MicroRNA156ab regulates apple plant growth and drought tolerance by targeting transcription factor MsSPL13.

Drought stress substantially reduces the productivity of apple plants and severely restricts the development of apple industry. Malus sieversii, wild apples with excellent drought resistance, is a valuable wild resource for a rootstock improvement of cultivated apple (Malus domestica). miRNAs and their targets play essential roles in plant growth and stress responses, but their roles in drought stress responses in apple are unknown. Here, we demonstrate that microRNA156ab is upregulated in M. sieversii in response to drought stress. Overexpressing msi-miR156ab promoted auxin accumulation, maintained the growth of apple plants, and increased plant resistance to osmotic stress. Antioxidant enzyme activities and proline contents were also increased in miR156ab-OE transgenic apple lines, which improved drought resistance. The squamosa promoter binding protein-like transcription factor MsSPL13 is the target of msi-miR156ab, as demonstrated by 5'-RACE and dual luciferase assays. Heterologous expression of MsSPL13 decreased auxin contents and inhibited growth in Arabidopsis (Arabidopsis thaliana) under normal and stress conditions. The activities of antioxidant enzymes were also suppressed in MsSPL13-OE transgenic Arabidopsis, reducing drought resistance. We showed that MsSPL13 regulates the expression of the auxin-related genes MsYUCCA5, PIN-FORMED7 (MsPIN7), and Gretchen Hagen3-5 (MsGH3-5) by binding to the GTAC cis-elements in their promoters, thereby regulating auxin metabolism. Finally, we demonstrated that the miR156ab-SPL13 module is involved in mediating the difference in auxin metabolism and stress responses between M. sieversii and M26 (M. domestica) rootstocks. Overall, these findings reveal that the miR156ab-SPL13 module enhances drought stress tolerance in apples by regulating auxin metabolism and antioxidant enzyme activities.

Two B-box proteins, PavBBX6/9, positively regulate light-induced anthocyanin accumulation in sweet cherry.

Anthocyanin production in bicolored sweet cherry (Prunus avium cv. Rainier) fruit is induced by light exposure, leading to red coloration. The phytohormone abscisic acid (ABA) is essential for this process, but the regulatory relationships that link light and ABA with anthocyanin-associated coloration are currently unclear. In this study, we determined that light treatment of bicolored sweet cherry fruit increased anthocyanin accumulation and induced ABA production and that ABA participates in light-modulated anthocyanin accumulation in bicolored sweet cherry. Two B-box (BBX) genes, PavBBX6/9, were highly induced by light and ABA treatments, as was anthocyanin accumulation. The ectopic expression of PavBBX6 or PavBBX9 in Arabidopsis (Arabidopsis thaliana) increased anthocyanin biosynthesis and ABA accumulation. Overexpressing PavBBX6 or PavBBX9 in sweet cherry calli also enhanced light-induced anthocyanin biosynthesis and ABA accumulation. Additionally, transient overexpression of PavBBX6 or PavBBX9 in sweet cherry peel increased anthocyanin and ABA contents, whereas silencing either gene had the opposite effects. PavBBX6 and PavBBX9 directly bound to the G-box elements in the promoter of UDP glucose-flavonoid-3-O-glycosyltransferase (PavUFGT), a key gene for anthocyanin biosynthesis, and 9-cis-epoxycarotenoid dioxygenase 1 (PavNCED1), a key gene for ABA biosynthesis, and enhanced their activities. These results suggest that PavBBX6 and PavBBX9 positively regulate light-induced anthocyanin and ABA biosynthesis by promoting PavUFGT and PavNCED1 expression, respectively. Our study provides insights into the relationship between the light-induced ABA biosynthetic pathway and anthocyanin accumulation in bicolored sweet cherry fruit.

bZIP Transcription Factor PavbZIP6 Regulates Anthocyanin Accumulation by Increasing Abscisic Acid in Sweet Cherry.

Basic leucine zipper (bZIP) transcription factors (TFs) play a crucial role in anthocyanin accumulation in plants. In addition to bZIP TFs, abscisic acid (ABA) increases anthocyanin biosynthesis. Therefore, this study aimed to investigate whether bZIP TFs are involved in ABA-induced anthocyanin accumulation in sweet cherry and elucidate the underlying molecular mechanisms. Specifically, the BLAST method was used to identify bZIP genes in sweet cherry. Additionally, we examined the expression of ABA- and anthocyanin-related genes in sweet cherry following the overexpression or knockdown of a bZIP candidate gene. In total, we identified 54 bZIP-encoding genes in the sweet cherry genome. Basic leucine zipper 6 (bZIP6) showed significantly increased expression, along with increased anthocyanin accumulation in sweet cherry. Additionally, yeast one-hybrid and dual-luciferase assays indicated that PavbZIP6 enhanced the expression of anthocyanin biosynthetic genes (PavDFR, PavANS, and PavUFGT), thereby increasing anthocyanin accumulation. Moreover, PavbZIP6 interacted directly with the PavBBX6 promoter, thereby regulating PavNCED1 to promote abscisic acid (ABA) synthesis and enhance anthocyanin accumulation in sweet cherry fruit. Conclusively, this study reveals a novel mechanism by which PavbZIP6 mediates anthocyanin biosynthesis in response to ABA and contributes to our understanding of the mechanism of bZIP genes in the regulation of anthocyanin biosynthesis in sweet cherry.

Abscisic acid-responsive transcription factors PavDof2/6/15 mediate fruit softening in sweet cherry.

Softening is a key step during fruit ripening that is modulated by the interplay between multiple phytohormones. The antagonistic action of abscisic acid (ABA) and auxin determines the rate of fruit ripening and softening. However, the transcription factors that integrate ABA and auxin signals to regulate fruit softening remain to be determined. In this study, we identified several DNA-binding with One Finger (Dof) transcription factors essential for ABA-promoted fruit softening, based on transcriptome analysis of two sweet cherry (Prunus avium L.) varieties with different fruit firmness. We show that PavDof6 directly binds to the promoters of genes encoding cell wall-modifying enzymes to activate their transcription, while PavDof2/15 directly repress their transcription. Transient overexpression of PavDof6 and PavDof2/15 in sweet cherry fruits resulted in precocious and delayed softening, respectively. In addition, we show that the auxin response factor PavARF8, the expression of whose encoding gene is repressed by ABA, activates PavDof2/15 transcription. Furthermore, PavDof2/6/15 and PavARF8 directly bind to the 9-cis-epoxycarotenoid dioxygenase 1 (PavNCED1) promoter and regulate its expression, forming a feedback mechanism for ABA-mediated fruit softening. These findings unveil the physiological framework of fruit softening and establish a direct functional link between the ABA-PavARF8-PavDofs module and cell-wall-modifying genes in mediating fruit softening.

Genome-Wide Identification of the SQUAMOSA Promoter-Binding Protein-like (SPL) Transcription Factor Family in Sweet Cherry Fruit.

Plant-specific SQUAMOSA promoter-binding protein-like (SPL) transcription factors play important regulatory roles during plant growth and development, fruit ripening, inflorescence branching, and biotic and abiotic stresses. However, there have been no identification or systematic studies of the SPL gene family in the sweet cherry. In this study, 12 SPL genes were identified in the sweet cherry reference genome, which were distributed over 6 chromosomes and classified into six groups according to phylogenetic relationships with other SPL gene families. Nine PavSPLs were highly expressed at green fruit stages and dramatically decreased at the onset of fruit ripening, which implied that they were important regulators during fruit development and ripening. The expression patterns of PavSPL genes under ABA, GA, and MeJA treatments showed that the PavSPLs were involved in the process of fruit ripening. A subcellular localization experiment proved that PavSPL4 and PavSPL7 proteins were localized in the nucleus. The genome-wide identification of the SPL gene family provided new insights while establishing an important foundation for sweet cherry studies.

The Apple microR171i-SCARECROW-LIKE PROTEINS26.1 Module Enhances Drought Stress Tolerance by Integrating Ascorbic Acid Metabolism.

Drought stress severely restricts crop yield and quality. Small noncoding RNAs play critical roles in plant growth, development, and stress responses by regulating target gene expression, but their roles in drought stress tolerance in apple (Malus domestica) are poorly understood. Here, we identified various small noncoding RNAs and their targets from the wild apple species Malus sieversii via high-throughput sequencing and degradome analysis. Forty known microRNAs (miRNAs) and eight new small noncoding RNAs were differentially expressed in response to 2 or 4 h of drought stress treatment. We experimentally verified the expression patterns of five selected miRNAs and their targets. We established that one miRNA, mdm-miR171i, specifically targeted and degraded SCARECROW-LIKE PROTEINS26 1 (MsSCL26 1) transcripts. Both knockout of mdm-miR171i and overexpression of MsSCL26 1 improved drought stress tolerance in the cultivated apple line 'GL-3' by regulating the expression of antioxidant enzyme genes, especially that of MONODEHYDROASCORBATE REDUCTASE, which functions in metabolism under drought stress. Transient expression analysis demonstrated that MsSCL26.1 activates MsMDHAR transcription by positively regulating the activity of the P1 region in its promoter. Therefore, the miR171i-SCL26 1 module enhances drought stress tolerance in apple by regulating antioxidant gene expression and ascorbic acid metabolism.

Genome-Wide Identification of the B-BOX Genes that Respond to Multiple Ripening Related Signals in Sweet Cherry Fruit.

B-BOX proteins are zinc finger transcription factors that play important roles in plant growth, development, and abiotic stress responses. In this study, we identified 15 PavBBX genes in the genome database of sweet cherry. We systematically analyzed the gene structures, clustering characteristics, and expression patterns of these genes during fruit development and in response to light and various hormones. The PavBBX genes were divided into five subgroups. The promoter regions of the PavBBX genes contain cis-acting elements related to plant development, hormones, and stress. qRT-PCR revealed five upregulated and eight downregulated PavBBX genes during fruit development. In addition, PavBBX6, PavBBX9, and PavBBX11 were upregulated in response to light induction. We also found that ABA, BR, and GA3 contents significantly increased in response to light induction. Furthermore, the expression of several PavBBX genes was highly correlated with the expression of anthocyanin biosynthesis genes, light-responsive genes, and genes that function in multiple hormone signaling pathways. Some PavBBX genes were strongly induced by ABA, GA, and BR treatment. Notably, PavBBX6 and PavBBX9 responded to all three hormones. Taken together, BBX proteins likely play major roles in regulating anthocyanin biosynthesis in sweet cherry fruit by integrating light, ABA, GA, and BR signaling pathways.

Expression of the Malus sieversii NF-YB21 Encoded Gene Confers Tolerance to Osmotic Stresses in Arabidopsis thaliana.

Drought is the main environmental factor that limits the yield and quality of apples (Malus × domestica) grown in arid and semi-arid regions. Nuclear factor Ys (NF-Ys) are important transcription factors involved in the regulation of plant growth, development, and various stress responses. However, the function of NF-Y genes is poorly understood in apples. Here, we identified 43 NF-Y genes in the genome of apples and conducted an initial functional characterization of the apple NF-Y. Expression analysis of NF-Y members in M. sieversii revealed that a large number of NF-Ys were highly expressed in the roots compared with the leaves, and a large proportion of NF-Y genes responded to drought treatment. Furthermore, heterologous expression of MsNF-YB21, which was significantly upregulated by drought, led to a longer root length and, thus, conferred improved osmotic and salt tolerance in Arabidopsis. Moreover, the physiological analysis of MsNF-YB21 overexpression revealed enhanced antioxidant systems, including antioxidant enzymes and compatible solutes. In addition, genes encoding catalase (AtCAT2, AtCAT3), superoxide dismutase (AtFSD1, AtFSD3, AtCSD1), and peroxidase (AtPER12, AtPER42, AtPER47, AtPER51) showed upregulated expression in the MsNF-YB21 overexpression lines. These results for the MsNF-Y gene family provide useful information for future studies on NF-Ys in apples, and the functional analysis of MsNF-YB21 supports it as a potential target in the improvement of apple drought tolerance via biotechnological strategies.

Genome-Wide Identification of the Xyloglucan endotransglucosylase/Hydrolase (XTH) and Polygalacturonase (PG) Genes and Characterization of Their Role in Fruit Softening of Sweet Cherry.

Fruit firmness is an important economical trait in sweet cherry (Prunus avium L.) where the change of this trait is related to cell wall degradation. Xyloglucan endotransglycosylase/hydrolase (XTH) and polygalacturonases (PGs) are critical cell-wall-modifying enzymes that occupy a crucial position in fruit ripening and softening. Herein, we identified 18 XTHs and 45 PGs designated PavXTH1-18 and PavPG1-45 based on their locations in the genome of sweet cherry. We provided a systematical overview of PavXTHs and PavPGs, including phylogenetic relationships, conserved motifs, and expression profiling of these genes. The results showed that PavXTH14, PavXTH15 and PavPG38 were most likely to participated in fruit softening owing to the substantial increment in expression during fruit development and ripening. Furthermore, the phytohormone ABA, MeJA, and ethephon significantly elevated the expression of PavPG38 and PavXTH15, and thus promoted fruit softening. Importantly, transient expression PavXTH14, PavXTH15 and PavPG38 in cherry fruits significantly reduced the fruit firmness, and the content of various cell wall components including hemicellulose and pectin significantly changed correspondingly in the transgenic fruit. Taken together, these results present an extensive analysis of XTHs and PGs in sweet cherry and provide potential targets for breeding softening-resistant sweet cherry cultivars via manipulating cell wall-associated genes.

Eye Movement Evidence of Attentional Bias for Substance-Related Cues in Heroin Dependents on Methadone Maintenance Therapy.

BACKGROUND: Attentional biases toward substance-related stimuli might play a contributing role in addictive behaviors. OBJECTIVES: This study investigated the selective attention to substance-related stimuli in heroin dependents receiving methadone maintenance therapy. METHODS: Thirty outpatients receiving methadone maintenance treatment for heroin dependence and 38 healthy controls completed a visual probe task with concurrent eye movement monitoring. RESULTS: The results showed that the heroin group reacted faster to probes associated with substance-related pictures than neutral pictures, and they directed more initial fixations and maintained longer initial fixation durations toward substance-related pictures than neutral pictures. However, attentional bias was not correlated with addiction severity in the heroin group. CONCLUSION: These findings suggest that attentional bias towards substance-related cues occurs in heroin dependents, although this bias might not be associated with the severity of drug-using behavior.

Microgels sense wounds' temperature, pH and glucose.

Wound healing concerns almost all bed-side related diseases. With our increasing comprehension of healing nature, the physical and chemical natures behind the wound microenvironment have been decoupled. Wound care demands timely screening and prompt diagnosis of wound complications such as infection and inflammation. Biosensor by the way of exhaustive collection, delivery, and analysis of data, becomes indispensable to arrive at an ideal healing upshot and controlling complications by capturing in-situ wound status. Electrochemical based sensors carry some potential unstable performance subjected to the electrical circuitry and power access and contamination. The colorimetric sensors are free from those concerns. We report that microsensors designed from O/W/O of capillary fluids can continuously monitor wound temperature, pH and glucose concentration. We combined three different types of microgels to encapsulate liquid crystals of cholesterol, nontoxic fuel litmus and two glucose-sensitizing enzymes. A smartphone applet was then developed to convert wound healing images to RGB of digitalizing data. The microgel dressing effectively demonstrates the local temperature change, pH and glucose levels of the wound in high resolution where a microgel is a 'pixel'. They are highly responsive, reversible and accurate. Monitoring multiple physicochemical and physiological indicators provides tremendous potential with insight into healing processing.

Understanding cellulose pyrolysis via ab initio deep learning potential field.

Comprehensive and dynamic studies of cellulose pyrolysis reaction mechanisms are crucial in designing experiments and processes with enhanced safety, efficiency, and sustainability. The details of the pyrolysis mechanism are not readily available from experiments but can be better described via molecular dynamics (MD) simulations. However, the large size of cellulose molecules challenges accurate ab initio MD simulations, while existing reactive force field parameters lack precision. In this work, precise ab initio deep learning potentials field (DPLF) are developed and applied in MD simulations to facilitate the study of cellulose pyrolysis mechanisms. The formation mechanism and production rate of both valuable and greenhouse products from cellulose at temperatures larger than 1073 K are comprehensively described. This study underscores the critical role of advanced simulation techniques, particularly DLPF, in achieving efficient and accurate understanding of cellulose pyrolysis mechanisms, thus promoting wider industrial applications.

Extracellular Matrices as Bioactive Materials for In Situ Tissue Regeneration.

Bioactive materials based on a nature-derived extracellular matrix (NECM) represent a category of biomedical devices with versatile therapeutic applications in the realms of tissue repair and engineering. With advancements in decellularization technique, the inherent bioactive molecules and the innate nano-structural and mechanical properties are preserved in three-dimensional scaffolds mainly composed of collagens. Techniques such as electrospinning, three-dimensional printing, and the intricate fabrication of hydrogels are developed to mimic the physical structures, biosignalling and mechanical cues of ECM. Until now, there has been no approach that can fully account for the multifaceted properties and diverse applications of NECM. In this review, we introduce the main proteins composing NECMs and explicate the importance of them when used as therapeutic devices in tissue repair. Nano-structural features of NECM and their applications regarding tissue repair are summarized. The origins, degradability, and mechanical property of and immune responses to NECM are also introduced. Furthermore, we review their applications, and clinical features thereof, in the repair of acute and chronic wounds, abdominal hernia, breast deformity, etc. Some typical marketed devices based on NECM, their indications, and clinical relevance are summarized.

Synthesis and Functionalization of Graphene Materials for Biomedical Applications: Recent Advances, Challenges, and Perspectives.

Since its discovery in 2004, graphene is increasingly applied in various fields owing to its unique properties. Graphene application in the biomedical domain is promising and intriguing as an emerging 2D material with a high surface area, good mechanical properties, and unrivalled electronic and physical properties. This review summarizes six typical synthesis methods to fabricate pristine graphene (p-G), graphene oxide (GO), and reduced graphene oxide (rGO), followed by characterization techniques to examine the obtained graphene materials. As bare graphene is generally undesirable in vivo and in vitro, functionalization methods to reduce toxicity, increase biocompatibility, and provide more functionalities are demonstrated. Subsequently, in vivo and in vitro behaviors of various bare and functionalized graphene materials are discussed to evaluate the functionalization effects. Reasonable control of dose (<20 mg kg-1 ), sizes (50-1000 nm), and functionalization methods for in vivo application are advantageous. Then, the key biomedical applications based on graphene materials are discussed, coupled with the current challenges and outlooks of this growing field. In a broader sense, this review provides a comprehensive discussion on the synthesis, characterization, functionalization, evaluation, and application of p-G, GO, and rGO in the biomedical field, highlighting their recent advances and potential.

miR164g-MsNAC022 acts as a novel module mediating drought response by transcriptional regulation of reactive oxygen species scavenging systems in apple.

Under drought stress, reactive oxygen species (ROS) overaccumulate as a secondary stress that impairs plant performance and thus severely reduces crop yields. The mitigation of ROS levels under drought stress is therefore crucial for drought tolerance. MicroRNAs (miRNAs) are critical regulators of plant development and stress responses. However, the complex molecular regulatory mechanism by which they function during drought stress, especially in drought-triggered ROS scavenging, is not fully understood. Here, we report a newly identified drought-responsive miRNA, miR164g, in the wild apple species Malus sieversii and elucidate its role in apple drought tolerance. Our results showed that expression of miR164g is significantly inhibited under drought stress and it can specifically cleave transcripts of the transcription factor MsNAC022 in M. sieversii. The heterologous accumulation of miR164g in Arabidopsis thaliana results in enhanced sensitivity to drought stress, while overexpression of MsNAC022 in Arabidopsis and the cultivated apple line 'GL-3' (Malus domestica Borkh.) lead to enhanced tolerance to drought stress by raising the ROS scavenging enzymes activity and related genes expression levels, particularly PEROXIDASE (MsPOD). Furthermore, we showed that expression of MsPOD is activated by MsNAC022 in transient assays. Interestingly, Part1 (P1) region is the key region for the positive regulation of MsPOD promoter by MsNAC022, and the different POD expression patterns in M. sieversii and M. domestica is attributed to the specific fragments inserted in P1 region of M. sieversii. Our findings reveal the function of the miR164g-MsNAC022 module in mediating the drought response of M. sieversii and lay a foundation for breeding drought-tolerant apple cultivars.

Development of high-performance biodegradable rigid polyurethane foams using all bioresource-based polyols: Lignin and soy oil-derived polyols.

Development of biodegradable polyurethane materials is the most promising in the wider context of the "greening" of industrial chemistry. To tackle this challenge, a novel biodegradable polyurethane foam from all bioresource-based polyols (lignin and soy oil-derived polyols) and polymeric methyldiphenyl diisocyanate (pMDI) have been synthesized via a one-pot and self-rising process. All these foam samples have the internal cellular morphology and microstructure. FTIR result exhibits characteristic peaks of polyurethane, and indicates covalent bonds between soy-based polyurethane and lignin, and the lignin powders can react with pMDI via active -H and -CNO. In addition, hydrogen bonding also plays an important role in forming the 3D structures. These interactions and chemical bonds made the prepared foam samples form the 3D macromolecular structure with improved mechanical, thermal, and biodegradable properties. The reaction process is time-saving and cost-effective as it requires no blowing agent and minimum processing steps, while exploring the potential of using the higher content of nature bioresource constituents.

The influence of trait empathy on reactive aggression: An ERP study.

This study mainly investigated the electrophysiological mechanism underlying the effect of trait empathy on reactive aggression using event-related potentials (ERPs). Firstly, we computed the correlation between trait empathy and trait aggression in 413 male undergraduates. The trait empathy was positively correlated with trait aggression, whereas the perspective taking (cognitive empathy) and the empathic concern (affective empathy) dimensions of trait empathy were negatively correlated with physical aggression. Subsequently, we selected 50 young male undergraduates respectively with high empathy (25) and low empathy (25) to complete a competitive reaction time task (a modified version of the Taylor Aggression Paradigm) against two mock opponents in high and low provocation conditions. The high empathy group selected lower average punishments against their opponents than the low empathy group. In the decision phase, high provocation elicited an enlarged ERP negativity compared to low provocation in the high empathy group. This observation was absent in the low empathy group. In the outcome phase, a feedback related negativity was observed in both groups. The low empathy group, but not the high empathy group, showed a significant average amplitude difference for losing and winning trials. The lack of a difference in ERP negativity amplitude in high empathy group suggests that punishing and being punished by opponents have a similar negative valence for this group.

[Experimental study on treatment of immune ulcerative colitis in mice by Changlu Enema].

OBJECTIVE: To observe the therapeutic effect of Chinese compound, Changlu Enema (CE), on immune ulcerative colitis (UC) in mice. METHODS: Mice were randomly divided into the normal group, the model group, the CE high dose (CE-H) and low dose (CE-L) group and the salicylazosulfapyridine (SASP) group. All mice, except those in the normal group, were made into UC model by colonic mucosa protein immunization. After 21 days of medication, the changes of UC activity index and body weight in mice were observed, and the condition of colonic inflammation and histomorphological changes in colonic tissue were observed also. RESULTS: UC activity index was lower and body weight was higher in the two CE groups than those in the model group respectively, showing significant difference (P < 0.05 or P < 0.01); pathological examination showed the pathological changes and inflammatory response in colonic tissue were relieved significantly after treatment, and the improvement in the CE-H group was better than that in the SASP group. CONCLUSION: TCM compound CE is markedly effective in treating UC rats.