Soil salinization severely restricts the sustainable development of the pear industry. Pyrus betulifolia, a vital native salt-tolerant rootstock in China, holds great significance for investigating stress resistance mechanisms. Plant-specific DNA-binding One Zinc Finger (Dof) transcription factors act as pivotal regulators in stress adaptation. However, their functions in P. betulifolia remain largely unexplored. In this study, we identified 43 PbeDof members within the P. betulifolia genome and classified them into eight subfamilies via phylogenetic analysis. Gene structure and conserved motif analyses revealed that PbeDof members within the same subfamily share similar exon-intron organizations and protein architecture, suggesting evolutionary conservation. Promoter analysis indicated that PbeDof genes are rich in cis-acting elements related to light, phytohormones (especially ABA and MeJA), and stress responses, implying their potential roles in diverse biological processes. Chromosomal localization and collinearity analyses revealed that segmental duplication was the primary driver of this family’s expansion. Combined transcriptomic profiling and qRT-PCR assays demonstrated that PbeDof9.1 is predominantly expressed in roots and is strongly induced by salt stress. Subcellular localization confirmed that PbeDof9.1 targets the nucleus. Functional characterization indicated that heterologous overexpression of PbeDof9.1 in Arabidopsis thaliana significantly enhances salt tolerance at germination and seedling stages. Notably, under 175 mM NaCl stress, the transgenic lines exhibited a superior root system architecture, with primary root length and lateral root numbers being approximately 1.5-fold higher than those of the wild type. Furthermore, homologous overexpression in pear calli confirmed that PbeDof9.1 mitigates oxidative damage by boosting the activities of peroxidase (POD) and catalase (CAT) to scavenge reactive oxygen species (ROS), thereby reducing malondialdehyde (MDA) accumulation. Collectively, this study characterizes the PbeDof family and establishes PbeDof9.1 as a key candidate gene for the genetic improvement of salt tolerance in pear rootstocks.

Apoptosis is a highly conserved process that eliminates unwanted or damaged cells in both physiological and pathological conditions. Dysregulation of apoptosis leads to developmental abnormalities and various diseases, such as neurodegeneration and cancer. Drosophila inhibitor of apoptosis 1 (Diap1) plays a crucial role in cell survival by inhibiting caspases and preventing apoptosis. However, under stress conditions, the prodeath proteins Rpr, Hid, and Grim (RHG) induce apoptosis by antagonizing Diap1. Despite being a key component of the apoptotic pathway, the mechanism that controls the stability of Diap1 remains unknown. Here, we find that loss of hdac3 results in the activation of apoptosis, which is completely blocked by expressing Diap1. Although Hdac3 localizes in both the cell cytoplasm and nucleus, only the cytoplasmic Hdac3 is able to suppress apoptosis induced by hdac3 deficiency, RHG overexpression, or x-ray irradiation. This finding indicates that Hdac3 exerts an antiapoptotic role independent of its canonical epigenetic functions. Loss of hdac3 decreases Diap1 protein, which is rescued by introducing cytoplasmic Hdac3. The deacetylase activity is necessary for Hdac3 to suppress apoptosis. Mechanistically, Hdac3 interacts with Diap1 to remove the acetyl group from K315 on Diap1, thereby increasing its stability. Compared with the wild-type Diap1, the acetyl-deficient mutant Diap1-K315R exhibits stronger stability and antiapoptotic activity. Last, RHG proteins compete with Hdac3 for Diap1 interaction, directing Diap1 toward degradation and triggering apoptosis. Together, these findings not only reveal the involvement of Diap1 acetylation modification in apoptosis regulation but also clarify the role of Hdac3 in apoptosis.

Since chronic inflammation is a typical feature of polycystic ovary syndrome (PCOS), both clinical and experimental studies have demonstrated that resveratrol (RES) can effectively alleviate it. However, the underlying mechanism remains unclear. To further investigate this, granulosa cells (GCs) derived from PCOS patients, lipopolysaccharide (LPS)-treated human granulosa cells (KGN), LPS-induced chronic inflammation mouse models, and dehydroepiandrosterone (DHEA)-induced PCOS mouse models were treated with RES. The expression of inflammatory cytokines, including interleukin (IL)-6, IL-1β, chemoattractant protein-1 (MCP-1), and cyclooxygenase-2 (COX2), as well as absent in melanoma 2 (AIM2), was examined. Additionally, ovarian morphological changes in these mouse models were assessed using hematoxylin-eosin (HE) staining. The results showed that the expression of these inflammatory cytokines and AIM2 increased significantly in GCs derived from PCOS patients and LPS-induced KGN cells, as well as in the ovaries of LPS-induced chronic inflammation and DHEA-induced PCOS mouse models. Furthermore, blocking AIM2 in LPS-treated KGN cells and mice with LPS-induced inflammation or PCOS significantly reduced the upregulation of inflammatory cytokines, similar to the results observed following RES treatment. In addition, LPS-induced phosphorylation of the JAK2/STAT3 pathway in KGN cells was completely abolished by RES treatment. Notably, LPS-induced upregulation of AIM2 and these inflammatory cytokines was completely reversed by blocking the JAK2/STAT3 pathway using AZD-1480 and SH-4-54, respectively. Further in vivo studies showed that ovarian morphological and estrous cycle disturbances in DHEA-induced PCOS mouse models were effectively ameliorated by RES and A151. In conclusion, RES alleviates chronic inflammation in PCOS by inhibiting AIM2 via blocking the JAK2/STAT3 pathway. Our findings suggest that the targeted inhibition of AIM2 could represent a novel therapeutic approach for PCOS.

Salmonella enterica can persist in low-moisture foods and shows enhanced dry-heat resistance under low water activity, posing significant food safety challenges. However, the genetic basis of extreme dry-heat resistance and its relationship with other processing stresses remain unclear. In this study, twelve S. enterica strains were screened for dry-heat treatment at 60 °C and 80 °C, with S. Infantis CICC21649 identified as the most resistant strain. Comparative genomics and transcriptional analysis identified candidate genes related to envelope integrity and regulation, including gtrB and dam. Deletion of the chromosomal gtrB homolog reduced dry-heat resistance, producing an additional 0.91-log10 reduction relative to the parent strain at 80 °C. Deletion of dam caused broader stress sensitivity, reducing resistance to both dry heat and high hydrostatic pressure, with the stronger phenotype observed under high hydrostatic pressure. Proteomic analysis of the chromosomal gtrB homolog mutant revealed broad alterations in envelope-associated proteins, transport functions, oxidative stress pathways, and central metabolism under dry-heat stress. These findings indicate that the chromosomal gtrB homolog is an important contributor to extreme dry-heat resistance, whereas dam contributes to resistance against both dry-heat and high hydrostatic pressure, likely through a broader regulatory role in stress adaptation. These results reveal distinct structural and regulatory layers underlying stress adaptation in S. enterica and provide practical guidance for low-moisture food processing by highlighting the need to account for strain-dependent and stress-specific resistance during process validation.

Body weight and body measurements are key indicators of growth and economic efficiency in pigs, but conventional weighing is labor-intensive and stressful, increasing disease risk and necessitating non-contact estimation. We measured five dimensions (body length, chest circumference, abdominal circumference, body width, and body height) in 811 Suzi black pigs and constructed six multiple linear regression models using different combinations. All models had R2 > 0.91, with adjusted R2 also exceeding 0.91, and the model combining length, chest, and abdominal circumference gave the lowest RMSE, balancing accuracy and practicality. Separately, we performed GWAS on 165 genotyped individuals (100 K SNP chip and GBS) for age (as a growth rate proxy), body weight, and the five measurements. No SNP reached genome-wide significance (p < 1.86 × 10-6), but three suggestive loci (p < 1.39 × 10-5) were detected: SNP 4_12 319 200 for age (35.55% variance), a pleiotropic SNP 1_60 912 826 associated with length, chest, and abdominal circumference (42.10%, 53.06%, and 45.97% variance), and SNP 1_60 638 159 for abdominal circumference. Positional mapping identified EPHA7 as the nearest candidate gene. Enrichment analyses revealed focal adhesion, receptor tyrosine kinase, IgSF-CAM, integrin, and PI3K-Akt pathways, with EPHA7 and FYN as key regulators. Notably, the three traits in the best model mapped to the same pleiotropic locus, suggesting a shared genetic basis. This study provides a practical estimation tool and suggestive markers, supporting non-contact weighing systems and molecular breeding.

Ovulatory dysfunction is a typical symptom of polycystic ovary syndrome (PCOS) and is widely recognized as a major obstacle to fertility. Numerous studies have reported a close association between the programmed cell death (PCD) of granulosa cells (GCs) and ovulatory dysfunction in PCOS. Recently, a novel type of PCD exhibiting features of pyroptosis, apoptosis, and necroptosis was identified and termed PANoptosis. However, whether the PANoptosis occurs in PCOS GCs and plays a role in ovulatory dysfunction remains unclear. In this study, we found that the expression levels of PANoptosis-related molecules (NLRP3, CASP1, IL-1β, CASP3, MLKL), the PANoptosis regulator AIM2, IFN-γ, the IFN-responsive transcription factor 1 (IRF1), and HAS2 were significantly increased in GCs collected from PCOS patients, DHT-treated KGN cells, and the ovaries of DHEA-induced PCOS mouse models. Most importantly, KGN cells treated with a combination IFN-γ and lipopolysaccharide (LPS) exhibited similar effects. While, blocking IRF1 (using RNA interference-based knockdown), HAS2 (using 4-MU), JAK2 (using AZD-1480), or STAT3 (using SH-4-54) completely abolished these effects. Mechanistically, we revealed that elevated IFN-γ promotes AIM2 expression and AIM2-PANoptosome assembly through the JAK2/STAT3/IRF1/HAS2 axis. Further in vivo studies showed that inhibiting HAS2 (using 4-MU) and AIM2 (using A151) increased the number of oocytes retrieved from DHEA-induced PCOS mice. Taken together, our results suggest that AIM2-PANoptosis occurs in the GCs of PCOS and is triggered by the IFN-γ/IRF1/HAS2 signalling pathway, thereby resulting in ovulatory dysfunction. This study therefore provides valuable potential therapeutic targets for the treatment of this typical PCOS symptom.

The proliferation and migration of porcine trophectoderm (pTr) cells are crucial processes during the early stages of embryo implantation in sows. The effects of deoxynivalenol (DON) and chlorogenic acid (CGA), a plant-derived compound, on pTr cells are currently unclear. In this study, pTr cells were treated with DON at different times (24, 48, and 72 h) and different concentrations (0.5, 1, and 2 μg/mL) to construct a pathological model of DON-induced pTr cells by detecting the expression levels of genes related to cell proliferation, migration, and oxidative stress, as well as the cell viability and the cell migration ability. Subsequently, CGA intervention experiments revealed that CGA could promote the proliferation, migration, and antioxidant ability of pTr cells and alleviate the damage induced by DON in pTr cells. Finally, RNA-seq technology combined with experiments illustrated that CGA might alleviate the damage of DON-induced pTr cells by regulating the PI3K/AKT signaling pathway. In conclusion, this study explored the toxicological effect of DON and the alleviation effect of CGA on DON at the pTr cells level, which provided new insights and an experimental basis for using CGA to alleviate the reproductive toxicity induced by DON.

Winter rapeseed is susceptible to freezing stress during winter, making it difficult to overwinter safely and resulting in a reduction of yield and quality. DNA methylation, the main epigenetic modification, can regulate plant responses to various stresses. However, the regulatory mechanism of DNA methylation in response to freezing stress in winter rapeseed remains unclear. This study investigates how DNA methylation regulates gene expression and protein abundance in response to freezing stress, revealing key regulatory pathways involved in rapeseed cold tolerance. A total of 6776 unique differentially methylated genes (DMGs), 4285 unique differentially expressed genes (DEGs), and 269 unique differentially abundant proteins (DAPs) were identified between the two cultivars under T1 and T2 freezing stress. Function enrichment analysis revealed that these genes were involved in signal transduction, biosynthesis of unsaturated fatty acids, sugar metabolism, peroxidase, peroxisome, photosynthesis, and additional pathways. An integrative analysis of methylome, transcriptome, and proteome showed that only nine genes were shared among all three datasets, and they were closely related to cold tolerance metabolism in rapeseed. The findings provide molecular insights into rapeseed freezing tolerance, which can be applied in breeding programs to enhance cold resistance in oilseed crops.

African swine fever (ASF), caused by ASF virus (ASFV) infection, is a highly contagious and severe hemorrhagic viral disease with significant economic implications for the global pig farming industry. Currently, there are no effective vaccines or antiviral drugs available for controlling ASF epidemics. Developing effective strategies to combat this epidemic has become imperative. In this study, by screening a chemical library with 400 small molecule compounds, we identified that hycanthone, a previously used anthelminthic drug, exhibits the most potent inhibition on ASFV replication in PAMs, with an IC50 value of 0.57 mu mol/L. Hycanthone showed dose-dependent antiviral activity, reducing the mRNA levels of ASFV p30 and p72 proteins and decreasing viral DNA synthesis. Mechanistically, hycanthone attenuates ASFV replication by suppressing the cellular AKT phosphorylation and inducing viral DNA damage through interacting with ASFV AP endonuclease. Our findings suggest that hycanthone could serve as a promising therapeutic candidate for controlling ASFV infections.

Kynurenine (KYN) is a primary tryptophan derivative found in the human body and fermented foods. Previous studies have shown that KYN is an aryl hydrocarbon receptor (AHR) agonist and is important in regulating various physiological activities, including female reproduction. Progesterone is a vital steroid hormone that facilitates embryo implantation and maintains pregnancy. However, whether KYN affects its biosynthesis remains unclear. To gain understanding, in vitro luteinized porcine granulosa luteal (pGL) cells were treated with KYN. The results showed that KYN disrupted progesterone biosynthesis by decreasing the expression of steroidogenic acute regulatory protein (STAR) and 3beta-hydroxysteroid dehydrogenase (HSD3B) in pGL cells. In addition, the expression of three transcription factors of STAR and HSD3B (GATA4, GATA6, and CEBPB) decreased after KYN treatment. Furthermore, the AHR blockade results showed comparable effects to those of KYN treatment, and subsequent knockdown experiments confirmed these results. These findings suggest that KYN inhibits progesterone biosynthesis in pGL cells by downregulating GATA4, GATA6, and CEBPB expression through AHR. Thus, our results showed for the first time a previously unknown connection between KYN and progesterone biosynthesis.

Numerous studies have demonstrated that lipopolysaccharide (LPS) stimulates TGF-β1 expression. Although studies have implicated the NF-κB/METTL3/METTL14 transactivation/m6A-dependent and AMPK-dependent signaling pathways are engaged in this process in a variety of cell types, the underlying regulatory mechanism in murine macrophages is still not fully understood. To address this issue, in vitro studies were performed using the murine macrophage cell line, RAW264.7. The results showed that LPS challenge resulted in a significant increase in TGF-β1 expression at both mRNA and protein levels. Subsequent studies revealed that the MAPK (including p38, Erk1/2, and JNK) and NF-κB signaling pathways were activated in response to LPS stimulation, but only blocking the Erk1/2 singling pathway completely abolished LPS-induced TGF-β1 expression. Further studies revealed that the levels of a downstream regulator of the Erk1/2 pathway, EGR1, were significantly increased after LPS treatment, and its knockdown significantly reduced LPS-induced Tgf-β1 expression levels. Finally, dual luciferase reporter and ChIP-PCR assays confirmed that EGR1 is a key transcription factor in the regulation of Tgf-β1 expression by binding to its promoter region in response to LPS stimulation. In conclusion, we elucidated the molecular events by which LPS regulates TGF-β1 expression in murine macrophages through the Erk1/2-EGR1 signaling pathway. These findings provide a conceptually novel pathway for LPS-induced TGF-β1 expression beyond the known NF-κB/METTL3/METTL14 transactivation/m6A-dependent and AMPK-dependent signaling pathways.

Abstract Kynurenine (KYN) is a primary tryptophan derivative found in the human body and fermented foods. Previous studies have shown that KYN is an Aryl hydrocarbon receptor (AHR) agonist and is important in regulating various physiological activities, including female reproduction. Progesterone is a vital steroid hormone that facilitates embryo implantation and maintains pregnancy. However, whether KYN affects its biosynthesis remains unclear. To gain understanding, in vitro luteinized porcine granulosa luteal (pGL) cells were treated with KYN. The results showed that KYN disrupted progesterone biosynthesis by decreasing the expression of STAR and HSD3B in pGL cells. In addition, the expression of three transcription factors of STAR and HSD3B (GATA4, GATA6, and CEBPB) decreased after KYN treatment. Furthermore, the AHR blockade results showed comparable to those of KYN treatment, and subsequent knockdown experiments confirmed these results. These findings suggest that KYN inhibits progesterone biosynthesis in pGL cells by downregulating GATA4, GATA6, and CEBPB expression through AHR. Thus, our results showed for the first time, a previously unknown connection between KYN and progesterone biosynthesis.

Backgrounds: Objective of this study is to conduct a genome-wide association study (GWAS) of first-parity reproductive traits in Suzi pigs to identify significant single-nucleotide polymorphisms (SNPs) or candidate genes influencing these traits. Methods: This research employed technologies including the Zhongxin 50K SNP chip, simplified genome sequencing, resequencing, and the 100K SNP liquid chip to perform genome-wide SNP detection on 898 Suzi sows. Genotype data and phenotypic data were combined to do GWAS, gene annotation, and enrichment analysis. Results: Results showed that this study obtained phenotypes of 33 first-parity reproductive traits from 574 sows. GWAS results indicated there were 10 first-parity reproductive traits significantly associated with SNPs, and these traits were AFS, AFF, NNB, NH, NW, NS, NM, ND, PB, and CCN. These 10 traits were significantly associated with 60 SNPs, with 15 (25%) located on chromosome 2-the highest proportion. The SNPs significantly associated with AFS and AFF were largely identical. Genome-wide variance component analysis revealed that among the 10 traits with significantly associated SNPs in GWAS, there were 5 traits that exhibited genome-wide heritability ≥ 0.01. Trait of NM showed the highest heritability (0.65–0.7). These significantly associated SNPs annotated 20 candidate genes, including ADAMTS19, PROP1, ZNF354B, PCARE, LUZP2, VIRMA, EPHA5, AAAS, SLCO3A1-SV2B, KIF18A-BDNF, SERGEF, DYNLRB2, HNF4G, CATSPERD, HSD11B1L, DICER1, RARG, PCDHAC2, KRT79, and HSD17B2. GO analysis of candidate genes revealed that the top three biological processes were cell adhesion, positive regulation of cell projection organization, and positive regulation of neuron projection development. KEGG results showed the top three pathways were inositol phosphate metabolism, glutamatergic synapse, and phosphatidylinositol signaling system. Conclusions: These findings provide a foundation for the reproductive breeding of Suzi pigs and offer new insights into biological breeding in pigs.

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.

BackgroundPolycystic ovary syndrome (PCOS) is a heterogeneous metabolic and endocrine disorder that causes anovulatory infertility and abnormal folliculogenesis in women of reproductive age. Several studies have revealed inflammation in PCOS follicles, and recent evidence suggests that Berberine (BBR) effectively reduces inflammatory responses in PCOS, however, the underlying mechanisms remain unclear.PurposeTo determine the underlying mechanisms by which BBR alleviates inflammation in PCOS.Study designPrimary human GCs from healthy women and women with PCOS, and KGN cells were used for in vitro studies. ICR mice were used for in vivo studies.MethodsGene expression was measured using RT-qPCR. HAS2, inflammatory cytokines, and serum hormones were assayed by ELISA. Protein expression profiles were assayed by Western blot. Chronic low-grade inflammatory mouse models were developed by intraperitoneal injection with LPS, and PCOS mouse models were established by subcutaneous intraperitoneal injection of DHEA. BBR and 4-MU were administered by gavage. Ovarian morphologic changes were evaluated using H&E staining. HAS2 expression in the ovary was assayed using Western blot and immunohistochemistry.ResultsOur results confirmed that HAS2 expression and hyaluronan (HA) accumulation are closely associated with inflammatory responses in PCOS. Data obtained from in vitro studies showed that HAS2 and inflammatory genes (e.g., MCP-1, IL-1β, and IL-6) are significantly upregulated in PCOS samples and LPS-induced KGN cells compared to their control groups. In addition, these effects were reversed by blocking HAS2 expression or HA synthesis using BBR or 4-MU, respectively. Furthermore, HAS2 overexpression induces the expression of inflammatory genes in PCOS. These results were further confirmed in LPS- and DHEA-induced mouse models, where inflammatory genes were reduced by BBR or 4-MU, and ovarian morphology was restored.ConclusionsOur results define previously unknown links between HAS2 and chronic low-grade inflammation in the follicles of women with PCOS. BBR exerts its anti-inflammatory effects by down-regulating HAS2. This study provides a novel therapeutic target for alleviating ovarian inflammation in women with PCOS.

Here, cytosine methylation in the whole genome of pear flower buds was mapped at a single-base resolution. There was 19.4% methylation across all sequenced C sites in the Pyrus pyrifolia cultivar ‘Sucui 1’ flower bud genome. Meantime, the CG, CHG, and CHH sequence contexts (where H = A, T or C) exhibited 47.4%, 33.3%, and 11.9% methylation, respectively. Methylation in different gene regions was revealed through combining methylome and transcriptome analysis, which presented various transcription trends. Genes with methylated promoters exhibited lower expression levels than genes with non-methylated promoters, while body-methylated genes displayed an obvious negative correlation with their transcription levels. The methylation profiles of auxin- and cytokinin-related genes were estimated. And some of them proved to be hypomethylated, with increased transcription levels, in wizened buds. More specifically, the expression of the genes PRXP73, CYP749A22, and CYP82A3 was upregulated as a result of methylation changes in their promoters. Finally, auxin and cytokinin concentrations were higher in wizened flower buds than in normal buds. The exogenous application of paclobutrazol (PP333) in the field influenced the DNA methylation status of some genes and changed their expression level, reducing the proportion of wizened flower buds in a concentration-dependent manner. Overall, our results demonstrated the relationship between DNA methylation and gene expression in wizened flower buds of P. pyrifolia cultivar ‘Sucui 1’, which was associated with changes in auxin and cytokinin concentrations.

The genomic landscape of cold-tolerant winter rapeseed (Brassica napus, L) has been poorly characterized. We assembled a high-quality reference genome of a prominent cold-tolerant winter rapeseed cultivar, NTS57, and performed phylogenetic and pan-genomic analyses by integrating other reported B. napus accessions. The transcriptome analysis revealed that microtubule-associated biological pathways were much more active in NTS57 under cold stress than in the cold-sensitive variety. Whole genome methylation data analysis revealed that DNA demethylation on protein-coding genes and repetitive elements, especially at CHH sites, is essential for cold response in winter rapeseed.

Breeding early maturing cultivars is one of the most important objectives in pear breeding. Very early maturing pears provide an excellent parental material for crossing, but the immature embryo and low seed germination of their hybrid progenies often limit the selection and breeding of new early maturing pear cultivars. In this study, we choose a very early maturing pear cultivar ‘Pearl Pear’ as the study object and investigate the effects of cold stratification, the culture medium, and the seed coat on the germination and growth of early maturing pear seeds. Our results show that cold stratification (4 °C) treatment could significantly improve the germination rates of early maturing pear seeds. A total of 100 days of cold-temperature treatment in 4 °C and in vitro germination on White medium increased the germination rate to 84.54%. We also observed that seed coat removal improved the germination of early maturing pear seeds, with middle seed coat removal representing the optimal method, with a high germination rate and low contamination. The results of our study led to the establishment of an improved protocol for the germination of early maturing pear, which will greatly facilitate the breeding of new very early maturing pear cultivars.

【Objective】The purpose of this study was to explore the genetic polymorphism of swine leukocyte antigen-1（SLA-1）and analyze its disease resistance potential,in order to provide a theoretical basis for molecular breeding of Suzi pig．【Method】The SLA-1gene was amplified by PCR using blood cell cDNA in Suzi pig as a template and sequenced.Similarity analysis and phylogenetic tree construction were conducted using DNAStar and Mega 5.0 software,respectively.NetMHCpan 4.1server was used to analyze the ability and characteristics of SLA-1molecule binding to African swine fever virus（ASFV）antigenic epitopes.The physical and chemical properties,hydrophilic and hydrophobic properties,transmembrane region,structure and function of the encoded proteins were predicted by biological software．【Result】Six SLA-1allele sequences of Suzi pig were successfully amplified,and namely SLA-1*sz01,SLA-1*sz02,SLA-1*sz03,SLA-1*sz04,SLA-1*sz05and SLA-1*sz06.The highly variable sites were mainly located in theα1andα2regions of the peptide binding groove（PBG）.The similarity comparison results showed that the similarity of the six SLA-1alleles nucleotide and amino acid sequences of Suzi pig were between 92.9% to 96.9% and 87.0% to 94.8%,respectively.The similarity of the six SLA-1alleles nucleotide and amino acid sequences of Suzi pig with different pig breeds were between92.8%to 98.8%and 84.3%to 98.0%,respectively.The prediction results of the binding ability of ASFV derived polypeptides showed that the SLA-1*sz01protein had a strong ASFV antigen presentation ability,and had highly affinity with B354L protein polypeptide fragment FIDKTTVLY in ASFV.Prediction of functional domain and tertiary structure showed that SLA-1*sz01was related to the immune system and had a classic SLA-1tertiary structure．【Conclusion】In this study,six SLA-1gene sequences were obtained,among which SLA-1*sz01could restrict the binding of more ASFV-derived peptides,and had the highest affinity with FIDKTTVLY,which could be used as a candidate gene for disease resistance breeding of Suzi pig.The results provided a reference basis for the research and development of ASFV vaccine.

【Objective】Here,two ecotypes of P.betulaefolia from Huaguo Mountain,Lianyungang （the salt-tolerant ecotype,D） and Purple Mountain,Nanjing （the common ecotype,U） were collected for this research.The purpose of this study was to analyze the role of transcription factor genes in the roots of two ecotypes of P.betulaefolia differing in terms of salt stress.Transcription factors involving in the regulation of the salt tolerance of different P.betulaefolia ecotypes were identified on the grounds of differential expression under salt stress and the relationship between the methylation status and the relative expression level of relevant tolerance genes after exposure to salt stress was investigated.【Method】The 90-day-old P.betulaefolia seedlings were grown hydroponically in Hoagland’s nutrient solution supplemented with 200 mmol·L -1 NaCl,with seedlings grown in Hoagland’s nutrient solution as the control.The sodium ion content in the tissues was determined by flame graphite furnace atomic absorption spectrometry.Whole-genome DNA methylation analysis and transcriptome sequencing were performed on three replicates for the following four root samples:ecotype D and ecotype U,each grown in the presence or absence of salt stress.Bioinformatics analysis of transcription factor gene expression under salt stress at the levels of transcriptional regulation and epigenetic methylation were carried out using transcriptome sequencing data and whole-genome DNA methylation results,respectively.Then,McrBC-PCR and real-time fluorescence quantitative PCR （qPCR） were used to confirm the levels of methylation and transcription of differential transcription factor genes.【Result】After exogenous NaCl treatment for 24 h,the concentration of sodium ions in P.betulaefolia roots increased significantly,with the increase in sodium ion concentration in the salt-tolerant ecotype being significantly less than that in the common ecotype.In the whole seedling,the final salt concentration of tolerant ecotype was only 73.1%of that of the common ecotype.Whereas,in the roots,the sodium content of the salt-tolerant ecotype was 1.1 times of that in the common ecotype.These results indicated that the salt-tolerant ecotype could store more sodium ions in roots and limit their upward transport after salt stress.A total of 2 682 transcription factor （TF） genes from 69 gene families were detected in roots.Among them,243 TF genes displayed differential expression in response to salt stress,including 37 AP2/ERF,19 bHLH,7 bZIP,10 HD-Zip,30 MYB,18 NAC,8 WRKY,and 23 ZFP family genes.The global methylation level of transcription factor genes in the genome of the salt-tolerant rootstock ecotype decreased,whereas the overall methylation level of these genes in the common ecotype increased after exposure to 200 mmol·L -1 NaCl.The differentially methylated regions in both ecotypes were mainly in the position of gene promoters,with the type of differentially methylated sequences being mostly mCHH,constituting more than 93%of the sum of all three types of methylated sequences.The expression levels of twenty-three transcription factor genes,which belonged to the AP2/ERF,bHLH,DREB,GRAS,GT factor,HB Zip,MYB,NAC,Trihelix,and zinc-finger ZFP gene families,were upregulated,and their methylation levels were downregulated in both two ecotypes in response to salt stress.These genes may be involved in the regulation of sodium uptake and accumulation in roots under salt stress.The expression patterns and promoter methylation of representative candidate genes identified by bioinformatics analysis were confirmed by qPCR and McrBC-qPCR.【Conclusion】The differentially expressed genes in roots of P.betulaefolia under salt stress included 243 transcription factor genes in both ecotypes.The methylation changes in DNA sequences in eight transcription factor genes （PbERF2,PbGT3,PbZAT10.1,PbSCL33,PbDREB1,PbZAT10.2,PbERF53,and PbNAC72） were correlated with their transcriptional activity.Our results provided preliminary experimental evidence for supporting a relationship between promoter DNA methylation and expression of TF genes in P.betulaefolia in response to salt stress as part of the molecular role of TFs involved in the regulation of salt tolerance among different P.betulaefolia ecotypes,which would increase our understanding of the role of epigenetics in the response of woody trees to abiotic stress.