Mitochondria-Specific Fluorescent Probe for Revealing the Interaction between Mitochondria and Lysosomes during Apoptosis.

The mitochondria, as one of the essential organelles in cells, are closely associated with numerous biological processes. Therefore, the realization of clear and real-time imaging for tracking mitochondria is of profound significance. Here, we present a mitochondria-targeting fluorescent probe, N(CH2)3-PD-NEt, for the real-time fluorescence imaging of mitochondria in living cells. Using the probe, the fluorescence changes of mitochondria stimulated by different drugs were successfully observed by fluorescence imaging. In addition, the dynamic processes of mitochondria and lysosomes during apoptosis were also explored. Importantly, we observed several novel dynamic interaction patterns between mitochondria and lysosomes. Among them, the most prominent pattern involved the noncontact movements of two lysosomes, that is, one lysosome gradually approached the other lysosome over time, eventually coming into contact and merging with it while gradually combining with mitochondria to form new mitochondria. Notably, the protrusions of the mitochondria became increasingly evident during this process. Meanwhile, we successfully observed the dynamic changes of mitochondria with SIM super-resolution imaging. The study provides promising help for the in-depth study of the dynamic processes of mitochondrial physiology and pathology and the study of the interactions between organelles.

Genome-wide analysis of MdPLATZ genes and their expression during axillary bud outgrowth in apple (Malus domestica Borkh.).

BACKGROUND: Branching is a plastic character that affects plant architecture and spatial structure. The trait is controlled by a variety of plant hormones through coordination with environmental signals. Plant AT-rich sequence and zinc-binding protein (PLATZ) is a transcription factor that plays an important role in plant growth and development. However, systematic research on the role of the PLATZ family in apple branching has not been conducted previously. RESULTS: In this study, a total of 17 PLATZ genes were identified and characterized from the apple genome. The 83 PLATZ proteins from apple, tomato, Arabidopsis, rice, and maize were classified into three groups based on the topological structure of the phylogenetic tree. The phylogenetic relationships, conserved motifs, gene structure, regulatory cis-acting elements, and microRNAs of the MdPLATZ family members were predicted. Expression analysis revealed that MdPLATZ genes exhibited distinct expression patterns in different tissues. The expression patterns of the MdPLATZ genes were systematically investigated in response to treatments that impact apple branching [thidazuron (TDZ) and decapitation]. The expression of MdPLATZ1, 6, 7, 8, 9, 15, and 16 was regulated during axillary bud outgrowth based on RNA-sequencing data obtained from apple axillary buds treated by decapitation or exogenous TDZ application. Quantitative real-time PCR analysis showed that MdPLATZ6 was strongly downregulated in response to the TDZ and decapitation treatments, however, MdPLATZ15 was significantly upregulated in response to TDZ, but exhibited little response to decapitation. Furthermore, the co-expression network showed that PLATZ might be involved in shoot branching by regulating branching-related genes or mediating cytokinin or auxin pathway. CONCLUSION: The results provide valuable information for further functional investigation of MdPLATZ genes in the control of axillary bud outgrowth in apple.

Strategy for Fluorescence/Photoacoustic Signal Maximization Using Dual-Wavelength-Independent Excitation.

Dual-mode imaging of fluorescence-photoacoustics has emerged as a promising technique for biomedical applications. However, conventional dual-mode imaging is based on single-wavelength excitation, which often results in opposing fluorescence and photoacoustic signals due to competing photophysical processes in one agent, rendering the maximization of both signals infeasible. To meet this challenge, we herein propose a new strategy by using the dual-excitation approach, where one excitation wavelength generates a fluorescence signal and the other produces a photoacoustic signal, thus achieving simultaneous maximization of both signals in one fluorescence-photoacoustic molecule. Based on this strategy, three dye molecules were employed for comparison, and it was surprising to find that QHD dye with two types of excitation wavelengths could generate fluorescence and photoacoustic signals, respectively. Furthermore, this strategy was successfully implemented in dual-mode imaging of rheumatoid arthritis mice. Importantly, this study emphasizes a new design guideline for the maximization of fluorescence-photoacoustic signals by using dual-wavelength-independent excitation.

Visualization of Lysosomal Dynamics during Autophagy by Fluorescent Probe.

Lysosomes are one of the important organelles within cells, and their dynamic movement processes are associated with many biological events. Therefore, real-time monitoring of lysosomal dynamics processes has far-reaching implications. A lysosome-targeted fluorescent probe N(CH2)3-BD-PZ is proposed for real-time monitoring of lysosomal kinetic motility. Using this probe, the dynamic process of lysosomes under starvation induction was successfully explored through fluorescence imaging. Importantly, we observed a new pattern of lysosomal dynamic movement, in which an irregular lysosome was slowly cleaved into two different-sized touching lysosomes and then fused to form a new round lysosome. This research provides a powerful fluorescence tool to understand the dynamic motility of intracellular lysosomes under fluorescence imaging.

A Lightweight Recurrent Grouping Attention Network for Video Super-Resolution.

Effective aggregation of temporal information of consecutive frames is the core of achieving video super-resolution. Many scholars have utilized structures such as sliding windows and recurrences to gather the spatio-temporal information of frames. However, although the performances of constructed video super-resolution models are improving, the sizes of the models are also increasing, exacerbating the demand on the equipment. Thus, to reduce the stress on the device, we propose a novel lightweight recurrent grouping attention network. The parameters of this model are only 0.878 M, which is much lower than the current mainstream model for studying video super-resolution. We have designed a forward feature extraction module and a backward feature extraction module to collect temporal information between consecutive frames from two directions. Moreover, a new grouping mechanism is proposed to efficiently collect spatio-temporal information of the reference frame and its neighboring frames. The attention supplementation module is presented to further enhance the information gathering range of the model. The feature reconstruction module aims to aggregate information from different directions to reconstruct high-resolution features. Experiments demonstrate that our model achieves state-of-the-art performance on multiple datasets.

Molecular mechanism of MdWUS2-MdTCP12 interaction in mediating cytokinin signaling to control axillary bud outgrowth.

Shoot branching is an important factor that influences the architecture of apple trees and cytokinin is known to promote axillary bud outgrowth. The cultivar 'Fuji', which is grown on ~75% of the apple-producing area in China, exhibits poor natural branching. The TEOSINTE BRANCHED1/CYCLOIDEA/PCF (TCP) family genes BRANCHED1/2 (BRC1/2) are involved in integrating diverse factors that function locally to inhibit shoot branching; however, the molecular mechanism underlying the cytokinin-mediated promotion of branching that involves the repression of BRC1/2 remains unclear. In this study, we found that apple WUSCHEL2 (MdWUS2), which interacts with the co-repressor TOPLESS-RELATED9 (MdTPR9), is activated by cytokinin and regulates branching by inhibiting the activity of MdTCP12 (a BRC2 homolog). Overexpressing MdWUS2 in Arabidopsis or Nicotiana benthamiana resulted in enhanced branching. Overexpression of MdTCP12 inhibited axillary bud outgrowth in Arabidopsis, indicating that it contributes to the regulation of branching. In addition, we found that MdWUS2 interacted with MdTCP12 in vivo and in vitro and suppressed the ability of MdTCP12 to activate the transcription of its target gene, HOMEOBOX PROTEIN 53b (MdHB53b). Our results therefore suggest that MdWUS2 is involved in the cytokinin-mediated inhibition of MdTCP12 that controls bud outgrowth, and hence provide new insights into the regulation of shoot branching by cytokinin.

MicroRNA-1224 Splicing CircularRNA-Filip1l in an Ago2-Dependent Manner Regulates Chronic Inflammatory Pain via Targeting Ubr5.

Dysfunctions of gene transcription and translation in the nociceptive pathways play the critical role in development and maintenance of chronic pain. Circular RNAs (circRNAs) are emerging as new players in regulation of gene expression, but whether and how circRNAs are involved in chronic pain remain elusive. We showed here that complete Freund's adjuvant-induced chronic inflammation pain significantly increased circRNA-Filip1l (filamin A interacting protein 1-like) expression in spinal neurons of mice. Blockage of this increase attenuated complete Freund's adjuvant-induced nociceptive behaviors, and overexpression of spinal circRNA-Filip1l in naive mice mimicked the nociceptive behaviors as evidenced by decreased thermal and mechanical nociceptive threshold. Furthermore, we found that mature circRNA-Filip1l expression was negatively regulated by miRNA-1224 via binding and splicing of precursor of circRNA-Filip1l (pre-circRNA-Filip1l) in the Argonaute-2 (Ago2)-dependent manner. Increase of spinal circRNA-Filip1l expression resulted from the decrease of miRNA-1224 expression under chronic inflammation pain state. miRNA-1224 knockdown or Ago2 overexpression induced nociceptive behaviors in naive mice, which was prevented by the knockdown of spinal circRNA-Filip1l. Finally, we demonstrated that a ubiquitin protein ligase E3 component n-recognin 5 (Ubr5), validated as a target of circRNA-Filip1l, plays a pivotal role in regulation of nociception by spinal circRNA-Filip1l. These data suggest that miRNA-1224-mediated and Ago2-dependent modulation of spinal circRNA-Filip1l expression regulates nociception via targeting Ubr5, revealing a novel epigenetic mechanism of interaction between miRNA and circRNA in chronic inflammation pain.SIGNIFICANCE STATEMENT circRNAs are emerging as new players in regulation of gene expression. Here, we found that the increase of circRNA-Filip1l mediated by miRNA-1224 in an Ago2-dependent way in the spinal cord is involved in regulation of nociception via targeting Ubr5 Our study reveals a novel epigenetic mechanism of interaction between miRNA and circRNA in chronic inflammation pain.

Molecular role of cytokinin in bud activation and outgrowth in apple branching based on transcriptomic analysis.

KEY MESSAGE: Axillary bud activation and outgrowth were dependent on local cytokinin, and that bud activation preceded the activation of cell cycle and cell growth genes in apple branching. Cytokinin is often applied to apple trees to produce more shoot branches in apple seedlings. The molecular response of apple to the application of cytokinin, and the relationship between bud activation and cell cycle in apple branching, however, are poorly understood. In this study, RNA sequencing was used to characterize differential expression genes in axillary buds of 1-year grafted "Fuji" apple at 4 and 96 h after cytokinin application. And comparative gene expression analyses were performed in buds of decapitated shoots and buds of the treatment of biosynthetic inhibitor of cytokinin (Lovastatin) on decapitated shoots. Results indicated that decapitation and cytokinin increased ZR content in buds and internodes at 4-8 h, and induced bud elongation at 96 h after treatment, relative to buds in shoots receiving the Lovastatin treatment. RNA-seq analysis indicated that differential expression genes in auxin and cytokinin signal transduction were significantly enriched at 4 h, and DNA replication was enriched at 96 h. Cytokinin-responsive type-A response regulator, auxin polar transport, and axillary meristem-related genes were up-regulated at 4 h in the cytokinin and decapitation treatments, while qRT-PCR analysis showed that cell cycle and cell growth genes were up-regulated after 8 h. Collectively, the data indicated that bud activation and outgrowth might be dependent on local cytokinin synthesis in axillary buds or stems, and that bud activation preceded the activation of cell cycle genes during the outgrowth of ABs in apple shoots.

Exogenous application of GA3 inactively regulates axillary bud outgrowth by influencing of branching-inhibitors and bud-regulating hormones in apple (Malus domestica Borkh.).

Although gibberellin (GA) has been reported to control branching, little is known about how GA mediates signals regulating the outgrowth of axillary buds (ABs). In the current study, the effect of the exogenous application of 5.0 mM GA3 on ABs outgrowth on 1-year-old 'Nagafu No. 2'/T337/M. robusta Rehd. apple trees was investigated and compared to the bud-activating treatments, 5 mM BA or decapitation. Additionally, the expression of genes related to bud-regulating signals and sucrose levels in ABs was examined. Results indicated that GA3 did not promote ABs' outgrowth, nor down-regulate the expression of branching repressors [MdTCP40, MdTCP33, and MdTCP16 (homologs of BRANCHED1 and BRC2)], which were significantly inhibited by the BA and decapitation treatments. MdSBP12 and MdSBP18, the putative transcriptional activators of these genes, which are expressed at lower levels in BA-treated and decapitated buds, were up-regulated in the GA3 treatment in comparison to the BA treatment. Additionally, GA3 did not up-regulate the expression of CK response- and auxin transport-related genes, which were immediately induced by the BA treatment. In addition, GA3 also up-regulated the expression of several Tre6P biosynthesis genes and reduced sucrose levels in ABs. Sucrose levels, however, were still higher than what was observed in BA-treated buds, indicating that sucrose may not be limiting in GA3-controlled AB outgrowth. Although GA3 promoted cell division, it was not sufficient to induce AB outgrowth. Conclusively, some branching-inhibiting genes and bud-regulating hormones are associated with the inability of GA3 to activate AB outgrowth.

Genome Identification of B-BOX Gene Family Members in Seven Rosaceae Species and Their Expression Analysis in Response to Flower Induction in Malus domestica.

BBX proteins play important roles in regulating plant growth and development including photomorphogenesis, photoperiodic regulation of flowering, and responses to biotic and abiotic stresses. At present, the genomes of seven Rosaceae fruit species have been fully sequenced. However, little is known about the BBX gene family and their evolutionary history in these Rosaceae species. Therefore, in this study total, 212 BBX genes were investigated from seven Rosaceae species (67 from Malus × domestica, 40 from Pyruscommunis, 22 from Rosa Chinesis, 20 from Prunuspersica, 21 from Fragariavesca, 22 from Prunusavium, and 20 from Rubusoccidentalis). The chemical properties, gene structures, and evolutionary relationships of the BBX genes were also studied. All the BBX genes were grouped into six subfamilies on the basis of their phylogenetic relationships and structural features. Analysis of gene structure, segmental and tandem duplication, gene phylogeny, and tissue-specific expression with the ArrayExpress database showed their diversification in function, quantity, and structure. The expression profiles of 19 MdBBX genes in different tissues were evaluated through qRT-PCR. These genes showed distinct transcription level among the tested tissues (bud, flower, fruit, stem, and leaf). Moreover, expression patterns of 19 MdBBX genes were examined during flowering induction time under flowering-related hormones and treatments (GA3, 6-BA, and sucrose). The expressions of the candidates BBX genes were affected and showed diverse expression profile. Furthermore, changes in response to these flowering-related hormones and treatment specifying their potential involvement in flowering induction. Based on these findings, BBX genes could be used as potential genetic markers for the growth and development of plants particularly in the area of functional analysis, and their involvement in flower induction in fruit plants.

Hydroxymethylation of microRNA-365-3p Regulates Nociceptive Behaviors via Kcnh2.

DNA 5-hydroxylmethylcytosine (5hmC) catalyzed by ten-eleven translocation methylcytosine dioxygenase (TET) occurs abundantly in neurons of mammals. However, the in vivo causal link between TET dysregulation and nociceptive modulation has not been established. Here, we found that spinal TET1 and TET3 were significantly increased in the model of formalin-induced acute inflammatory pain, which was accompanied with the augment of genome-wide 5hmC content in spinal cord. Knockdown of spinal TET1 or TET3 alleviated the formalin-induced nociceptive behavior and overexpression of spinal TET1 or TET3 in naive mice produced pain-like behavior as evidenced by decreased thermal pain threshold. Furthermore, we found that TET1 or TET3 regulated the nociceptive behavior by targeting microRNA-365-3p (miR-365-3p). Formalin increased 5hmC in the miR-365-3p promoter, which was inhibited by knockdown of TET1 or TET3 and mimicked by overexpression of TET1 or TET3 in naive mice. Nociceptive behavior induced by formalin or overexpression of spinal TET1 or TET3 could be prevented by downregulation of miR-365-3p, and mimicked by overexpression of spinal miR-365-3p. Finally, we demonstrated that a potassium channel, voltage-gated eag-related subfamily H member 2 (Kcnh2), validated as a target of miR-365-3p, played a critical role in nociceptive modulation by spinal TET or miR-365-3p. Together, we concluded that TET-mediated hydroxymethylation of miR-365-3p regulates nociceptive behavior via Kcnh2. SIGNIFICANCE STATEMENT: Mounting evidence indicates that epigenetic modifications in the nociceptive pathway contribute to pain processes and analgesia response. Here, we found that the increase of 5hmC content mediated by TET1 or TET3 in miR-365-3p promoter in the spinal cord is involved in nociceptive modulation through targeting a potassium channel, Kcnh2. Our study reveals a new epigenetic mechanism underlying nociceptive information processing, which may be a novel target for development of antinociceptive drugs.

DNA Hydroxymethylation by Ten-eleven Translocation Methylcytosine Dioxygenase 1 and 3 Regulates Nociceptive Sensitization in a Chronic Inflammatory Pain Model.

BACKGROUND: Ten-eleven translocation methylcytosine dioxygenase converts 5-methylcytosine in DNA to 5-hydroxymethylcytosine, which plays an important role in gene transcription. Although 5-hydroxymethylcytosine is enriched in mammalian neurons, its regulatory function in nociceptive information processing is unknown. METHODS: The global levels of 5-hydroxymethylcytosine and ten-eleven translocation methylcytosine dioxygenase were measured in spinal cords in mice treated with complete Freund's adjuvant. Immunoblotting, immunohistochemistry, and behavioral tests were used to explore the downstream ten-eleven translocation methylcytosine dioxygenase-dependent signaling pathway. RESULTS: Complete Freund's adjuvant-induced nociception increased the mean levels (± SD) of spinal 5-hydroxymethylcytosine (178 ± 34 vs. 100 ± 21; P = 0.0019), ten-eleven translocation methylcytosine dioxygenase-1 (0.52 ± 0.11 vs. 0.36 ± 0.064; P = 0.0088), and ten-eleven translocation methylcytosine dioxygenase-3 (0.61 ± 0.13 vs. 0.39 ± 0.08; P = 0.0083) compared with levels in control mice (n = 6/group). The knockdown of ten-eleven translocation methylcytosine dioxygenase-1 or ten-eleven translocation methylcytosine dioxygenase-3 alleviated thermal hyperalgesia and mechanical allodynia, whereas overexpression cytosinethem in naïve mice (n = 6/group). Down-regulation of spinal ten-eleven translocation methylcytosine dioxygenase-1 and ten-eleven translocation methylcytosine dioxygenase-3 also reversed the increases in Fos expression (123 ± 26 vs. 294 ± 6; P = 0.0031; and 140 ± 21 vs. 294 ± 60; P = 0.0043, respectively; n = 6/group), 5-hydroxymethylcytosine levels in the Stat3 promoter (75 ± 16.1 vs. 156 ± 28.9; P = 0.0043; and 91 ± 19.1 vs. 156 ± 28.9; P = 0.0066, respectively; n = 5/group), and consequent Stat3 expression (93 ± 19.6 vs. 137 ± 27.5; P = 0.035; and 72 ± 15.2 vs. 137 ± 27.5; P = 0.0028, respectively; n = 5/group) in complete Freund's adjuvant-treated mice. CONCLUSIONS: This study reveals a novel epigenetic mechanism for ten-eleven translocation methylcytosine dioxygenase-1 and ten-eleven translocation methylcytosine dioxygenase-3 in the modulation of spinal nociceptive information via targeting of Stat3.

Transcriptome analysis reveals the effects of sugar metabolism and auxin and cytokinin signaling pathways on root growth and development of grafted apple.

BACKGROUND: The root architecture of grafted apple (Malus spp.) is affected by various characteristics of the scions. To provide information on the molecular mechanisms underlying this influence, we examined root transcriptomes of M. robusta rootstock grafted with scions of wild-type (WT) apple (M. spectabilis) and a more-branching (MB) mutant at the branching stage. RESULTS: The growth rate of rootstock grafted MB was repressed significantly, especially the primary root length and diameter, and root weight. Biological function categories of differentially expressed genes were significantly enriched in processes associated with hormone signal transduction and intracellular activity, with processes related to the cell cycle especially down-regulated. Roots of rootstock grafted with MB scions displayed elevated auxin and cytokinin contents and reduced expression of MrPIN1, MrARF, MrAHP, most MrCRE1 genes, and cell growth-related genes MrGH3, MrSAUR and MrTCH4. Although auxin accumulation and transcription of MrPIN3, MrALF1 and MrALF4 tended to induce lateral root formation in MB-grafted rootstock, the number of lateral roots was not significantly changed. Sucrose, fructose and glucose contents were not decreased in MB-grafted roots compared with those bearing WT scions, but glycolysis and tricarboxylic acid cycle metabolic activities were repressed. Root resistance and nitrogen metabolism were reduced in MB-grafted roots as well. CONCLUSIONS: Our findings suggest that root growth and development of rootstock are mainly influenced by sugar metabolism and auxin and cytokinin signaling pathways. This study provides a basis that the characteristics of scions are related to root growth and development, resistance and activity of rootstocks.

Decolourization of azo dyes by a newly isolated Klebsiella sp. strain Y3, and effects of various factors on biodegradation.

In this study, we isolated and characterized a new strain of Klebsiella sp. Y3, which was capable of decolourizing azo dyes under anaerobic conditions. The effects of physico-chemical parameters on the Methyl Red degradation by the strain were determined. The results indicated that strain Y3 exhibited a good decolourization ability in the range of pH from 4 to 9, temperature from 30 °C to 42 °C and salinity from 1% to 4%. A broad spectrum of azo dyes with different structures could be decolourized by the strain. The isolate decolourized Methyl Red, Congo Red, Orange I and Methyl Orange by almost 100% (100 mg/L) in 48 h. The culture exhibited an ability to decolourize repeated additions of dye, showing that the strain could be used for multiple cycles of biodegradation. Azo dyes at high concentrations could be tolerated and degraded by Y3. An almost complete mineralization of Methyl Red and Congo Red at the concentration of 800 mg/L was observed within 48 h. The high degradation potential of this bacterium supports its use in the treatment of industrial wastewater containing azo dyes.

Ultrathin two-dimensional (2D) manganese-based metal-organic framework nanosheets for selective photocatalytic oxidation of thioether.

The efficiency of photocatalysts depends largely on the accessibility of reaction species to the active centre, the electron transfer and geometric matching between the active surface of the catalyst and reaction species. In this work, we successfully synthesized and designed one two-dimensional Mn(II) MOF with [Mn2(H2L1)(H2O)2(DMF)2]n·(CH3CH2OH)n (HSTC 3) by using MnCl2·4H2O and 5,5'-(anthracene-9,10-diyl)diisophthalic acid (H4L1), in which the adjacent layers are stacked with weak interactions, and the huge gap leads to the interpenetration between layers to form a 2D + 2D → 3D interpenetration frame. Based on the particularity of the structure of HSTC 3, ultrasonic wall breaking methods were tried to successfully peel HSTC 3 into nanosheets (HSTC 3-NS), thus achieving a significant improvement in a series of optoelectronic properties due to exposure to more active centres for HSTC 3-NS. These results significantly enhance the photocatalytic selective oxidation of thioether. This study provides a new insight into the post-synthesis modification of MOF photocatalyst and their application in photocatalytic organic synthesis.

Proton-coupled electron transfer and Lewis acid recognition at self-assembled monolayers of an oxo-bridged diruthenium(III) complex functionalized with two disulfide anchors.

A new µ-oxo-bis(µ-acetato)diruthenium(III) complex bearing two pyridyl disulfide ligands {[Ru2(µ-O)(µ-OAc)2(bpy)2(L(py-SS))2](PF6)2 (OAc = CH3CO2(-), bpy = 2,2'-bipyridine, L(py-SS) = (C5H4N)CH2NHC(O)(CH2)4CH(CH2)2SS) (1)} has been synthesized to prepare self-assembled monolayers (SAMs) on the Au(111) electrode surface. The SAMs have been characterized by contact-angle measurements, reflection-absorption surface infrared spectroscopy, cyclic voltammetry, and reductive desorption experiments. The SAMs exhibited proton-coupled electron transfer (PCET) reactions when the electrochemistry was studied in aqueous electrolyte solution (0.1 M NaClO4 with Britton-Robinson buffer to adjust the solution pH). The potential-pH plot (Pourbaix diagram) in the pH range from 1 to 12 has established that the dinuclear ruthenium moiety was involved in the interfacial PCET processes with four distinct redox states: Ru(III)Ru(III)(µ-O), Ru(II)Ru(III)(µ-OH), Ru(II)Ru(II)(µ-OH), and Ru(II)Ru(II)(µ-OH2). We also demonstrated that the interfacial redox processes were modulated by the addition of Lewis acids such as BF3 or Al(3+) to the electrolyte media, in which the externally added Lewis acids interacted with µ-O of the dinuclear moiety within the SAMs.

ASSVd infection inhibits the vegetative growth of apple trees by affecting leaf metabolism.

Apple scar skin viroid (ASSVd) can infect apple trees and cause scar skin symptoms. However, the associated physiological mechanisms are unclear in young saplings. In this study, ASSVd-infected and control 'Odysso' and 'Tonami' apple saplings were examined to clarify the effects of ASSVd on apple tree growth and physiological characteristics as well as the leaf metabolome. The results indicated that leaf ASSVd contents increased significantly after grafting and remained high in the second year. Leaf size, tree height, stem diameter, branch length, and leaf photosynthetic efficiency decreased significantly in viroid-infected saplings. In response to the ASSVd infection, the chlorophyll a and b contents decreased significantly in 'Odysso', but were unchanged in 'Tonami'. Moreover, the N, P, K, Fe, Mn, and Ca contents decreased significantly in the leaves of viroid-infected 'Odysso' or 'Tonami'. Similarly, the CAT and POD contents decreased significantly in the viroid-infected saplings, but the SOD content increased in the viroid-infected 'Tonami' saplings. A total of 15 and 40 differentially abundant metabolites were respectively identified in the metabolome analyses of 'Odysso' and 'Tonami' leaves. Specifically, in the viroid-infected 'Odysso' and 'Tonami' samples, the L-2-aminobutyric acid, 6″-O-malonyldaidzin, and D-xylose contents increased, while the coumarin content decreased. These metabolites are related to the biosynthesis of isoflavonoids and phenylpropanoids as well as the metabolism of carbohydrates and amino acids. These results imply that ASSVd affects apple sapling growth by affecting physiological characteristics and metabolism of apple leaves. The study data may be useful for future investigations on the physiological mechanisms underlying apple tree responses to ASSVd.

Volatile metabolome and floral transcriptome analyses reveal the volatile components of strongly fragrant progeny of Malus × robusta.

Floral fragrance is an important trait that contributes to the ornamental properties and pollination of crabapple. However, research on the physiological and molecular biology of the floral volatile compounds of crabapple is rarely reported. In this study, metabolomic and transcriptomic analyses of the floral volatile compounds of standard Malus robusta flowers (Mr), and progeny with strongly and weakly fragrant flowers (SF and WF, respectively), were conducted. Fifty-six floral volatile compounds were detected in the plant materials, mainly comprising phenylpropane/benzene ring-type compounds, fatty acid derivatives, and terpene compounds. The volatile contents were significantly increased before the early flowering stage (ES), and the contents of SF flowers were twice those of WF and Mr flowers. Odor activity values were determined for known fragrant volatiles and 10-11 key fragrant volatiles were identified at the ES. The predominant fragrant volatiles were methyl benzoate, linalool, leaf acetate, and methyl anthranilate. In the petals, stamens, pistil, and calyx of SF flowers, 26 volatiles were detected at the ES, among which phenylpropane/benzene ring-type compounds were the main components accounting for more than 75% of the total volatile content. Functional analysis of transcriptome data revealed that the phenylpropanoid biosynthesis pathway was significantly enriched in SF flowers. By conducting combined analyses between volatiles and differentially expressed genes, transcripts of six floral scent-related genes were identified and were associated with the contents of the key fragrant volatiles, and other 23 genes were potentially correlated with the key volatile compounds. The results reveal possible mechanisms for the emission of strong fragrance by SF flowers, and provide a foundation for improvement of the floral fragrance and development of new crabapple cultivars.

Chromatin-associated OGT promotes the malignant progression of hepatocellular carcinoma by activating ZNF263.

Reversible and dynamic O-GlcNAcylation regulates vast networks of highly coordinated cellular and nuclear processes. Although dysregulation of the sole enzyme O-GlcNAc transferase (OGT) was shown to be associated with the progression of hepatocellular carcinoma (HCC), the mechanisms by which OGT controls the cis-regulatory elements in the genome and performs transcriptional functions remain unclear. Here, we demonstrate that elevated OGT levels enhance HCC proliferation and metastasis, in vitro and in vivo, by orchestrating the transcription of numerous regulators of malignancy. Diverse transcriptional regulators are recruited by OGT in HCC cells undergoing malignant progression, which shapes genome-wide OGT chromatin cis-element occupation. Furthermore, an unrecognized cooperation between ZNF263 and OGT is crucial for activating downstream transcription in HCC cells. We reveal that O-GlcNAcylation of Ser662 is responsible for the chromatin association of ZNF263 at candidate gene promoters and the OGT-facilitated HCC malignant phenotypes. Our data establish the importance of aberrant OGT activity and ZNF263 O-GlcNAcylation in the malignant progression of HCC and support the investigation of OGT as a therapeutic target for HCC.