Hypermethylation in the promoter regions of flavonoid pathway genes is associated with skin color fading during 'Daihong' apple fruit development.

Apple fruit skin color fading is not well understood although the molecular mechanism of skin color formation is well known. The red-fleshed apple cultivar 'Daihong' (DH) exhibited fading skin color during fruit development despite having a heterozygous R6 allele but lacking Red-TE for red fruit skin. In this study, transcriptomic analysis revealed the expression level of MdMYB10 increased with fruit development whereas reduced expression levels of MdMYBPA1, MdCHS, MdANS, MdUFGT, MdLAR, and MdANR were observed, consistent with decreased levels of chalcone, anthocyanin, catechin, epicatechin, and procyanidin B2. Whole-genome bisulfite sequencing (WGBS) indicated a global gain in cytosine methylation levels and increased methylation in 5' and 3' flanking regions of genes and transposable elements (TEs), and in TE bodies in all CG, CHG and CHH contexts, especially the mCHH context, during fruit development. The increased DNA methylation was attributed to reduced expression levels of DNA demethylase genes, including MdDME1, MdROS1, and MdROS2. Association analysis revealed a significant negative correlation between promoter methylation levels of MdCHS, MdCHI, MdMYBPA1, and their respective transcript levels, as well as a negative correlation between promoter methylation levels of MdCHS, MdCHI, MdANR, and MdFLS, and the content of chalcones, naringenin-7-glucoside, epicatechin, and quercetin. Treatment with the DNA demethylation agent 5-aza-2'-deoxycytidine verified the negative correlation between DNA methylation and gene expression within the flavonoid pathway. These findings suggest that hypermethylation in promoter regions of genes of the flavonoid biosynthesis pathway is associated with the reduction of gene expression and flavonoid content, and fruit skin color fading during DH apple development.

PbHsfC1a-coordinates ABA biosynthesis and H2O2 signalling pathways to improve drought tolerance in Pyrus betulaefolia.

Abiotic stresses have had a substantial impact on fruit crop output and quality. Plants have evolved an efficient immune system to combat abiotic stress, which employs reactive oxygen species (ROS) to activate the downstream defence response signals. Although an aquaporin protein encoded by PbPIP1;4 is identified from transcriptome analysis of Pyrus betulaefolia plants under drought treatments, little attention has been paid to the role of PIP and ROS in responding to abiotic stresses in pear plants. In this study, we discovered that overexpression of PbPIP1;4 in pear callus improved tolerance to oxidative and osmotic stresses by reconstructing redox homeostasis and ABA signal pathways. PbPIP1;4 overexpression enhanced the transport of H2O2 into pear and yeast cells. Overexpression of PbPIP1;4 in Arabidopsis plants mitigates the stress effects caused by adding ABA, including stomatal closure and reduction of seed germination and seedling growth. Overexpression of PbPIP1;4 in Arabidopsis plants decreases drought-induced leaf withering. The PbPIP1;4 promoter could be bound and activated by TF PbHsfC1a. Overexpression of PbHsfC1a in Arabidopsis plants rescued the leaf from wilting under drought stress. PbHsfC1a could bind to and activate AtNCED4 and PbNCED4 promoters, but the activation could be inhibited by adding ABA. Besides, PbNCED expression was up-regulated under H2O2 treatment but down-regulated under ABA treatment. In conclusion, this study revealed that PbHsfC1a is a positive regulator of abiotic stress, by targeting PbPIP1;4 and PbNCED4 promoters and activating their expression to mediate redox homeostasis and ABA biosynthesis. It provides valuable information for breeding drought-resistant pear cultivars through gene modification.

Two adjacent NAC transcription factors regulate fruit maturity date and flavor in peach.

Although maturity date (MD) is an essential factor affecting fresh fruit marketing and has a pleiotropic effect on fruit taste qualities, the underlying mechanisms remain largely unclear. In this study, we functionally characterized two adjacent NAM-ATAF1/2-CUC2 (NAC) transcription factors (TFs), PpNAC1 and PpNAC5, both of which were associated with fruit MD in peach. PpNAC1 and PpNAC5 were found capable of activating transcription of genes associated with cell elongation, cell wall degradation and ethylene biosynthesis, suggesting their regulatory roles in fruit enlargement and ripening. Furthermore, PpNAC1 and PpNAC5 had pleiotropic effects on fruit taste due to their ability to activate transcription of genes for sugar accumulation and organic acid degradation. Interestingly, both PpNAC1 and PpNAC5 orthologues were found in fruit-producing angiosperms and adjacently arranged in all 91 tested dicots but absent in fruitless gymnosperms, suggesting their important roles in fruit development. Our results provide insight into the regulatory roles of NAC TFs in MD and fruit taste.

MADS-box protein PpDAM6 regulates chilling requirement-mediated dormancy and bud break in peach.

Bud dormancy is crucial for winter survival and is characterized by the inability of the bud meristem to respond to growth-promotive signals before the chilling requirement (CR) is met. However, our understanding of the genetic mechanism regulating CR and bud dormancy remains limited. This study identified PpDAM6 (DORMANCY-ASSOCIATED MADS-box) as a key gene for CR using a genome-wide association study analysis based on structural variations in 345 peach (Prunus persica (L.) Batsch) accessions. The function of PpDAM6 in CR regulation was demonstrated by transiently silencing the gene in peach buds and stably overexpressing the gene in transgenic apple (Malus × domestica) plants. The results showed an evolutionarily conserved function of PpDAM6 in regulating bud dormancy release, followed by vegetative growth and flowering, in peach and apple. The 30-bp deletion in the PpDAM6 promoter was substantially associated with reducing PpDAM6 expression in low-CR accessions. A PCR marker based on the 30-bp indel was developed to distinguish peach plants with non-low and low CR. Modification of the H3K27me3 marker at the PpDAM6 locus showed no apparent change across the dormancy process in low- and non-low- CR cultivars. Additionally, H3K27me3 modification occurred earlier in low-CR cultivars on a genome-wide scale. PpDAM6 could mediate cell-cell communication by inducing the expression of the downstream genes PpNCED1 (9-cis-epoxycarotenoid dioxygenase 1), encoding a key enzyme for ABA biosynthesis, and CALS (CALLOSE SYNTHASE), encoding callose synthase. We shed light on a gene regulatory network formed by PpDAM6-containing complexes that mediate CR underlying dormancy and bud break in peach. A better understanding of the genetic basis for natural variations of CR can help breeders develop cultivars with different CR for growing in different geographical regions.

The transcription factor PbrMYB24 regulates lignin and cellulose biosynthesis in stone cells of pear fruits.

Lignified stone cell content is a key factor used to evaluate fruit quality, influencing the economic value of pear (Pyrus pyrifolia) fruits. However, our understanding of the regulatory networks of stone cell formation is limited due to the complex secondary metabolic pathway. In this study, we used a combination of co-expression network analysis, gene expression profiles, and transcriptome analysis in different pear cultivars with varied stone cell content to identify a hub MYB gene, PbrMYB24. The relative expression of PbrMYB24 in fruit flesh was significantly correlated with the contents of stone cells, lignin, and cellulose. We then verified the function of PbrMYB24 in regulating lignin and cellulose formation via genetic transformation in homologous and heterologous systems. We constructed a high-efficiency verification system for lignin and cellulose biosynthesis genes in pear callus. PbrMYB24 transcriptionally activated multiple target genes involved in stone cell formation. On the one hand, PbrMYB24 activated the transcription of lignin and cellulose biosynthesis genes by binding to different cis-elements [AC-I (ACCTACC) element, AC-II (ACCAACC) element and MYB-binding sites (MBS)]. On the other hand, PbrMYB24 bound directly to the promoters of PbrMYB169 and NAC STONE CELL PROMOTING FACTOR (PbrNSC), activating the gene expression. Moreover, both PbrMYB169 and PbrNSC activated the promoter of PbrMYB24, enhancing gene expression. This study improves our understanding of lignin and cellulose synthesis regulation in pear fruits through identifying a regulator and establishing a regulatory network. This knowledge will be useful for reducing the stone cell content in pears via molecular breeding.

Auxin inhibits lignin and cellulose biosynthesis in stone cells of pear fruit via the PbrARF13-PbrNSC-PbrMYB132 transcriptional regulatory cascade.

Stone cells are often present in pear fruit, and they can seriously affect the fruit quality when present in large numbers. The plant growth regulator NAA, a synthetic auxin, is known to play an active role in fruit development regulation. However, the genetic mechanisms of NAA regulation of stone cell formation are still unclear. Here, we demonstrated that exogenous application of 200 µM NAA reduced stone cell content and also significantly decreased the expression level of PbrNSC encoding a transcriptional regulator. PbrNSC was shown to bind to an auxin response factor, PbrARF13. Overexpression of PbrARF13 decreased stone cell content in pear fruit and secondary cell wall (SCW) thickness in transgenic Arabidopsis plants. In contrast, knocking down PbrARF13 expression using virus-induced gene silencing had the opposite effect. PbrARF13 was subsequently shown to inhibit PbrNSC expression by directly binding to its promoter, and further to reduce stone cell content. Furthermore, PbrNSC was identified as a positive regulator of PbrMYB132 through analyses of co-expression network of stone cell formation-related genes. PbrMYB132 activated the expression of gene encoding cellulose synthase (PbrCESA4b/7a/8a) and lignin laccase (PbrLAC5) binding to their promotors. As expected, overexpression or knockdown of PbrMYB132 increased or decreased stone cell content in pear fruit and SCW thickness in Arabidopsis transgenic plants. In conclusion, our study shows that the 'PbrARF13-PbrNSC-PbrMYB132' regulatory cascade mediates the biosynthesis of lignin and cellulose in stone cells of pear fruit in response to auxin signals and also provides new insights into plant SCW formation.

Self S-RNase inhibits ABF-LRX signaling to arrest pollen tube growth to achieve self-incompatibility in pear.

Gametophytic self-incompatibility (GSI) has been widely studied in flowering plants, but studies of the mechanisms underlying pollen tube growth arrest by self S-RNase in GSI species are limited. In the present study, two leucine-rich repeat extensin genes in pear (Pyrus bretschneideri), PbLRXA2.1 and PbLRXA2.2, were identified based on transcriptome and quantitative real-time PCR analyses. The expression levels of these two LRX genes were significantly higher in the pollen grains and pollen tubes of the self-compatible cultivar 'Jinzhui' (harboring a spontaneous bud mutation) than in those of the self-incompatible cultivar 'Yali'. Both PbLRXA2.1 and PbLRXA2.2 stimulated pollen tube growth and attenuated the inhibitory effects of self S-RNase on pollen tube growth by stabilizing the actin cytoskeleton and enhancing cell wall integrity. These results indicate that abnormal expression of PbLRXA2.1 and PbLRXA2.2 is involved in the loss of self-incompatibility in 'Jinzhui'. The PbLRXA2.1 and PbLRXA2.2 promoters were directly bound by the ABRE-binding factor PbABF.D.2. Knockdown of PbABF.D.2 decreased PbLRXA2.1 and PbLRXA2.2 expression and inhibited pollen tube growth. Notably, the expression of PbLRXA2.1, PbLRXA2.2, and PbABF.D.2 was repressed by self S-RNase, suggesting that self S-RNase can arrest pollen tube growth by restricting the PbABF.D.2-PbLRXA2.1/PbLRXA2.2 signal cascade. These results provide novel insight into pollen tube growth arrest by self S-RNase.

Multi-omics analyses reveal MdMYB10 hypermethylation being responsible for a bud sport of apple fruit color.

Apple bud sports offer a rich resource for clonal selection of numerous elite cultivars. The accumulation of somatic mutations as plants develop may potentially impact the emergence of bud sports. Previous studies focused on somatic mutation in the essential genes associated with bud sports. However, the rate and function of genome-wide somatic mutations that accumulate when a bud sport arises remain unclear. In this study, we identified a branch from a 10-year-old tree of the apple cultivar 'Oregon Spur II' as a bud sport. The mutant branch showed reduced red coloration on fruit skin. Using this plant material, we assembled a high-quality haplotype reference genome consisting of 649.61 Mb sequences with a contig N50 value of 2.04 Mb. We then estimated the somatic mutation rate of the apple tree to be 4.56 × 10 -8 per base per year, and further identified 253 somatic single-nucleotide polymorphisms (SNPs), including five non-synonymous SNPs, between the original type and mutant samples. Transcriptome analyses showed that 69 differentially expressed genes between the original type and mutant fruit skin were highly correlated with anthocyanin content. DNA methylation in the promoter of five anthocyanin-associated genes was increased in the mutant compared with the original type as determined using DNA methylation profiling. Among the genetic and epigenetic factors that directly and indirectly influence anthocyanin content in the mutant apple fruit skin, the hypermethylated promoter of MdMYB10 is important. This study indicated that numerous somatic mutations accumulated at the emergence of a bud sport from a genome-wide perspective, some of which contribute to the low coloration of the bud sport.

The effect of different carbon sources on biofouling in membrane fouling simulators: microbial community and implications.

Biofouling is a problem affecting the operation of nanofiltration systems due to the complexity of the carbon matrix affecting bacteria and biofilm growth. This study used membrane fouling simulators to investigate the effects of five different carbon sources on the biofouling of nanofiltration membranes. For all the carbon sources analyzed, the increase in pressure drop was most accelerated for acetate. The use of acetate as the single carbon source produced less adenosine triphosphate but more extracellular polymers than glucose. The microbial community was analyzed using 16 s rRNA. The use of more than a single carbon source produced an increase in bacteria diversity even at similar concentrations. The relative abundance of proteobacteria was the highest at the phylum level (95%) when a single carbon source was added. Additionally, it was found that the use of different carbon sources produced a shift in the microbial community, affecting the biofouling and pressure drop on membranes.

Transposon insertions regulate genome-wide allele-specific expression and underpin flower colour variations in apple (Malus spp.).

Allele-specific expression (ASE) can lead to phenotypic diversity and evolution. However, the mechanisms regulating ASE are not well understood, particularly in woody perennial plants. In this study, we investigated ASE genes in the apple cultivar 'Royal Gala' (RG). A high quality chromosome-level genome was assembled using a homozygous tetra-haploid RG plant, derived from anther cultures. Using RNA-sequencing (RNA-seq) data from RG flower and fruit tissues, we identified 2091 ASE genes. Compared with the haploid genome of 'Golden Delicious' (GD), a parent of RG, we distinguished the genomic sequences between the two alleles of 817 ASE genes, and further identified allele-specific presence of a transposable element (TE) in the upstream region of 354 ASE genes. These included MYB110a that encodes a transcription factor regulating anthocyanin biosynthesis. Interestingly, another ASE gene, MYB10 also showed an allele-specific TE insertion and was identified using genome data of other apple cultivars. The presence of the TE insertion in both MYB genes was positively associated with ASE and anthocyanin accumulation in apple petals through analysis of 231 apple accessions, and thus underpins apple flower colour evolution. Our study demonstrated the importance of TEs in regulating ASE on a genome-wide scale and presents a novel method for rapid identification of ASE genes and their regulatory elements in plants.

microRNA172 targets APETALA2 to regulate flavonoid biosynthesis in apple (Malus domestica).

MicroRNA172 (miR172) plays a role in regulating a diverse range of plant developmental processes, including flowering, fruit development and nodulation. However, its role in regulating flavonoid biosynthesis is unclear. In this study, we show that transgenic apple plants over-expressing miR172 show a reduction in red coloration and anthocyanin accumulation in various tissue types. This reduction was consistent with decreased expression of APETALA2 homolog MdAP2_1a (a miR172 target gene), MdMYB10, and targets of MdMYB10, as demonstrated by both RNA-seq and qRT-PCR analyses. The positive role of MdAP2_1a in regulating anthocyanin biosynthesis was supported by the enhanced petal anthocyanin accumulation in transgenic tobacco plants overexpressing MdAP2_1a, and by the reduction in anthocyanin accumulation in apple and cherry fruits transfected with an MdAP2_1a virus-induced-gene-silencing construct. We demonstrated that MdAP2_1a could bind directly to the promoter and protein sequences of MdMYB10 in yeast and tobacco, and enhance MdMYB10 promotor activity. In Arabidopsis, over-expression of miR172 reduced flavonoid (including anthocyanins and flavonols) concentration and RNA transcript abundance of flavonoid genes in plantlets cultured on medium containing 7% sucrose. The anthocyanin content and RNA abundance of anthocyanin genes could be partially restored by using a synonymous mutant of MdAP2_1a, which had lost the miR172 target sequences at mRNA level, but not restored by using a WT MdAP2_1a. These results indicate that miR172 inhibits flavonoid biosynthesis through suppressing the expression of an AP2 transcription factor that positively regulates MdMYB10.

Significant improvement of apple (Malus domestica Borkh.) transgenic plant production by pre-transformation with a Baby boom transcription factor.

BABY BOOM (BBM) is a member of the APETALA2/ETHYLENE RESPONSE FACTOR (AP2/ERF) family and its expression has been shown to improve herbaceous plant transformation and regeneration. However, this improvement has not been shown clearly for tree species. This study demonstrated that the efficiency of transgenic apple (Malus domestica Borkh.) plant production was dramatically increased by ectopic expression of the MdBBM1 gene. "Royal Gala" apple plants were first transformed with a CaMV35S-MdBBM1 construct (MBM) under kanamycin selection. These MBM transgenic plants exhibited enhanced shoot regeneration from leaf explants on tissue culture media, with most plants displaying a close-to-normal phenotype compared with CaMV35S-GUS transgenic plants when grown under greenhouse conditions, the exception being that some plants had slightly curly leaves. Thin leaf sections revealed the MBM plants produced more cells than the GUS plants, indicating that ectopic-expression of MdBBM1 enhanced cell division. Transcriptome analysis showed that mRNA levels for cell division activators and repressors linked to hormone (auxin, cytokinin and brassinosteroid) signalling pathways were enhanced and reduced, respectively, in the MBM plants compared with the GUS plants. Plants of eight independent MBM lines were compared with the GUS plants by re-transforming them with an herbicide-resistant gene construct. The number of transgenic plants produced per 100 leaf explants was 0-3% for the GUS plants, 3-8% for five MBM lines, and 20-30% for three MBM lines. Our results provided a solution for overcoming the barriers to transgenic plant production in apple, and possibly in other trees.

The MADS-box gene FveSEP3 plays essential roles in flower organogenesis and fruit development in woodland strawberry.

Flower and fruit development are two key steps for plant reproduction. The ABCE model for flower development has been well established in model plant species; however, the functions of ABCE genes in fruit crops are less understood. In this work, we identified an EMS mutant named R27 in woodland strawberry (Fragaria vesca), showing the conversion of petals, stamens, and carpels to sepaloid organs in a semidominant inheritance fashion. Mapping by sequencing revealed that the class E gene homolog FveSEP3 (FvH4_4g23530) possessed the causative mutation in R27 due to a G to E amino acid change in the conserved MADS domain. Additional fvesep3CR mutants generated by CRISPR/Cas9 displayed similar phenotypes to fvesep3-R27. Overexpressing wild-type or mutated FveSEP3 in Arabidopsis suggested that the mutation in R27 might cause a dominant-negative effect. Further analyses indicated that FveSEP3 physically interacted with each of the ABCE proteins in strawberry. Moreover, both R27 and fvesep3CR mutants exhibited parthenocarpic fruit growth and delayed fruit ripening. Transcriptome analysis revealed that both common and specific differentially expressed genes were identified in young fruit at 6-7 days post anthesis (DPA) of fvesep3 and pollinated wild type when compared to unpollinated wild type, especially those in the auxin pathway, a key hormone regulating fruit set in strawberry. Together, we provided compelling evidence that FveSEP3 plays predominant E functions compared to other E gene homologs in flower development and that FveSEP3 represses fruit growth in the absence of pollination and promotes fruit ripening in strawberry.

A systems genetics approach reveals PbrNSC as a regulator of lignin and cellulose biosynthesis in stone cells of pear fruit.

BACKGROUND: Stone cells in fruits of pear (Pyrus pyrifolia) negatively influence fruit quality because their lignified cell walls impart a coarse and granular texture to the fruit flesh. RESULTS: We generate RNA-seq data from the developing fruits of 206 pear cultivars with a wide range of stone cell contents and use a systems genetics approach to integrate co-expression networks and expression quantitative trait loci (eQTLs) to characterize the regulatory mechanisms controlling lignocellulose formation in the stone cells of pear fruits. Our data with a total of 35,897 expressed genes and 974,404 SNPs support the identification of seven stone cell formation modules and the detection of 139,515 eQTLs for 3229 genes in these modules. Focusing on regulatory factors and using a co-expression network comprising 39 structural genes, we identify PbrNSC as a candidate regulator of stone cell formation. We then verify the function of PbrNSC in regulating lignocellulose formation using both pear fruit and Arabidopsis plants and further show that PbrNSC can transcriptionally activate multiple target genes involved in secondary cell wall formation. CONCLUSIONS: This study generates a large resource for studying stone cell formation and provides insights into gene regulatory networks controlling the formation of stone cell and lignocellulose.

PpIAA1 and PpERF4 form a positive feedback loop to regulate peach fruit ripening by integrating auxin and ethylene signals.

The signaling pathways of both auxin and ethylene regulate peach fruit ripening via the Aux/IAA and ERF transcription factors, respectively. However, the molecular mechanisms that coordinate both auxin and ethylene signals during peach fruit ripening remain unclear. In this study, we show that PpIAA1 and PpERF4 act as key players in a positive feedback loop, and promote peach fruit ripening by directly binding to and enhancing the activity of target gene promoters. PpIAA1 increased the expression of the ethylene biosynthesis gene PpACS1. Furthermore, PpERF4 enhanced the transcription of PpACO1 and PpIAA1 genes by binding to their promoters. Additionally, PpIAA1 and PpERF4 bound to each other to form a complex, which then enhanced the transcription of abscisic acid biosynthesis genes (PpNCED2 and PpNCED3) and the fruit softening gene (PpPG1) to levels higher than those achieved by each transcription factor individually. Moreover, overexpression of PpIAA1 in tomato accelerated fruit ripening and shortened the fruit shelf-life by increasing the production of ethylene and the expression levels of ripening regulator genes. Collectively, these results advance our understanding of the molecular mechanisms underlying peach fruit ripening and softening via auxin and ethylene signaling pathways.

The NAM/ATAF1/2/CUC2 transcription factor PpNAC.A59 enhances PpERF.A16 expression to promote ethylene biosynthesis during peach fruit ripening.

Peach is a typical climacteric fruit that releases ethylene during fruit ripening. Several studies have been conducted on the transcriptional regulation of ethylene biosynthesis in peach fruit. Herein, an ethylene response factor, PpERF.A16, which was induced by exogenous ethylene, could enhance ethylene biosynthesis by directly inducing the expression of 1-aminocyclopropane-1-carboxylic acid synthase (PpACS1) and 1-aminocyclopropane-1-carboxylic acid oxidase (PpACO1) genes. Moreover, the NAM/ATAF1/2/CUC2 (NAC) transcription factor (TF) PpNAC.A59 was coexpressed with PpERF.A16 in all tested peach cultivars. Interestingly, PpNAC.A59 can directly interact with the promoter of PpERF.A16 to induce its expression but not enhance LUC activity driven by any promoter of PpACS1 or PpACO1. Thus, PpNAC.A59 can indirectly mediate ethylene biosynthesis via the NAC-ERF signaling cascade to induce the expression of both PpACS1 and PpACO1. These results enrich the genetic network of fruit ripening in peach and provide new insight into the ripening mechanism of other perennial fruits.

Primary Bitter Taste of Citrus is Linked to a Functional Allele of the 1,2-Rhamnosyltransferase Gene Originating from Citrus grandis.

1,2-Rhamnosyltransferase (1,2RhaT) catalyzes the final step of production of flavanone neohesperidoside (FNH) that is responsible for the primary bitter taste of citrus fruits. In this study, species-specific flavonoid profiles were determined in 87 Citrus accessions by identifying eight main flavanone glycosides (FGs). Accumulation of FNHs was completely correlated to the presence of the 1,2RhaT gene in 87 citrus accessions analyzed using a novel 1,2RhaT-specific DNA marker. Pummelo (Citrus grandis) was identified as the genetic origin for a function allele of 1,2RhaT that underpinned FNH-bitterness in modern citrus cultivars. In addition, genes encoding six MYB and five bHLH transcription factors were shown to coexpress with 1,2RhaT and other flavonoid pathway genes related to FNH accumulation, indicating that these transcription factors may affect the fruit taste of citrus. This study provides a better understanding of bitterness formation in Citrus varieties and a genetic marker for the early selection of nonbitterness lines in citrus breeding programs.

Two loss-of-function alleles of the glutathione S-transferase (GST) gene cause anthocyanin deficiency in flower and fruit skin of peach (Prunus persica).

Flower and fruit colors are important agronomic traits. To date, there is no forward genetic evidence that the glutathione S-transferase (GST) gene is responsible for the white flower color in peach (Prunus persica). In this study, genetic analysis indicated that the white-flower trait is monogenetic, is recessive to the non-white allele, and shows pleiotropic effects with non-white-flowered types. The genetic locus underpinning this trait was mapped onto chromosome 3 between 0.421951 and 3.227115 Mb by using bulked segregant analysis in conjunction with whole-genome sequencing, and was further mapped between 0 and 1.178149 Mb by using the backcross 1 (BC1 ) population. Finally, the locus was fine-mapped within 535.974- and 552.027-kb intervals by using 151 F2 individuals and 75 individuals from a BC1 self-pollinated (BC1 S1 ) population, respectively. Pp3G013600, encoding a GST that is known to transport anthocyanin, was identified within the mapping interval. The analysis of genome sequence data showed Pp3G013600 in white flowers has a 2-bp insertion or a 5-bp deletion in the third exon. These variants likely render the GST non-functional because of early stop codons that reduce the protein length from 215 amino acids to 167 and 175 amino acids, respectively. Genetic markers based on these variants validated a complete correlation between the GST loss-of-function alleles and white flower in 128 peach accessions. This correlation was further confirmed by silencing of Pp3G013600 using virus-induced gene silencing technology, which reduced anthocyanin accumulation in peach fruit. The new knowledge from this study is useful for designing peach breeding programs to generate cultivars with white flower and fruit skin.

Unraveling a genetic roadmap for improved taste in the domesticated apple.

Although taste is an important aspect of fruit quality, an understanding of its genetic control remains elusive in apple and other fruit crops. In this study, we conducted genomic sequence analysis of 497 Malus accessions and revealed erosion of genetic diversity caused by apple breeding and possible independent domestication events of dessert and cider apples. Signatures of selection for fruit acidity and size, but not for fruit sugar content, were detected during the processes of both domestication and improvement. Furthermore, we found that single mutations in major genes affecting fruit taste, including Ma1, MdTDT, and MdSOT2, dramatically decrease malate, citrate, and sorbitol accumulation, respectively, and correspond to important domestication events. Interestingly, Ma1 was identified to have pleiotropic effects on both organic acid content and sugar:acid ratio, suggesting that it plays a vital role in determining fruit taste. Fruit taste is unlikely to have been negatively affected by linkage drag associated with selection for larger fruit that resulted from the pyramiding of multiple genes with minor effects on fruit size. Collectively, our study provides new insights into the genetic basis of fruit quality and its evolutionary roadmap during apple domestication, pinpointing several candidate genes for genetic manipulation of fruit taste in apple.

Diverse Functions of IAA-Leucine Resistant PpILR1 Provide a Genic Basis for Auxin-Ethylene Crosstalk During Peach Fruit Ripening.

Auxin and ethylene play critical roles in the ripening of peach (Prunus persica) fruit; however, the interaction between these two phytohormones is complex and not fully understood. Here, we isolated a peach ILR gene, PpILR1, which encodes an indole-3-acetic acid (IAA)-amino hydrolase. Functional analyses revealed that PpILR1 acts as a transcriptional activator of 1-amino cyclopropane-1-carboxylic acid synthase (PpACS1), and hydrolyzes auxin substrates to release free auxin. When Cys137 was changed to Ser137, PpILR1 failed to show hydrolase activity but continued to function as a transcriptional activator of PpACS1 in tobacco and peach transient expression assays. Furthermore, transgenic tomato plants overexpressing PpILR1 exhibited ethylene- and strigolactone-related phenotypes, including premature pedicel abscission, leaf and petiole epinasty, and advanced fruit ripening, which are consistent with increased expression of genes involved in ethylene biosynthesis and fruit ripening, as well as suppression of branching and growth of internodes (related to strigolactone biosynthesis). Collectively, these results provide novel insights into the role of IAA-amino acid hydrolases in plants, and position the PpILR1 protein at the junction of auxin and ethylene pathways during peach fruit ripening. These results could have substantial implications on peach fruit cultivation and storage in the future.