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Small secreted peptides (SSPs) play important roles in regulating plants' growth and development in response to external stimulus, but the genes and functions of SSPs in many species are still unknown. Therefore, it is particularly significant to characterize and annotate SSP genes in plant genomes. As a widely used stock of pears, Pyrus betulifolia has strong resistance to biotic and abiotic stresses. In this study, we analyzed the SSPs genes in the genome of P. betulifolia according to their characteristics and homology. A total of 1195 SSP genes were identified, and most of them are signaling molecules. Among these, we identified a new SSP, subtilase peptide 3 (SUBPEP3), which derived from the PA region of preSUBPEP3, increasing the expression level under salt stress. Both adding synthetic peptide SUBPEP3 to the culture medium of pears and the overexpression of SUBPEP3 in tobacco can improve the salt tolerance of plants. In summary, we annotated the SSP genes in the P. betulifolia genome and identified a small secreted peptide SUBPEP3 that regulates the salt tolerance of P. betulifolia, which provides an important theoretical basis for further revealing the function of SSPs. [ABSTRACT FROM AUTHOR]

SMXL genes constitute a conserved gene family that is ubiquitous in angiosperms and involved in regulating various plant processes, including branching, leaf elongation, and anthocyanin biosynthesis, but little is known about their molecular functions in pear branching. Here, we performed genome-wide identification and investigation of the SMXL genes in 16 angiosperms and analyzed their phylogenetics, structural features, conserved motifs, and expression patterns. In total, 121 SMXLs genes were identified and were classified into four groups. The number of non-redundant SMXL genes in each species varied from 3 (Amborella trichopoda Baill.) to 18 (Glycine max Merr.) and revealed clear gene expansion events over evolutionary history. All the SMXL genes showed conserved structures, containing no more than two introns. Three-dimensional protein structure prediction revealed distinct structures between but similar structures within groups. A quantitative real-time PCR analysis revealed different expressions of 10 SMXL genes from pear branching induced by fruit-thinning treatment. Overall, our study provides a comprehensive investigation of SMXL genes in the Rosaceae family, especially pear. The results offer a reference for understanding the evolutionary history of SMXL genes and provide excellent candidates for studying fruit tree branching regulation, and in facilitating pear pruning and planting strategies. [ABSTRACT FROM AUTHOR]

Pear black spot disease, caused by Alternaria alternata, is a devastating disease in pears and leads to enormous economic losses worldwide. In this investigation, we isolated a Streptomyces odonnellii SZF-179 from the rhizosphere soil of pear plants in China. Indoor confrontation experiments results showed that both SZF-179 and its aseptic filtrate had excellent inhibitory effects against A. alternata. Afterwards, the main antifungal compound of SZF-179 was identified as polyene, with thermal and pH stability in the environment. A microscopic examination of A. alternata mycelium showed severe morphological abnormalities caused by SZF-179. Protective studies showed that SZF-179 fermentation broth could significantly reduce the diameter of the necrotic lesions on pear leaves by 42.25%. Furthermore, the potential of fermentation broth as a foliar treatment to control black leaf spot was also evaluated. Disease indexes of 'Hosui' and 'Wonwhang' pear plants treated with SZF-179 fermentation broth were lower than that of control plants. Overall, SZF-179 is expected to be developed into a safe and broad-spectrum biocontrol agent. No studies to date have evaluated the utility of S. odonnellii for the control of pear black spot disease; our study fills this research gap. Collectively, our findings provide new insights that will aid the control of pear black spot disease, as well as future studies of S. odonnellii strains. [ABSTRACT FROM AUTHOR]

Valsa pyri-induced pear Valsa canker is among the most prevalent diseases to impact pear quality and yields. Biocontrol strategies to control plant disease represent an attractive alternative to the application of fungicides. In this study, the potential utility of Bacillus atrophaeus strain HF1 was assessed as a biocontrol agent against pear Valsa canker. Strain HF1 suppressed V. pyri mycelium growth by 61.20% and induced the development of malformed hyphae. Both culture filtrate and volatile organic compounds (VOCs) derived from strain HF1 were able to antagonize V. pyri growth. Treatment with strain HF1-derived culture filtrate or VOCs also induced the destruction of hyphal cell membranes. Headspace mixtures prepared from strain HF1 were analyzed, leading to the identification of 27 potential VOCs. Of the thirteen pure chemicals tested, iberverin, hexanoic acid, and 2-methylvaleraldehyde exhibited the strongest antifungal effects on V. pyri, with respective EC50 values of 0.30, 6.65, and 74.07 μL L−1. Fumigation treatment of pear twigs with each of these three compounds was also sufficient to prevent the development of pear Valsa canker. As such, these results demonstrate that B. atrophaeus strain HF1 and the volatile compounds iberverin, hexanoic acid, and 2-methylvaleraldehyde exhibit promise as novel candidate biocontrol agents against pear Valsa canker. [ABSTRACT FROM AUTHOR]

Proteins encoded by the G-box regulating factor (GRF, also called 14-3-3) gene family are involved in protein–protein interactions and mediate signaling transduction, which play important roles in plant growth, development, and stress responses. However, there were no detailed investigations of the GRF gene family in pear at present. In this study, we identified 25 GRF family members in the pear genome. Based on a phylogenetic analysis, the 25 GRF genes were clustered into two groups; the ε group and the non-ε group. Analyses of the exon–intron structures and motifs showed that the gene structures were conserved within each of the ε and non-ε groups. Gene duplication analysis indicated that most of the PbGRF gene expansion that occurred in both groups was due to WGD/segmental duplication. Phosphorylation sites analysis showed that the main phosphorylation sites of PbGRF proteins were serine residues. For gene expression, five PbGRF genes (PbGRF7, PbGRF11, PbGRF16, PbGRF21, and PbGRF23) were highly expressed in fruits, and PbGRF18 was highly expressed in all tissues. Further analysis revealed that eight PbGRF genes were significantly differentially expressed after treatment with different sugars; the expression of PbGRF7, PbGRF8, and PbGRF11 significantly increased, implying the involvement of these genes in sugar signaling. In addition, subcellular localization studies showed that the tested GRF proteins localize to the plasma membrane, and transgenic analysis showed that PbGRF18 can increase the sugar content in tomato leaves and fruit. The results of our research establish a foundation for functional determination of PbGRF proteins, and will help to promote a further understanding of the regulatory network in pear fruit development. [ABSTRACT FROM AUTHOR]

PEAR proteins are a type of plant-specific DNA binding with one finger (Dof) transcription factors that play a key role in the regulation of plant growth, especially during phloem cell growth and seed germination in Arabidopsis. However, the identification, characteristics and function of PEAR proteins, particularly in woody plants, need to be further studied. In the present study, 43 candidate PEAR proteins harboring the conserved Zf-Dof domain were obtained in Populus yunnanensis. Based on phylogenetic and structural analysis, 10 representative PEAR candidates were selected, belonging to different phylogenetic groups. The functions of PEAR proteins in the stress response, signal transduction, and growth regulation of stem cambium and roots undergoing vigorous cell division in Arabidopsis were revealed based on their expression patterns as characterized by qRT-PCR analysis, in accordance with the results of cis-element analysis. In vitro experiments showed that the interaction of transcription factor (E2F) and cyclin indirectly reflects the growth regulation function of PEAR through light signaling and cell-cycle regulation. Therefore, our results provide new insight into the identity of PEAR proteins and their function in stress resistance and vigorous cell division regulation of tissues in P. yunnanensis, which may serve as a basis for further investigation of the functions and characteristics of PEAR proteins in other plants. [ABSTRACT FROM AUTHOR]

Stable and high expression of introduced genes is a prerequisite for using transgenic trees. Transgene stacking enables combining several valuable traits, but repeated transformation increases the risk of unintended effects. This work studied the stability and intron-mediated enhancement of uidA gene expression in leaves and different anatomical parts of pear fruits during field trials over 14 years. The stability of reporter and herbicide resistance transgenes in retransformed pear plants, as well as possible unintended effects using high-throughput phenotyping tools, were also investigated. The activity of β-glucuronidase (GUS) varied depending on the year, but silencing did not occur. The uidA gene was expressed to a maximum in seeds, slightly less in the peel and peduncles, and much less in the pulp of pear fruits. The intron in the uidA gene stably increased expression in leaves and fruits by approximately twofold. Retransformants with the bar gene showed long-term herbicide resistance and exhibited no consistent changes in leaf size and shape. The transgenic pear was used as rootstock and scion, but grafted plants showed no transport of the GUS protein through the graft in the greenhouse and field. This longest field trial of transgenic fruit trees demonstrates stable expression under varying environmental conditions, the expression-enhancing effect of intron and the absence of unintended effects in single- and double-transformed woody plants. [ABSTRACT FROM AUTHOR]

Grafting is widely used to improve the stress tolerance and the fruit yield of horticultural crops. Ribonucleoprotein complexes formed by mRNAs and proteins play critical roles in the communication between scions and stocks of grafted plants. In Pyrus betulaefolia, ankyrin was identified previously to promote the long-distance movement of the ribonucleoprotein complex(PbWoxT1-PbPTB3) by facilitating callose degradation at plasmodesmata. However, the mechanism of the ankyrin-mediated callose degradation remains elusive. In this study, we discovered a β-1,3-glucanase (EC 3.2.1.39, PbPDBG) using ankyrin as a bait from plasmodesmata by co-immunoprecipitation and mass spectrometry. Ankyrin was required for the plasmodesmata-localization of PbPDBG. The grafting and bombardment experiments indicated that overexpressing PbPDBG resulted in decreased callose content at plasmodesmata, and thereby promoting the long-distance transport of the ribonucleoprotein complex. Altogether, our findings revealed that PbPDBG was the key factor in ankyrin-mediated callose degradation at plasmodesmata. [ABSTRACT FROM AUTHOR]

The 9-cis-epoxycarotenoid dioxygenase (NCED) is a key enzyme for the process of ABA synthesis that plays key roles in a variety of biological processes. In the current investigation, genome-wide identification and comprehensive analysis of the NCED gene family in 'Kuerle Xiangli' (Pyrus sinkiangensis Yu) were conducted using the pear genomic sequence. In total, nineteen members of PbNCED genes were identified from the whole genome of pear, which are not evenly distributed over the scaffolds, and most of which were focussed in the chloroplasts. Sequence analysis of promoters showed many cis-regulatory elements, which presumably responded to phytohormones such as abscisic acid, auxin, etc. Synteny block indicated that the PbNCED genes have experienced strong purifying selection. Multiple sequence alignment demonstrated that these members are highly similar and conserved. In addition, we found that PbNCED genes were differentially expressed in various tissues, and three PbNCED genes (PbNCED1, PbNCED2, and PbNCED13) were differentially expressed in response to exogenous Gibberellin (GA3) and Paclobutrazol (PP333). PbNCED1 and PbNCED13 positively promote ABA synthesis in sepals after GA3 and PP333 treatment, whereas PbNCED2 positively regulated ABA synthesis in ovaries after GA3 treatment, and PbNCED13 positively regulated ABA synthesis in the ovaries after PP333 treatment. This study was the first genome-wide report of the pear NCED gene family, which could improve our understanding of pear NCED proteins and provide a solid foundation for future cloning and functional analyses of this gene family. Meanwhile, our results also give a better understanding of the important genes and regulation pathways related to calyx abscission in 'Kuerle Xiangli'. [ABSTRACT FROM AUTHOR]

The pear is an important fruit tree in temperate areas, but due to its sensitivity, fruit yield and quality are often affected by disease and pest attacks. Pear genotypes from a germplasm collection comprising 13 Pyrus species, 17 Romanian varieties, and 50 non-Romanian varieties from a worldwide assortment were investigated in this study. Throughout four years, response to attack of the principal pathogens and pests was investigated phenotypically under natural conditions of infection and infestation. SSR markers were used to analyze the genetic diversity of the genotypes. A standardized method for the evaluation of responses to biotic stressors was proposed, which highlighted significant differences between genotypes. The species and varieties with the lowest degrees of attack (DA%), calculated based on the frequency and intensity of attack, were identified for pear scab (Venturia pyrina), septoria (Septoria pyricola), fire blight (Erwinia amylovora), and psyllids (Psylla sp.). These accessions could provide valuable sources of genes of interest to develop resistant varieties in new pear breeding programs. By combining phenotypic and molecular analyses, significant information was obtained that can be exploited to generate high variability for selection through artificial hybridization by harnessing accessions with complementary molecular fingerprints and high genetic distances. [ABSTRACT FROM AUTHOR]

Calcium/calmodulin-dependent protein kinase (CaMK), a key downstream target protein in the Ca2+ signaling pathway of eukaryotes, plays an important regulatory role in the growth, development and pathogenicity of plant fungi. Three AaCaMKs (AaCaMK1, AaCaMK2 and AaCaMK3) with conserved PKC_like superfamily domains, ATP binding sites and ACT sites have been cloned from Alternaria alternata, However, their regulatory mechanism in A. alternata remains unclear. In this study, the function of the AaCaMKs in the development, infection structure differentiation and pathogenicity of A. alternata was elucidated through targeted gene disruption. The single disruption of AaCaMKs had no impact on the vegetative growth and spore morphology but significantly influenced hyphae growth, sporulation, biomass accumulation and melanin biosynthesis. Further expression analysis revealed that the AaCaMKs were up-regulated during the infection structure differentiation of A. alternata on hydrophobic and pear wax substrates. In vitro and in vivo analysis further revealed that the deletion of a single AaCaMKs gene significantly reduced the A. alternata conidial germination, appressorium formation and infection hyphae formation. In addition, pharmacological analysis confirmed that the CaMK specific inhibitor, KN93, inhibited conidial germination and appressorium formation in A. alternata. Meanwhile, the AaCaMKs genes deficiency significantly reduced the A. alternata pathogenicity. These results demonstrate that AaCaMKs regulate the development, infection structure differentiation and pathogenicity of A. alternata and provide potential targets for new effective fungicides. [ABSTRACT FROM AUTHOR]

Climate change is deeply impacting the food chain production, lowering quality and yield. In this context, the international scientific community has dedicated many efforts to enhancing resilience and sustainability in agriculture. Italy is among the main European producers of several fruit trees; therefore, national research centers and universities undertook several initiatives to maintain the specificity of the 'Made in Italy' label. Despite their importance, fruit crops are suffering from difficulties associated with the conventional breeding approaches, especially in terms of financial commitment, land resources availability, and long generation times. The 'new genomic techniques' (NGTs), renamed in Italy as 'technologies for assisted evolution' (TEAs), reduce the time required to obtain genetically improved cultivars while precisely targeting specific DNA sequences. This review aims to illustrate the role of the Italian scientific community in the use of NGTs, with a specific focus on Citrus, grapevine, apple, pear, chestnut, strawberry, peach, and kiwifruit. For each crop, the key genes and traits on which the scientific community is working, as well as the technological improvements and advancements on the regeneration of local varieties, are presented. Lastly, a focus is placed on the legal aspects in the European and in Italian contexts. [ABSTRACT FROM AUTHOR]

Superficial scald is a postharvest physiological disorder that occurs in pear during and after cold storage. In this study, the superficial scald index; α-farnesene and its oxidation products, conjugated trienols (CTols); phenolic content; and the expression of its related genes were investigated in two different pear cultivars, 'Wujiuxiang' (Pyrus communis L.) and 'Yali' (Pyrus bretschneideri R.), following 115 days of cold storage at 0 °C followed by 7 days of shelf life at 20 °C. The results indicated that the superficial scald occurred after 115 days of cold storage and became more severe during the shelf life of the 'Wujiuxiang' pear, whereas no scald was observed in 'Yali'. The α-farnesene levels increased rapidly at first and then decreased, while the CTols contents increased significantly in 'Wujiuxiang' as compared to 'Yali', and the expression levels of the genes involved in α-farnesene and CTols metabolism (HMGR1, HMGR2, GSTU7, GPX5, and GPX6), as well as the phenolic synthesis (PAL1, PAL2, C4H1, 4CL2, C3H, and ANR) of the peel, were significantly up-regulated at the onset of the superficial scald. In addition, the relative conductivity and contents of catechin and epicatechin were higher, and the expression level of the laccase gene (LAC7) significantly increased with the development of superficial scald, while lower contents of chlorogenic acid, arbutin, and isorhamnetin-3-3-glucoside, as well as the lower expression levels of a phenolic-synthesis-related gene (C4H3) and polyphenol oxidase genes (PPO1 and PPO5), were noticed in 'Wujiuxiang' as compared to 'Yali'. The results indicated that the onset and progression of superficial scald were associated with the accumulation of CTols, cell membrane breakdown, and higher catechin, epicatechin, and rutin contents, as well as the expression of associated genes of the peels of pear fruit. [ABSTRACT FROM AUTHOR]

Glycosylation is necessary for many processes of plant secondary metabolism. It can maintain plant homeostasis and is of great significance to normal plant growth and development. At present, the significance of glycosylation for lignin biosynthesis has been proven in some plants, but it has not yet been reported in pears. We used in situ hybridization, in vitro expression, substrate catalysis, transgenic Arabidopsisthaliana, and transient transformation of pear fruit in our investigation, which was predicated on the identification of a gene PbUGT72AJ2 that may be involved in lignin monolignol glycosylation according to our previous work. These results revealed that PbUGT72AJ2 transcripts were localized to some pulp cell walls, lignin deposition, and stone cell areas of pear fruit. The recombinant PbUGT72AJ2-pGEX4T-1 protein had activity against coniferyl alcohol and sinapyl alcohol, and its catalytic efficiency against coniferyl alcohol was higher than that against sinapyl alcohol. When PbUGT72AJ2 was transferred into Arabidopsisthaliana mutants, it was found that some characteristics of Arabidopsisthalianaugt72e3 mutants were restored. In Arabidopsisthaliana, overexpression of PbUGT72AJ2 enhanced the contents of coniferin and syringin, whereas lignification did not change significantly. Transient transformation of pear fruit showed that when PbUGT72AJ2 in pear fruit was silenced by RNA interference, the content of lignin and stone cells in pear fruit increased, whereas the gene PbUGT72AJ2 was overexpressed in pear fruit, and there was almost no change in the pear fruit compared with the control. Lignin deposition in pear fruit was closely related to stone cell development. In this study, we proved that PbUGT72AJ2 plays an important role in lignin deposition and stone cell development in pear fruit, which provides a molecular biological basis for improving pear fruit quality at the molecular level. [ABSTRACT FROM AUTHOR]

The SAUR (small auxin-up RNA) gene family is the biggest family of early auxin response genes in higher plants and has been associated with the control of a variety of biological processes. Although SAUR genes had been identified in several genomes, no systematic analysis of the SAUR gene family has been reported in Chinese white pear. In this study, comparative and systematic genomic analysis has been performed in the SAUR gene family and identified a total of 116 genes from the Chinese white pear. A phylogeny analysis revealed that the SAUR family could be classified into four groups. Further analysis of gene structure (introns/exons) and conserved motifs showed that they are diverse functions and SAUR-specific domains. The most frequent mechanisms are whole-genome duplication (WGD) and dispersed duplication (DSD), both of which may be important in the growth of the SAUR gene family in Chinese white pear. Moreover, cis-acting elements of the PbrSAUR genes were found in promoter regions associated with the auxin-responsive elements that existed in most of the upstream sequences. Remarkably, the qRT-PCR and transcriptomic data indicated that PbrSAUR13 and PbrSAUR52 were significantly expressed in fruit ripening. Subsequently, subcellular localization experiments revealed that PbrSAUR13 and PbrSAUR52 were localized in the nucleus. Moreover, PbrSAUR13 and PbrSAUR52 were screened for functional verification, and Dangshan pear and frandi strawberry were transiently transformed. Finally, the effects of these two genes on stone cells and lignin were analyzed by phloroglucinol staining, Fourier infrared spectroscopy, and qRT-PCR. It was found that PbrSAUR13 promoted the synthesis and accumulation of stone cells and lignin, PbrSAUR52 inhibited the synthesis and accumulation of stone cells and lignin. In conclusion, these results indicate that PbrSAUR13 and PbrSAUR52 are predominantly responsible for lignin inhibit synthesis, which provides a basic mechanism for further study of PbrSAUR gene functions. [ABSTRACT FROM AUTHOR]

Gibberellic acid (GA) is an important phytohormone that regulates every aspect of plant growth and development. While elements involved in GA signaling have been identified and, hence, their functions have been well studied in model plants, such as Arabidopsis and rice, very little is known in pear. We, therefore, analyzed the genes related to GA signaling from the recently sequenced genome of the wildtype 'duli' pear (Pyrus betulifolia Bunge), a widely used rootstock for grafting in pear cultivation in China due to its vigorous growth and resistance to abiotic and biotic stress. In total, 15 genes were identified, including five GA receptors PbGID1s (GA-INSENSTIVE DWARF 1), six GA negative regulators, PbDELLAs, and four GA positive regulators, PbSLYs. Exogenous application of GA could promote the expression of PbGID1s but inhibit that of PbDELLAs and PbSLYs in tissue culture 'duli' pear seedlings. The expression profiles of these genes in field-grown trees under normal growth conditions, as well as in tissue-cultured seedlings treated with auxin (IAA), GA, paclobutrazol (PAC), abscisic acid (ABA), and sodium chloride (NaCl), were also studied, providing further evidence of the involvement of these genes in GA signaling in 'duli' pear plants. The preliminary results obtained in this report lay a good foundation for future research into GA signaling pathways in pear. Importantly, the identification and preliminary functional verification of these genes could guide molecular breeding in order to obtain the highly desired dwarf pear rootstocks for high-density plantation to aid easy orchard management and high yielding of pear fruits. [ABSTRACT FROM AUTHOR]

Red-skinned pears are favored by people for their attractive appearance and abundance of anthocyanins. However, the molecular basis of anthocyanin biosynthesis in red pears remains elusive. Here, a comprehensive transcriptome analysis was conducted to explore the potential regulatory mechanism of anthocyanin biosynthesis in 'Red Zaosu' pear (Pyrus pyrifolia × Pyrus communis). Gene co-expression analysis and transcription factor mining identified 263 transcription factors, which accounted for 6.59% of the total number of transcription factors in the pear genome in two gene modules that are highly correlated with anthocyanin biosynthesis. Clustering, gene network modeling with STRING-DB, and local motif enrichment analysis (CentriMo) analysis suggested that PpPIF8 may play a role in anthocyanin biosynthesis. Furthermore, eight PIFs were identified in the pear genome, of which only PpPIF8 was rapidly induced by light. Functional studies showed that PpPIF8 localizes in the nucleus and is preferentially expressed in the tissue of higher levels of anthocyanin. The overexpression of PpPIF8 in pear peel and pear calli promotes anthocyanin biosynthesis and upregulates the expression of anthocyanin biosynthesis genes. Yeast-one hybrid and transgenic analyses indicated that PpPIF8 binds to the PpCHS promoter to induce PpCHS expression. The positive effect of PpPIF8 on anthocyanin biosynthesis is different from previously identified negative regulators of PyPIF5 and MdPIF7 in pear and apple. Taken together, our data not only provide a comprehensive view of transcription events during the coloration of pear peel, but also resolved the regulatory role of PpPIF8 in the anthocyanin biosynthesis pathway. [ABSTRACT FROM AUTHOR]

Dormancy is an adaptive mechanism that allows temperate deciduous plants to survive unfavorable winter conditions. In the present work, we investigated the possible function of abscisic acid (ABA) on the endodormancy process in pear. The ABA content increased during pear flower bud endodormancy establishment and decreased towards endodormancy release. In total, 39 putative genes related to ABA metabolism and signal transductions were identified from pear genome. During the para- to endodormancy transition, PpNCED-2 and PpNCED-3 had high expression levels, while PpCYP707As expression levels were low. However, during endodormancy, the expression of PpCYP707A-3 sharply increased with increasing cold accumulation. At the same time, the ABA content of pear buds declined, and the percentage of bud breaks rapidly increased. On the other hand, the expression levels of PpPYLs, PpPP2Cs, PpSnRK2s, and PpABI4/ABI5s were also changed during the pear flower bud dormancy cycle. Furthermore, exogenous ABA application to para-dormant buds significantly reduced the bud breaks and accelerated the transition to endodormancy. During the whole treatment time, the expression level of PpPP2C-12 decreased to a greater extent in ABA-treated buds than in control. However, the expression levels of PpSnRK2-1, PpSnRK2-4, and PpABI5-1 were higher in ABA-treated buds. Our results indicated that PpCYP707A-3 and PpNCEDs play pivotal roles on the regulation of endodormancy release, while ABA signal transduction pathway also appears to be involved in the process. The present work provided the basic information about the function of ABA-related genes during pear flower bud dormancy process. [ABSTRACT FROM AUTHOR]

Isorhamnetin glycosides are representative compounds of Opuntia ficus-indica that possess different biological activities. There is slight information about the changes in bioaccessibility induced by the glycosylation pattern of flavonoids, particularly for isorhamnetin. In this study, the bioaccessibility and permeability of isorhamnetin glycosides extracted from O. ficus-indica were contrasted with an isorhamnetin standard. Also, the plasma stability of these isorhamnetin glycosides after intravenous administration in rats was evaluated. Recoveries of isorhamnetin after oral and gastric digestion were lower than that observed for its glycosides. After intestinal digestion, isorhamnetin glycosides recoveries were reduced to less than 81.0%. The apparent permeability coefficient from apical (AP) to basolateral (BL) direction (Papp(AP-BL)) of isorhamnetin was 2.6 to 4.6-fold higher than those obtained for its glycosides. Isorhamnetin diglycosides showed higher Papp(AP-BL) values than triglycosides. Sugar substituents affected the Papp(AP-BL) of the triglycosides. Isorhamnetin glycosides were better retained in the circulatory system than the aglycone. After intravenous dose of the isorhamnetin standard, the elimination half-life was 0.64 h but increased to 1.08 h when the O. ficus-indica extract was administered. These results suggest that isorhamnetin glycosides naturally found in O. ficus-indica could be a controlled delivery system to maintain a constant plasmatic concentration of this important flavonoid to exert its biological effects in vivo. [ABSTRACT FROM AUTHOR]

Seed dormancy transition is a vital developmental process for seedling propagation and agricultural production. The process is precisely regulated by diverse endogenous genetic factors and environmental cues. Callery pear (Pyrus calleryana Decne) is an important rootstock species that requires cold stratification to break seed dormancy, but the mechanisms underlying pear seed dormancy release are not yet fully understood. Here, we analyzed the transcriptome profiles at three different stages of cold stratification in callery pear seeds using RNA sequencing combined with phytohormone and sugar content measurements. Significant alterations in hormone contents and carbohydrate metabolism were observed and reflected the dormancy status of the seeds. The expressions of genes related to plant hormone metabolism and signaling transduction, including indole-3-acetic acid (IAA) biosynthesis (ASAs, TSA, NITs, YUC, and AAO) genes as well as several abscisic acid (ABA) and gibberellic acid (GA) catabolism and signaling transduction genes (CYP707As, GA2ox, and DELLAs), were consistent with endogenous hormone changes. We further found that several genes involved in cytokinin (CTK), ethylene (ETH), brassionolide (BR), and jasmonic acid (JA) metabolism and signaling transduction were differentially expressed and integrated in pear seed dormancy release. In accordance with changes in starch and soluble sugar contents, the genes associated with starch and sucrose metabolism were significantly up-regulated during seed dormancy release progression. Furthermore, the expression levels of genes involved in lipid metabolism pathways were also up-regulated. Finally, 447 transcription factor (TF) genes (including ERF, bHLH, bZIP, NAC, WRKY, and MYB genes) were observed to be differentially expressed during seed cold stratification and might relate to pear seed dormancy release. Our results suggest that the mechanism underlying pear seed dormancy release is a complex, transcriptionally regulated process involving hormones, sugars, lipids, and TFs. [ABSTRACT FROM AUTHOR]

Anthocyanin accumulation in vacuoles results in red coloration in pear peels. Glutathione S-transferase (GST) proteins have emerged as important regulators of anthocyanin accumulation. Here, a total of 57 PcGST genes were identified in the European pear 'Bartlett' (Pyrus communis) through comprehensive genomic analysis. Phylogenetic analysis showed that PcGST genes were divided into 10 subfamilies. The gene structure, chromosomal localization, collinearity relationship, cis-elements in the promoter region, and conserved motifs of PcGST genes were analyzed. Further research indicated that glutamic acid (Glu) can significantly improve anthocyanin accumulation in pear peels. RNA sequencing (RNA-seq) analysis showed that Glu induced the expression of most PcGST genes, among which PcGST57 was most significantly induced. Further phylogenetic analysis indicated that PcGST57 was closely related to GST genes identified in other species, which were involved in anthocyanin accumulation. Transcript analysis indicated that PcGST57 was expressed in various tissues, other than flesh, and associated with peel coloration at different developmental stages. Silencing of PcGST57 by virus-induced gene silencing (VIGS) inhibited the expression of PcGST57 and reduced the anthocyanin content in pear fruit. In contrast, overexpression of PcGST57 improved anthocyanin accumulation. Collectively, our results demonstrated that PcGST57 was involved in anthocyanin accumulation in pear and provided candidate genes for red pear breeding. [ABSTRACT FROM AUTHOR]

The GATA gene family is one of the most important transcription factors (TFs). It extensively exists in plants, contributes to diverse biological processes such as the development process, and responds to environmental stress. Although the GATA gene family has been comprehensively and systematically studied in many species, less is known about GATA genes in Chinese pears (Pyrus bretschneideri). In the current study, the GATA gene family in the four Rosaceae genomes was identified, its structural characteristics identified, and a comparative analysis of its properties was carried out. Ninety-two encoded GATA proteins were authenticated in the four Rosaceae genomes (Pyrus bretschneideri, Prunus avium, Prunus mume, and Prunus persica) and categorized into four subfamilies (Ⅰ–Ⅳ) according to phylogeny. The majority of GATA genes contained one to two introns and conserved motif composition analysis revealed their functional divergence. Whole-genome duplications (WGDs) and dispersed duplication (DSD) played a key role in the expansion of the GATA gene family. The microarray indicated that, among P. bretschneideri, P. avium, P. mume and P. persica, GATA duplicated regions were more conserved between Pyrus bretschneideri and Prunus persica with 32 orthologous genes pairs. The physicochemical parameters, duplication patterns, non-synonymous (ka), and synonymous mutation rate (ks) and GO annotation ontology were performed using different bioinformatics tools. cis-elements respond to various phytohormones, abiotic/biotic stress, and light-responsive were found in the promoter regions of GATA genes which were induced via stimuli. Furthermore, subcellular localization of the PbGATA22 gene product was investigated, showing that it was present in the nucleus of tobacco (Nicotiana tabacum) epidermal cells. Finally, in silico analysis was performed on various organs (bud, leaf, stem, ovary, petal, and sepal) and different developmental stages of fruit. Subsequently, the expression profiles of PbGATA genes were extensively expressed under exogenous hormonal treatments of SA (salicylic acid), MeJA (methyl jasmonate), and ABA (abscisic acid) indicating that play important role in hormone signaling pathways. A comprehensive analysis of GATA transcription factors was performed through systematic biological approaches and comparative genomics to establish a theoretical base for further structural and functional investigations in Rosaceae species. [ABSTRACT FROM AUTHOR]

Bud endodormancy is an important, complex process subject to both genetic and epigenetic control, the mechanism of which is still unclear. The endogenous hormone abscisic acid (ABA) and its signaling pathway play important roles in the endodormancy process, in which the type 2C protein phosphatases (PP2Cs) is key to the ABA signal pathway. Due to its excellent effect on endodormancy release, hydrogen cyanamide (HC) treatment is considered an effective measure to study the mechanism of endodormancy release. In this study, RNA-Seq analysis was conducted on endodormant floral buds of pear (Pyrus pyrifolia) with HC treatment, and the HC-induced PP2C gene PpPP2C1 was identified. Next, software prediction, expression tests and transient assays revealed that lncRNA PpL-T31511-derived Pp-miRn182 targets PpPP2C1. The expression analysis showed that HC treatment upregulated the expression of PpPP2C1 and downregulated the expression of PpL-T31511 and Pp-miRn182. Moreover, HC treatment inhibited the accumulation of ABA signaling pathway-related genes and hydrogen peroxide (H2O2). Furthermore, overexpression of Pp-miRn182 reduced the inhibitory effect of PpPP2C1 on the H2O2 content. In summary, our study suggests that downregulation of PpL-T31511-derived Pp-miRn182 promotes HC-induced endodormancy release in pear plants through the PP2C-H2O2 pathway. [ABSTRACT FROM AUTHOR]

Commercialization of cactus pears based on their antioxidant properties can generate competitive advantages, and these can turn into business opportunities and the development of new products and a high-value ingredient for the food industry. This work evaluated the antioxidant activities (1,1-diphenyl-2-picrylhydrazyl radical-scavenging, protection against oxidation of a &bgr;-carotene-linoleic acid emulsion, and iron (II) chelation), the content of total phenolic compounds, ascorbic acid, betacyanin, betaxanthin and the stability of betacyanin pigments in presence of Cu (II)-dependent hydroxyl radicals (OH·), in 18 cultivars of purple, red, yellow and white cactus pear from six Mexican states. Our results indicated that the antiradical activities from yellow and white cactus pear cultivars were not significantly different (p < 0.05) and were lower than the average antiradical activities in red and purple cultivars. The red cactus pear from the state of Zacatecas showed the highest antioxidant activity. The free radical scavenging activity for red cactus pears was significantly correlated (p < 0.05) to the concentration of total phenolic compounds (R2 = 0.90) and ascorbic acid (R2 = 0.86). All 18 cultivars of cactus pears studied showed significant chelating activity of ferrous ions. The red and purple cactus pears showed a great stability when exposed to OH·. [ABSTRACT FROM AUTHOR]

Dye-sensitized solar cells (DSSCs) were assembled by using the bougainvillea flowers, red turnip and the purple wild Sicilian prickly pear fruit juice extracts as natural sensitizers of TiO2 films. The yellow orange indicaxanthin and the red purple betacyanins are the main components in the cocktail of natural dyes obtained from these natural products. The best overall solar energy conversion efficiency of 1.7% was obtained, under AM 1.5 irradiation, with the red turnip extract, that showed a remarkable current density (Jsc = 9.5 mA/cm2) and a high IPCE value (65% at λ = 470 nm). Also the purple extract of the wild Sicilian prickly pear fruit showed interesting performances, with a Jsc of 9.4 mA/cm2, corresponding to a solar to electrical power conversion of 1.26%. [ABSTRACT FROM AUTHOR]

Seedless fruit is a feature appreciated by consumers. The ovule abortion process is highly orchestrated and controlled by numerous environmental and endogenous signals. However, the mechanisms underlying ovule abortion in pear remain obscure. Here, we found that gibberellins (GAs) have diverse functions during ovules development between seedless pear '1913' and seeded pear, and that GA4+7 activates a potential programmed cell death process in '1913' ovules. After hormone analyses, strong correlations were determined among jasmonic acid (JA), ethylene and salicylic acid (SA) in seedless and seeded cultivars, and GA4+7 treatments altered the hormone accumulation levels in ovules, resulting in significant correlations between GA and both JA and ethylene. Additionally, SA contributed to ovule abortion in '1913'. Exogenously supplying JA, SA or the ethylene precursor 1-aminocyclopropane-1-carboxylic acid promoted 'Bartlett' seed death. The regulatory mechanism in which ethylene controls ovule death has been demonstrated; therefore, JA's role in regulating '1913' ovule abortion was investigated. A further study identified that the JA signaling receptor MYC2 bound the SENESCENCE-ASSOCIATED 39 promoter and triggered its expression to regulate ovule abortion. Thus, we established ovule abortion-related relationships between GA and the hormones JA, ethylene and SA, and we determined their synergistic functions in regulating ovule death. [ABSTRACT FROM AUTHOR]

Fruit acidity is one of the main determinants of fruit flavor and a target trait in fruit breeding. However, the genomic mechanisms governing acidity variation among different pear varieties remain poorly understood. In this study, two pear varieties with contrasting organic acid levels, 'Dangshansuli' (low-acidity) and 'Amute' (high-acidity), were selected, and a combination of transcriptome and population genomics analyses were applied to characterize their patterns of gene expression and genetic variation. Based on RNA-seq data analysis, differentially expressed genes (DEGs) involved in organic acid metabolism and accumulation were identified. Weighted correlation network analysis (WGCNA) revealed that nine candidate TCA (tricarboxylic acid)-related DEGs and three acid transporter-related DEGs were located in three key modules. The regulatory networks of the above candidate genes were also predicted. By integrating pear resequencing data, two domestication-related genes were found to be upregulated in 'Amute', and this trend was further validated for other pear varieties with high levels of organic acid, suggesting distinct selective sweeps during pear dissemination and domestication. Collectively, this study provides insight into organic acid differences related to expression divergence and domestication in two pear varieties, pinpointing several candidate genes for the genetic manipulation of acidity in pears. [ABSTRACT FROM AUTHOR]

Related to ABSCISIC ACID INSENSITIVE3/VIVIPAROUS1 (ABI3/VP1, RAV), transcription factors (TFs) belonging to the APETALA2/ETHYLENE RESPONSIVE FACTOR (AP2/ERF) TF family play critical roles in plant growth, development, and responses to abiotic and biotic stress. In this study, 11 novel RAV TFs were identified in pear (Pyrus bretschneideri Rehd). A phylogenetic analysis revealed that the TFs clustered into three groups with 10 conserved motifs, some of which were group- or subgroup-specific, implying that they are important for the functions of the RAVs in these clades. RAVs in Pyrus and Malus were closely related, and the former showed a collinear relationship. Analysis of their expression patterns in different tissues and at various growth stages and their responses to abiotic and biotic stress suggested that PbRAV6 and PbRAV7 play important roles in drought stress and salt stress, respectively. We investigated the function of RAVs in pear peel coloration using two red pear varieties with different color patterns and applying data from transcriptome analyses. We found that PbRAV6 participates in the regulation of pericarp color. These findings provide insight into a new TF family in pear and a basis for further studies on the response to drought stress and fruit coloration in this commercially important crop. [ABSTRACT FROM AUTHOR]

Watercore is a physiological disorder that commonly occurs in sand pear cultivars. The typical symptom of watercore tissue is transparency, and it is often accompanied by browning, breakdown and a bitter taste during fruit ripening. To better understand the molecular mechanisms of watercore affecting fruit quality, this study performed transcriptome and metabolome analyses on watercore pulp from "Akibae" fruit 125 days after flowering. The present study found that the "Akibae" pear watercore pulp contained higher sorbitol and sucrose than healthy fruit. Moreover, the structure of the cell wall was destroyed, and the content of pectin, cellulose and hemicellulose was significantly decreased. In addition, the content of ethanol and acetaldehyde was significantly increased, and the content of polyphenol was significantly decreased. Watercore induced up-regulated expression levels of sorbitol synthesis-related (sorbitol-6-phosphate dehydrogenase, S6PDH) and sucrose synthesis-related genes (sucrose synthesis, SS), whereas it inhibited the expression of sorbitol decomposition-related genes (sorbitol dehydrogenase, SDH) and sorbitol transport genes (sorbitol transporter, SOT). Watercore also strongly induced increased expression levels of cell wall-degrading enzymes (polygalactosidase, PG; ellulase, CX; pectin methylesterase, PME), as well as ethanol synthesis-related (alcohol dehydrogenase, ADH), acetaldehyde synthesis-related (pyruvate decarboxylase, PDC) and polyphenol decomposition-related genes (polyphenol oxidase, PPO). Moreover, the genes that are involved in ethylene (1-aminocyclopropane- 1-carboxylate oxidase, ACO; 1-aminocyclopropane- 1-carboxylate synthase, ACS) and abscisic acid (short-chain alcohol dehydrogenase, SDR; aldehyde oxidase, AAO) synthesis were significantly up-regulated. In addition, the bitter tasting amino acids, alkaloids and polyphenols were significantly increased in watercore tissue. Above all, these findings suggested that the metabolic disorder of sorbitol and sucrose can lead to an increase in plant hormones (abscisic acid and ethylene) and anaerobic respiration, resulting in aggravated fruit rot and the formation of bitter substances. [ABSTRACT FROM AUTHOR]

Despite recent progress, the application of CRISPR/Cas9 in perennial plants still has many obstacles to overcome. Our previous results with CRISPR/Cas9 in apple and pear indicated the frequent production of phenotypic and genotypic chimeras, after editing of the phytoene desaturase (PDS) gene conferring albino phenotype. Therefore, our first objective was to determine if adding an adventitious regeneration step from leaves of the primary transgenic plants (T0) would allow a reduction in chimerism. Among hundreds of adventitious buds regenerated from a variegated T0 line, 89% were homogeneous albino. Furthermore, the analysis of the target zone sequences of twelve of these regenerated lines (RT0 for "regenerated T0" lines) indicated that 99% of the RT0 alleles were predicted to produce a truncated target protein and that 67% of RT0 plants had less heterogeneous editing profiles than the T0. Base editors are CRISPR/Cas9-derived new genome-editing tools that allow precise nucleotide substitutions without double-stranded breaks. Hence, our second goal was to demonstrate the feasibility of CRISPR/Cas9 base editing in apple and pear using two easily scorable genes: acetolactate synthase—ALS (conferring resistance to chlorsulfuron) and PDS. The two guide RNAs under MdU3 and MdU6 promoters were coupled into a cytidine base editor harboring a cytidine deaminase fused to a nickase Cas9. Using this vector; we induced C-to-T DNA substitutions in the target genes; leading to discrete variation in the amino-acid sequence and generating new alleles. By co-editing ALS and PDS genes; we successfully obtained chlorsulfuron resistant and albino lines in pear. Overall; our work indicates that a regeneration step can efficiently reduce the initial chimerism and could be coupled with the application of base editing to create accurate genome edits in perennial plants. [ABSTRACT FROM AUTHOR]

Potassium (K) plays a crucial role in multiple physiological and developmental processes in plants. Its deficiency is a common abiotic stress that inhibits plant growth and reduces crop productivity. A better understanding of the mechanisms involved in plant responses to low K could help to improve the efficiency of K use in plants. However, such responses remain poorly characterized in fruit tree species such as pears (Pyrus sp). We analyzed the physiological and transcriptome responses of a commonly used pear rootstock, Pyrus betulaefolia, to K-deficiency stress (0 mM). Potassium deprivation resulted in apparent changes in root morphology, with short-term low-K stress resulting in rapidly enhanced root growth. Transcriptome analyses indicated that the root transcriptome was coordinately altered within 6 h after K deprivation, a process that continued until 15 d after treatment. Potassium deprivation resulted in the enhanced expression (up to 5-fold) of a putative high-affinity K+ transporter, PbHAK5 (Pbr037826.1), suggesting the up-regulation of mechanisms associated with K+ acquisition. The enhanced root growth in response to K-deficiency stress was associated with a rapid and sustained decrease in the expression of a transcription factor, PbMYB44 (Pbr015309.1), potentially involved in mediating auxin responses, and the increased expression of multiple genes associated with regulating root growth. The concentrations of several phytohormones including indoleacetic acid (IAA), ABA, ETH, gibberellin (GA3), and jasmonic acid (JA) were higher in response to K deprivation. Furthermore, genes coding for enzymes associated with carbon metabolism such as SORBITOL DEHYDROGENASE (SDH) and SUCROSE SYNTHASE (SUS) displayed greatly enhanced expression in the roots under K deprivation, presumably indicating enhanced metabolism to meet the increased energy demands for growth and K+ acquisition. Together, these data suggest that K deprivation in P. betulaefolia results in the rapid re-programming of the transcriptome to enhance root growth and K+ acquisition. These data provide key insights into the molecular basis for understanding low-K-tolerance mechanisms in pears and in other related fruit trees and identifying potential candidates that warrant further analyses. [ABSTRACT FROM AUTHOR]

Potentilla dickinsii var. glabrata and Spiraea insularis in the family Rosaceae are species endemic to Ulleung Island, Korea, the latter of which is listed as endangered. In this study, we characterized the complete plastomes of these two species and compared these with previously reported plastomes of other Ulleung Island endemic species of Rosaceae (Cotoneaster wilsonii, Prunus takesimensis, Rubus takesimensis, and Sorbus ulleungensis). The highly conserved complete plastomes of P. dickinsii var. glabrata and S. insularis are 158,637 and 155,524 base pairs with GC contents of 37% and 36.9%, respectively. Comparative phylogenomic analysis identified three highly variable intergenic regions (trnT-UGU/trnL-UAA, rpl32/trnL-UAG, and ndhF/rpl32) and one variable genic region (ycf1). Only 6 of the 75 protein-coding genes have been subject to strong positive selection. Phylogenetic analysis of 23 representative plastomes within the Rosaceae supported the monophyly of Potentilla and the sister relationship between Potentilla and Fragaria and indicated that S. insularis is sister to a clade containing Cotoneaster, Malus, Pyrus, and Sorbus. The plastome resources generated in this study will contribute to elucidating the plastome evolution of insular endemic Rosaceae on Ulleung Island and also in assessing the genetic consequences of anagenetic speciation for various endemic lineages on the island. [ABSTRACT FROM AUTHOR]

The glycogen synthase kinase 3/shaggy kinase (GSK3) is a serine/threonine kinase that plays important roles in brassinosteroid signaling, abiotic stress responses, cell division, and elongation, etc. In this study, we characterized seven grape GSK3 genes, showing high similarities with homologs from other species including Arabidopsis, white pear, apple, orange, and peach. Gene chip microarray data derived from an online database revealed very diverse developmental and tissue-specific expression patterns of VvSKs. VvSK3 and VvSK7 showed much higher expression levels in almost every tissue compared with other members. VvSK7 was highly enriched in young tissues like berries before the veraison stage, young leaves and green stems, etc., but immediately downregulated after these tissues entered maturation or senescence phases. Prediction of cis-elements in VvSK promoters indicated that VvSKs might be sensitive to light stimulation, which is further confirmed by the qPCR data. Constitutive overexpression of VvSK7 in Arabidopsis leads to dwarf plants that resembles BR-deficient mutants. The photosynthetic rate was significantly reduced in these plants, even though they accumulated more chlorophyll in leaves. Transient overexpression of VvSKs in tomatoes delayed the fruit ripening process, consistent with the observation in grapevine which blocks VvSKs by EBR- or BIKININ-promoted berry expansion and soluble solids accumulation. Data presented in the current study may serve as a theoretical basis for the future application of BRs or related compounds in quality grape production. [ABSTRACT FROM AUTHOR]

Anthocyanin biosynthesis exhibits a rhythmic oscillation pattern in some plants. To investigate the correlation between the oscillatory regulatory network and anthocyanin biosynthesis in pear, the anthocyanin accumulation and the expression patterns of anthocyanin late biosynthetic genes (ALBGs) were investigated in fruit skin of 'Red Zaosu' (Pyrus bretschneideri Rehd.). The anthocyanin accumulated mainly during the night over three continuous days in the fruit skin, and the ALBGs' expression patterns in 'Red Zaosu' fruit skin were oscillatory. However, the expression levels of typical anthocyanin-related transcription factors did not follow this pattern. Here, we found that the expression patterns of four PbREVEILLEs (PbRVEs), members of a class of atypical anthocyanin-regulated MYBs, were consistent with those of ALBGs in 'Red Zaosu' fruit skin over three continuous days. Additionally, transient expression assays indicated that the four PbRVEs promoted anthocyanin biosynthesis by regulating the expression of the anthocyanin biosynthetic genes encoding dihydroflavonol-4-reductase (DFR) and anthocyanidin synthase (ANS) in red pear fruit skin, which was verified using a dual-luciferase reporter assay. Moreover, a yeast one-hybrid assay indicated that PbRVE1a, 1b and 7 directly bound to PbDFR and PbANS promoters. Thus, PbRVEs promote anthocyanin accumulation at night by up-regulating the expression levels of PbDFR and PbANS in 'Red Zaosu' fruit skin. [ABSTRACT FROM AUTHOR]

Subgroup 4 R2R3 MYBs play vital roles in the regulation of anthocyanin biosynthesis. However, there is limited knowledge regarding the functions of MYB repressors in pear (Pyrus × bretschneideri). Here, PbMYB120 was identified as a potential regulator of anthocyanin biosynthesis. A phylogenetic analysis revealed that PbMYB120 was clustered into the FaMYB1-like clade of the subgroup 4 R2R3 MYBs. PbMYB120 was expressed higher in red peels than in green peels in five pear cultivars. PbMYB120 expression was positively correlated with anthocyanin accumulation. However, the transient overexpression of PbMYB120 led to the inhibition of anthocyanin accumulation and PbUFGT1 expression. Promoter binding and activation assays indicated that PbMYB120 binds to the promoter of PbUFGT1 and represses the promoter's activity. Thus, the inhibition of anthocyanin accumulation by PbMYB120 may be correlated with the repression of PbUFGT1. Furthermore, during anthocyanin induction, the expression levels of anthocyanin activators and PbMYB120 were upregulated. This study demonstrated that PbMYB120 was highly expressed in pear tissues having higher anthocyanin accumulations but acted as a repressor in the regulation of anthocyanin accumulation. PbMYB120 may work coordinately with anthocyanin activators and serve as a balancer of anthocyanin accumulation. [ABSTRACT FROM AUTHOR]

The development of dwarf fruit trees with smaller and compact characteristics leads to significantly increased fruit production, which is a major objective of pear (Pyrus bretschneideri) breeding. We identified the S-acylation activity of PbPAT14, an S-acyltransferase gene related to plant development, using a yeast (Saccharomyces cerevisiae) complementation assay, and also PbPAT14 could rescue the growth defect of the Arabidopsis mutant atpat14. We further studied the function of PbPAT14 by designing three guide RNAs for PbPAT14 to use in the CRISPR/Cas9 system. We obtained 22 positive transgenic pear lines via Agrobacterium-mediated transformation using cotyledons from seeds of Pyrus betulifolia ('Duli'). Six of these lines exhibited the dwarf yellowing phenotype and were homozygous mutations according to sequencing analysis. Ultrastructure analysis suggested that this dwarfism was manifested by shorter, thinner stems due to a reduction in cell number. A higher level of endogenous abscisic acid (ABA) and a higher transcript level of the ABA pathway genes in the mutant lines revealed that the PbPAT14 function was related to the ABA pathway. Overall, our experimental results increase the understanding of how PATs function in plants and help elucidate the mechanism of plant dwarfism. [ABSTRACT FROM AUTHOR]

C-repeat binding factor/dehydration-responsive element (CBF/DRE) transcription factors (TFs) participate in a variety of adaptive mechanisms, and are involved in molecular signaling and abiotic stress tolerance in plants. In pear (Pyrus pyrifolia) and other rosaceous crops, the independent evolution of CBF subfamily members requires investigation to understand the possible divergent functions of these proteins. In this study, phylogenetic analysis divided six PpyCBFs from the Asian pear genome into three clades/subtypes, and collinearity and phylogenetic analyses suggested that PpyCBF3 was the mother CBF. All PpyCBFs were found to be highly expressed in response to low temperature, salt, drought, and abscisic acid (ABA) as well as bud endodormancy, similar to PpyCORs (PpyCOR47, PpyCOR15A, PpyRD29A, and PpyKIN). Transcript levels of clade II PpyCBFs during low temperature and ABA treatments were higher than those of clades I and III. Ectopic expression of PpyCBF2 and PpyCBF3 in Arabidopsis enhanced its tolerance against abiotic stresses, especially to low temperature in the first case and salt and drought stresses in the latter, and resulted in lower reactive oxygen species (ROS) and antioxidant gene activities compared with the wild type. The increased expression of endogenous ABA-dependent and -independent genes during normal conditions in PpyCBF2- and PpyCBF3-overexpressing Arabidopsis lines suggested that PpyCBFs were involved in both ABA-dependent and -independent pathways. All PpyCBFs, especially the mother CBF, had high transactivation activities with 6XCCGAC binding elements. Luciferase and Y1H assays revealed the existence of phylogenetically and promoter-dependent conserved CBF–COR cascades in the pear. The presence of a previously identified CCGA binding site, combined with the results of mutagenesis of the CGACA binding site of the PpyCOR15A promoter, indicated that CGA was a core binding element of PpyCBFs. In conclusion, PpyCBF TFs might operate redundantly via both ABA-dependent and -independent pathways, and are strongly linked to abiotic stress signaling and responses in the Asian pear. [ABSTRACT FROM AUTHOR]

Self-incompatibility (SI) is a complex process, one out of several mechanisms that prevent plants from self-fertilizing to maintain and increase the genetic variability. This process leads to the rejection of the male gametophyte and requires the co-participation of numerous molecules. Plants have evolved two distinct SI systems, the sporophytic (SSI) and the gametophytic (GSI) systems. The two SI systems are markedly characterized by different genes and proteins and each single system can also be divided into distinct subgroups; whatever the mechanism, the purpose is the same, i.e., to prevent self-fertilization. In Malinae, a subtribe in the Rosaceae family, i.e., Pyrus communis and Malus domestica, the GSI requires the production of female determinants, known as S-RNases, which penetrate the pollen tube to interact with the male determinants. Beyond this, the penetration of S-RNase into the pollen tube triggers a series of responses involving membrane proteins, such as phospholipases, intracellular variations of cytoplasmic Ca2+, production of reactive oxygen species (ROS) and altered enzymatic activities, such as that of transglutaminase (TGase). TGases are widespread enzymes that catalyze the post-translational conjugation of polyamines (PAs) to different protein targets and/or the cross-linking of substrate proteins leading to the formation of cross-linked products with high molecular mass. When actin and tubulin are the substrates, this destabilizes the cytoskeleton and inhibits the pollen-tube's growth process. In this review, we will summarize the current knowledge of the relationship between S-RNase penetration, TGase activity and cytoskeleton function during GSI in the Malinae. [ABSTRACT FROM AUTHOR]

Pear (Pyrus L.) is an important commercial fruit in the world. The fruit size is one of the important characters in fruit quality. The previous research reported that the fruit size of pear was mainly caused by the number of cell in about 40 days after blossom (DAB) in nature. However, studies about the mechanisms underlying cell division in young fruit development are very limited in pear. Two pear accessions codenamed ‘GH59B’ with big fruit and ‘GH81S’ with small fruit in three stages were sampled and the RNA-seq high-throughput sequencing was used to evaluate changes of gene transcription levels in the early stage of fruit development. The difference of cell size among two samples was little in 40 DAB, implying that the difference of the fruit size was caused by the number of the cell. More than 274,517,982 high quality reads from six libraries of fruit development were sequenced. A total of 797 differentially expressed genes (DEGs) were identified. Three cytokinin dehydrogenase genes and two gibberellin 2-beta-dioxygenase gene were identified in the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways related to zeatin and gibberellin. Their expression was upregulated at 20 DAB in ‘GH81S’ and at 30 DAB in ‘GH59B’, suggesting that the small fruit size might be related to the early degradation of cytokinin and gibberellin inducing a short period of cell division. A total of 38 DEGs of transcription factors were found and 23 DEGs including NAC, ERF and bHLH transcription factors were highly related with cytokinin dehydrogenase and gibberellin dioxygenase genes. Altogether, the results of the present study provide information from a comprehensive gene expression analysis and insight into the molecular mechanism underlying the difference of fruit size in Pyrus pyrifolia. [ABSTRACT FROM AUTHOR]