among them, the transcriptomes in the roots of the susceptible genotype M.26 were reflected by overrepresented DEGs. MapMan analysis indicated that a large number of DEGs were involved in the response of apple plants to Fpmd MR5 stress. The important functional groups identified via gene ontology (GO) and the Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment were responsible for fundamental biological regulation, secondary metabolism, plant-pathogen recognition, and plant hormone signal transduction (ethylene and jasmonate). Furthermore, the expression of 33 up-regulated candidate genes (12 related to WRKY DNA-binding proteins, one encoding endochitinase, two encoding beta-glucosidases, ten related to pathogenesis-related proteins, and eight encoding ethylene-responsive transcription factors) were validated by quantitative real-time PCR. CONCLUSION: RNA-seq profiling was performed for the first time to analyze response of apple root to Fpmd MR5 infection. We found that the production of antimicrobial compounds and antioxidants enhanced plant resistance to pathogens, and pathogenesis-related protein (PR10 homologs, chitinase, and beta-glucosidase) may play unique roles in the defense response. These results provide new insights into the mechanisms of the apple root response to Fpmd MR5 infection.