IntroductionMarek’s disease, caused by the oncogenic Marek’s disease virus (MDV), remains a major threat to poultry production. Host immune responses and apoptosis pathways play critical roles in MDV pathogenesis, yet the underlying mechanisms are not fully understood.MethodsChicken embryo fibroblasts (CEF) were treated with poly I:C to assess its effect on MDV infection, followed by transcriptome profiling to identify differentially expressed genes. Functional assays using siRNA knockdown and overexpression of TNFRSF11B were performed to evaluate its impact on viral infection and apoptosis.ResultsPoly I:C treatment significantly reduced MDV infection in CEF. Transcriptome analysis identified 274 DEGs, among which TNFRSF11B was notably downregulated. Consistently, MDV infection suppressed TNFRSF11B expression in both CEF and spleen tissues. Functional assays revealed that knockdown of TNFRSF11B enhanced MDV infection, whereas its overexpression suppressed MDV infection. Furthermore, MDV infection induced substantial cell apoptosis, and TNFRSF11B knockdown further exacerbated this effect, as demonstrated by elevated CASP3 expression and higher apoptosis rates. A strong positive correlation was observed between MDV infection levels and apoptosis rates (R2 = 0.9895, p < 0.0001).DiscussionOur findings indicated TNFRSF11B could modulate MDV infection through collaborating with MDV to modulate cell apoptosis. This study provides new insights into the pathogenesis of MDV and potential antiviral strategies targeting MDV/TNFRSF11b—cell apoptosis.

IntroductionThe blood and meat spots in eggs are recognized as defects for egg quality. The frequency of blood and meat spots in brown-shell eggs is much higher than that in white-shell eggs in previous studies. However, the actual occurrence frequency and their effects on the microbial composition in eggs remain poorly understood.MethodsIn this study, we examined the frequency of blood and meat spots in brown-shell and white-shell eggs, respectively, from Rhode Island Red and White Leghorn chickens at seven ages.ResultsThe results showed that blood and meat spots in brown-shell eggs exhibit much higher average frequency (63.99%) than that in white-shell eggs (1.37%). Furthermore, we analyzed the relationship between the presence of blood and meat spots and the microbial community distribution in the egg albumen and yolk. Briefly, we selected brown-shell eggs (n = 112) from Rhode Island Red, among which 51 eggs showing blood/meat spots were classified as RIR_CASE, and 61 normal eggs without blood/meat spot were classified as RIR_CON. Additional white-eggshell eggs (n = 124) without blood/meat spots from White Leghorn were selected as WL_CON. 16S rRNA sequencing was performed in both egg white and yolk. The results indicated that neither egg white nor yolk is sterile, with Proteobacteria identified as the dominant bacterial phyla. The microbial alpha diversity in both egg white and yolk of RIR_CASE was significantly lower compared to RIR_CON and WL_CON. Beta diversity analysis showed that the Weighted UniFrac Distance between RIR_CASE and RIR_CON in the egg yolk group was significantly larger than the distance between WL_CON and RIR_CON. It suggested that the difference of microbial diversity was mainly caused by blood and meat spots other than by chicken breeds. LEfSe analysis identified eight microbial taxa closely linked to the presence of blood and meat spots in egg white or yolk. Moreover, through the combination of random forest analysis, we identified the unique microbial biomarkers Comamonas_F and Chryseobacterium in the egg white of the RIR_CASE group.DiscussionOur study indicates that eggs with blood and meat spots occur at a higher frequency in brown-shell chickens and are accompanied by a distinct microbial community distribution.

Blood and meat spots in egg white and yolk are regarded as defects affecting egg internal quality. The presence of blood and meat spots in egg directly reflects the health status of the hen’s ovary and oviduct. In this study, we analyzed mRNA and lncRNA profiles from 36 samples (ovary, magnum/isthmus, and uterus) collected from 12 Rhode Island Red hens with higher and lower incidence of blood and meat spots (BMS) in eggs, to elucidate the factors associated with BMS incidence. There were 398, 359, 548 differentially expressed genes (DEGs) and 194, 34, 170 differentially expressed lncRNAs (DElncRNAs) across the ovary, magnum/isthmus and uterus identified between high and low BMS incidence groups. Through WGCNA analysis, we identified phenotype-associated gene modules in the ovary, the key tissues where egg yolk formed, which indicated DEGs in the ovary was associated with BMS formation. Further analysis revealed 36, 28, 353 DEGs were co-expressed with DElncRNAs in the ovary, magnum/isthmus tissues, and uterus, respectively. Functional enrichment analysis indicated that co-expressed genes in ovary and magnum/isthmus were predominantly associated with inflammatory responses and immune regulation, which indicated inflammation response occurred in ovary tissue. Our study provided the transcriptome profiles of chickens with high and low BMS, which was helpful for in-depth research on molecular mechanisms underlying BMS formation.

Marek's disease (MD), caused by the oncogenic Marek's disease virus (MDV), is a lymphoproliferative disease in chickens and serves as a natural animal model for CD30-overexpressing lymphomas. The expression of CD30, a member of the tumor necrosis factor receptor superfamily, is significantly upregulated during lymphocyte transformation in MD. The MDV-encoded proto-oncogene Meq is known to bind to the chicken CD30 promoter, but the precise regulatory mechanisms and functional binding sites remain uncharacterized. This study aimed to elucidate the transcriptional regulation of CD30 by Meq. Initially, expression of CD30 was detected in spleens of MDV-uninfected chickens and tumorous spleens of MDV-infected chickens, and the results showed it was highly expressed in tumorous spleens of MDV-infected chickens. Knockdown of Meq in vitro resulted in a corresponding downregulation of CD30 expression, confirming a positive regulatory relationship between them. Subsequently, dual-luciferase reporter assays and chromatin immunoprecipitation (ChIP)-qPCR demonstrated that Meq directly bound to the CD30 promoter to activate its transcription, and the primary binding regions located at -1383 to -1162 bp and -97 to +75 bp relative to the transcription start site. Collectively, this study demonstrated Meq could bind to CD30 promoter at two binding regions and up-regulate its expression, and provided a critical foundation to investigate the role of the CD30 signaling pathway in MD-induced tumorigenesis.

Myocyte enhancer factor 2A (MEF2A) is a transcription factor that plays a critical role in cell proliferation, differentiation and apoptosis. In contrast to the wide characterization of its regulation mechanism in mammalian skeletal muscle, its role in chickens is limited. Especially, its wide target genes remain to be identified. Therefore, we utilized Cleavage Under Targets and Tagmentation (CUT&Tag) technology to reveal the genome-wide binding profile of MEF2A in chicken primary myoblasts thus gaining insights into its potential role in muscle development. Our results revealed that MEF2A binding sites were primarily distributed in intergenic and intronic regions. Within the promoter region, although only 8.87% of MEF2A binding sites were found, these binding sites were concentrated around the transcription start site (TSS). Following peak annotation, a total of 1903 genes were identified as potential targets of MEF2A. Gene Ontology (GO) enrichment analysis further revealed that MEF2A target genes may be involved in the regulation of embryonic development in multiple organ systems, including muscle development, gland development, and visual system development. Moreover, a comparison of the MEF2A target genes identified in chicken primary myoblasts with those in mouse C2C12 cells revealed 388 target genes are conserved across species, 1515 target genes are chicken specific. Among these conserved genes, ankyrin repeat and SOCS box containing 5 (ASB5), transmembrane protein 182 (TMEM182), myomesin 2 (MYOM2), leucyl and cystinyl aminopeptidase (LNPEP), actinin alpha 2 (ACTN2), sorbin and SH3 domain containing 1 (SORBS1), ankyrin 3 (ANK3), sarcoglycan delta (SGCD), and ORAI calcium release-activated calcium modulator 1 (ORAI1) exhibited consistent expression patterns with MEF2A during embryonic muscle development. Finally, TMEM182, as an important negative regulator of muscle development, has been validated to be regulated by MEF2A by dual-luciferase and quantitative real-time PCR (qPCR) assays. In summary, our study for the first time provides a wide landscape of MEF2A target genes in chicken primary myoblasts, which supports the active role of MEF2A in chicken muscle development.

Marek’s disease (MD), an immunosuppressive disease induced by the Marek’s disease virus (MDV), is regarded as an ideal model for lymphoma research to elucidate oncogenic and anti-oncogene genes. Using this model, we found that circRUNX2.2, derived from exon 6 of RUNX2, was significantly upregulated in MDV-infected tumorous spleens. In this study, we deeply analyzed the potential role of circRUNX2.2 in lymphoma cells. An open reading frame (ORF) in circRUNX2.2 with no stop codon was predicted, and small peptides (named circRUNX2.2-rt) presenting multiple ladder-like bands with different molecular weights encoded by circRUNX2.2 were detected via Western blotting assay. The polysome fraction assay reconfirmed the translation ability of circRUNX2.2, which could be detected in polysome fractions. Subsequent analysis verified that it translated in a rolling circle manner, rather than being assisted by the internal ribosome entry site (IRES) or m6A-mediated mechanism. Furthermore, we found that circRUNX2.2-rt was potently induced in MSB1 cells treated with sodium butyrate (NaB), which reactivated MDV and forced the MDV transition from the latent to reactivation phase. During this phase, MDV particles were clearly observed by electron microscopy, and the viral gene pp38 was also significantly upregulated. A biological function study showed that circRUNX2.2-rt promoted cell proliferation and cell cycle transition from the S to G2 phase and inhibited the apoptosis of MSB1. Further immunoprecipitation and mass spectrometry assays showed that 168 proteins potentially interacting with circRUNX2.2-rt were involved in multiple pathways related to cell cycle regulation, which proved that circRUNX2.2-rt could bind or recruit proteins to mediate the cell cycle.

Marek's disease (MD), an immunosuppression disease induced by Marek's disease virus (MDV), is one of the significant diseases affecting the health and productive performance of poultry. The roles of circular RNAs (circRNAs) in MD development were poorly understood. In this study, we found a circRNA derived from exon 6 of RUNX family transcription factor 2 (RUNX2) gene, named circRUNX2.2, was highly expressed in chicken tumorous spleens (TS) induced by MDV. Through fluorescence in situ hybridization and nuclear-cytoplasmic separation assay, we determined circRUNX2.2 was mainly located in the nucleus. Knockout experiments confirmed that the flanking complementary sequences (RCMs) mediated its circularization. Gain of function assay and dual luciferase reporter gene assay revealed that circRUNX2.2 could promote the expression of RUNX2 via binding with its promoter region. RNA antisense purification assay and mass spectrometry assay showed circRUNX2.2 could recruit proteins such as CHD9 protein. Knocking down CHD9 expression decreased the expression of RUNX2 gene, which confirmed the positive regulation that circRUNX2.2 on RUNX2 expression was probably facilitated via recruiting CHD9 protein. Functional experiments showed that circRUNX2.2 promoted the proliferation of the MD lymphoma-derived chicken cell line, MDCC-MSB1, which confirmed the potential oncogenic role of circRNX2.2 in tumor development. In conclusion, we found that the RUNX2-derived circRUNX2.2 can positively regulate the transcription of the parental gene RUNX2 in a cis-acting manner. The high expression of circRUNX2.2 in MD tumor tissues indicated that it might mediate MD lymphoma progression.

Background Skeletal muscle development is pivotal for animal growth and health. Recently, long noncoding RNAs (lncRNAs) were found to interact with chromatin through diverse roles. However, little is known about how lncRNAs act as chromatin-associated RNAs to regulate skeletal muscle development. Here, we aim to investigate the regulation of chromatin-associated RNA (MYH1G-AS) during skeletal muscle development. Methods We provided comprehensive insight into the RNA profile and chromatin accessibility of different myofibers, combining RNA sequencing (RNA-seq) with an assay for transposase-accessible chromatin with high-throughput sequencing (ATAC-seq). The dual-luciferase reporter assay and chromatin immunoprecipitation (ChIP) assay were used to analyze the transcriptional regulation mechanism of MYH1G-AS. ALKBH5 -mediated MYH1G-AS N 6 -methyladenosine (m 6 A) demethylation was assessed by a single-base elongation and ligation-based qPCR amplification method (SELECT) assay. Functions of MYH1G-AS were investigated through a primary myoblast and lentivirus/cholesterol-modified antisense oligonucleotide (ASO)-mediated animal model. To validate the interaction of MYH1G-AS with fibroblast growth factor 18 (FGF18) protein, RNA pull down and an RNA immunoprecipitation (RIP) assay were performed. Specifically, the interaction between FGF18 and SWI/SNF-related matrix-associated actin-dependent regulator of chromatin subfamily A member 5 (SMARCA5) protein was analyzed by coimmunoprecipitation (Co-IP) and a yeast two-hybrid assay. Results A total of 45 differentially expressed (DE) lncRNAs, with DE ATAC-seq peaks in their promoter region, were classified as open chromatin-associated lncRNAs. A skeletal muscle-specific lncRNA (MSTRG.15576.9; MYH1G-AS), which is one of the open chromatin-associated lncRNA, was identified. MYH1G-AS transcription is coordinately regulated by transcription factors (TF) SMAD3 and SP2. Moreover, SP2 represses ALKBH5 transcription to weaken ALKBH5 -mediated m 6 A demethylation of MYH1G-AS, thus destroying MYH1G-AS RNA stability. MYH1G-AS accelerates myoblast proliferation but restrains myoblast differentiation. Moreover, MYH1G-AS drives a switch from slow-twitch to fast-twitch fibers and causes muscle atrophy. Mechanistically, MYH1G-AS inhibits FGF18 protein stabilization to reduce the interaction of FGF18 to SMARCA5, thus repressing chromatin accessibility of the SMAD4 promoter to activate the SMAD4 -dependent pathway. Conclusions Our results reveal a new pattern of the regulation of lncRNA expression at diverse levels and help expound the regulation of m 6 A methylation on chromatin status. Graphical Abstract

1. Skeletal muscle is an important component of chicken carcass. In chickens, the number of muscle fibres is fixed during the embryonic period, and muscle development during the embryonic period determines the muscle development potential after hatching.2. Beijing-You (BY) and Cornish (CN) chickens show completely different growth rates and body types, and two breeds were used in this study to explore the role of lncRNAs in muscle development during different chicken embryonic periods. A systematic analysis of lncRNAs and mRNAs were conducted in the pectoral muscle tissues of BY and CN chickens at embryonic days 11 (ED11), 13 (ED13), 15 (ED15), 17 (ED17), and 1-day-old (D1) using RNA-seq. A total of 4,104 differentially expressed transcripts (DETs) were identified among the five stages, including 2,359 lncRNAs and 1,745 mRNAs.3. The number of DETs between the two breeds at ED17 (1,658 lncRNAs and 1,016 mRNAs) was much higher than the total number of DET at all the other stages (692 lncRNAs and 729 mRNAs), indicating that the two breeds show the largest difference in gene regulation at ED17.4. Correlation analysis was performed for all differentially expressed lncRNAs and mRNAs during the five periods. Forty-three, cis interaction pairs of lncRNA-mRNA related to chicken muscle development were predicted. The expression of four pairs was verified, and the results showed MSTRG.12395.2-FGFBP2 and MSTRG.18590.6-FMOD were significantly up-regulated in CN at ED11 compared to BY and might be important candidate genes for embryonic muscle development.

为了探究生物信息学软件识别禽白血病病毒E(Avian leukosis virus subgroup E,ALVE)基因插入位点的准确性,试验采用obsERVer流程和位点特异性PCR方法,测定了 3只BWEL-SPF鸡全基因组测序数据中的ALVE基因插入位点,并验证了 obsERVer流程预测的准确性.结果表明:在3只BWEL-SPF鸡中共识别到4个ALVE基因插入位点,分别为ALVE1、ALVE3、ALVE9和ALVE 15;插入位点上下游序列存在规律性,是由6个碱基的基序构成的重复序列;并且这4个插入位点均可以通过位点特异性PCR方法验证,吻合率为100％.说明obsERVer流程可以准确预测测序数据中的ALVE基因插入位点,在BWEL-SPF鸡中存在多个ALVE基因插入位点.

Marek's disease (MD) is a lymphoproliferative neoplastic disease caused by Marek's disease virus (MDV). Previous studies have showed that DNA methylation was involved in MD development, but systematic studies are still lacking. Herein, we performed whole genome bisulfite sequencing (WGBS) and RNA-seq in MDV-infected tumorous spleens (IN), noninfected spleens (NoIN), and survivor (SUR) spleens of chickens to identify the genes playing important roles in MD tumor transformation. We generated the first genome-wide DNA methylation profile of MDV-infected, noninfected, and survivor chickens. Combined the WGBS and RNA-Seq, we found that the expression of 25% differential expression genes (DEGs) were significantly correlated with methylation of CpG sites in their gene bodies or promoters. Further, we focused on the DEGs with differentially methylated regions (DMRs) on genes' body and promoter, and it showed the expression of 60% DEGs were significantly correlated with methylation of CpG sites in DMRs. Finally, we identified 8 genes, including CD4, CTLA4, DTL, HMGB1, LGMN, NUP210, RAD52, and ZAP70, and their expression was negatively correlated with methylation of DMRs in their promoters in both IN vs. NoIN and IN vs. SUR. These 8 genes showed specifically high expression in IN groups and clustered in module turquoise analyzed by WGCNA. Out of 8 genes, CD4 and HMGB1 were drop in QTLs associated with MD resistance. Thus, we overexpressed the 2 genes to simulate their high expression in the IN group and found they significantly promoted MDCC-MSB-1 cell proliferation, which revealed they might play promoting roles in MD tumorigenesis in IN due to their high expression induced by hypomethylation.

Avian leukosis virus subgroup E (ALVE) as a kind of endogenous retroviruses extensively exists in chicken genome. The insertion of ALVE has some effects on chicken production traits and appearance. Most of the work on ALVEs has been done with commercial breeds. We present here an investigation of ALVE elements in seven Chinese domestic breeds and four standard breeds. Firstly, we established an ALVE insertion site dataset by using the obsERVer pipeline to identify ALVEs from whole-genome sequence data of eleven chicken breeds, seven Chinese domestic breeds, including Beijing You (BY), Dongxiang (DX), Luxi Game (LX), Shouguang (SG), Silkie (SK), Tibetan (TB) and Wenchang (WC), four standard breeds, including White Leghorn (WL), White Plymouth Rock (WR), Cornish (CS), and Rhode Island Red (RIR). A total of 37 ALVE insertion sites were identified and 23 of them were novel. Most of these insertion sites were distributed in intergenic regions and introns. We then used locus-specific PCR to validate the insertion sites in an expanded population with 18~60 individuals in each breed. The results showed that all predicted integration sites in 11 breeds were verified by PCR. Some ALVE insertion sites were breeds specific, and 16 out of 23 novel ALVEs were found in only one Chinese domestic chicken breed. We randomly selected three ALVE insertions including ALVE_CAU005, ALVE_ros127, and ALVE_ros276, and obtained their insertion sequences by long-range PCR and Sanger sequencing. The insertion sequences were all 7525 bp, which were full-length ALVE insertion and all of them were highly homologous to ALVE1 with similarity of 99%. Our study identified the distribution of ALVE in 11 chicken breeds, which expands the current research on ALVE in Chinese domestic breeds.

Skeletal muscle is a regulator of the body's energy expenditure and metabolism. Abnormal regulation of skeletal muscle-specific genes leads to various muscle diseases. Long non-coding RNAs (lncRNAs) have been demonstrated to play important roles in muscle growth and muscle atrophy. To explore the potential function of muscle-associated lncRNA, we analyzed our previous RNA-sequencing data and selected the lncRNA (LncEDCH1) as the research object. In this study, we report that LncEDCH1 is specifically enriched in skeletal muscle, and its transcriptional activity is positively regulated by transcription factor SP1. LncEDCH1 regulates myoblast proliferation and differentiation in vitro. In vivo, LncEDCH1 reduces intramuscular fat deposition, activates slow-twitch muscle phenotype, and inhibits muscle atrophy. Mechanistically, LncEDCH1 binds to sarcoplasmic/ER calcium ATPase 2 (SERCA2) protein to enhance SERCA2 protein stability and increase SERCA2 activity. Meanwhile, LncEDCH1 improves mitochondrial efficiency possibly through a SERCA2-mediated activation of the AMPK pathway. Our findings provide a strategy for using LncEDCH1 as an effective regulator for the treatment of muscle atrophy and energy metabolism.

鸡主要组织相容性复合体(MHC)与排斥反应、淋巴细胞反应及抗原递呈有关,其编码基因簇位于16号染色体上,其上许多基因具有多态性,与疾病抗性或易感性密切相关.文章主要对鸡的16号染色体上基因分布、MHC-B区域基因、MHC单倍型检测方法及鸡MHC与常见疾病的关系等方面进行综述.

Background Marek’s disease (MD), a class II infectious, lymphoproliferative disease that mainly afflicts poultry, has been shown to cause wasting, limb paralysis, and often acute death. It is a neoplastic disease caused by a cell-binding herpesvirus that leads to the formation of tumors in various organs and tissues. Our previous reports have found that the microRNA, gga-miR-29b-3p, showed abnormal expression in MD lymphoma. However, it remains unknown whether gga-miR-29b-3p affects MD tumorigenesis. Methods The MD tumor cell line MSB1 was chosen to analyze the characteristics of gga-miR-29b-3p in tumors. Cell proliferation and migration were assessed by Cell Counting Kit-8 (CCK-8) and Transwell, respectively, and cell apoptosis and cycle were analyzed via fluorescent staining and flow cytometry, respectively. The regulation between gga-miR-29b-3p and its potential target genes was verified by dual luciferase results and loss-of-function assays. The effect of target genes was verified by examining the degree of RNA interference on MSB1 cells. Results Analysis revealed that gga-miR-29b-3p impaired the proliferation of the MSB1 MD tumor cell line, induced apoptosis without obvious effects on the cell cycle, and suppressed the expression of the invasion-associated MMP2 and MMP9 genes. It was concluded that DNMT3B is the direct target of gga-miR-29b-3p. As expected, the effects of DNMT3B knockdown with small interfering RNA (siRNA) on MSB1 cell proliferation, apoptosis, and cycle were associated with gga-miR-29b-3p overexpression. Moreover, BCL2 and BCL2L1 were downregulated and TNFSF10 was upregulated in both the gga-miR-29b-3p overexpression and DNMT3B knockdown groups. The expression levels of invasion-related genes were decreased post-DNMT3B knockdown. In both the gga-miR-29b-3p overexpression and DNMT3B knockdown conditions, a decrease in MEQ oncogene expression in MD virus was observed. Conclusions Overall, gga-miR-29b-3p was demonstrated to have a suppressive effect in MD lymphoma progression via the targeting of the DNMT3B gene. Gga-miR-29b-3p overexpression and DNMT3B knockdown inhibited MSB1 cell proliferation through suppressing the pro-apoptotic gene expression and elevating the anti-apoptotic gene expression in the apoptosis pathway. Our study provides a theoretical basis for targeted treatment of MD.

《动物遗传学》是遗传学重要分支之一，是高等农科院校动物科学专业必修课程，也是其他相关专业课程和动物遗传育种领域从业人员的重要理论基础。考虑到高校课程改革的要求、课时安排的调整和新时代“思政教育”的使命，教研组结合多年课程教学经验，对课程内容从多个角度进行优化和设计，旨在将《动物遗传学》打造成一门高效且独具特色的精品课程，为相关领域培养新时代“顶天”“立地”的优秀大学生。

Marek's disease (MD), an immunosuppressive disease induced by Marek's disease virus (MDV), provides an ideal model for studying diseases caused by a carcinogenic virus. CD79B is a B-cell antigen receptor complex-associated protein β-chain precursor which is involved in the activation, proliferation, differentiation of B-cell and the transmission of downstream signals. This study analyzed CD79B gene mRNA expression and methylation by two schemes #20 (5´ flanking to intron 1) and #27 (intron 2 to intron 3), between MDV-infected tumorous spleens (TS) and non-infected spleens (NS). Results showed that average methylation levels of CpGs in #20 and #27 were higher in TS than in NS (P<0.05), while, CD79B mRNA expression was lower in TS than in NS (P<0.01). Six of 40 CpG sites showed significantly (P<0.05) different methylation levels between TS and NS. Correlation analysis showed that the average methylation level rather than a single site methylation level in #20 affected (P<0.05) mRNA expression. Collectively, it was found that the change of CD79B gene expression after MDV infection might be partly explained by modification of DNA methylation.

Marek’s disease (MD), a highly contagious T cell lymphoid neoplasia disease of chickens, causes huge economic losses to the poultry industry. It is the only one tumor disease which can be prevented by vaccine in chickens; therefore, MD is considered to be an excellent model to study the pathogenesis of virus-induced cancer. Recently, abundant evidences have verified that miRNAs are regulators in the process of neoplastic transformation. In our previous study on miRNome analysis of MDV-induced lymphoma in chicken, we found that gga-miR-181a was downregulated drastically in MDV-infected spleens. To further investigate the role of gga-miR-181a in MDV-induced lymphomagenesis, we performed cell migration assay, and the results suggested that gga-miR-181a suppressed the migration of MDV-transformed lymphoid cell (MSB-1). Subsequently, luciferase reporter gene assay revealed that acidic nuclear phosphoprotein 32A (ANP32A) was a functional target gene of gga-miR181a. Real-time PCR and western blot assay showed that the mRNA and protein levels of ANP32A were downregulated in gga-miR-181a mimic group at 48-h and 96-h post-transfection, respectively, indicating that ANP32A was modulated by gga-miR-181a. All the results suggested that gga-miR-181a was an inhibitor in MSB-1 cell migration. ANP32A was a direct target gene of gga-miR-181a and they were implicated in MD lymphoma tumorigenesis.

目的:研究gga-miR-29b-3p对马立克氏病(MD)肿瘤转化细胞侵袭和原癌基因Meq表达的影响.方法:选取SPF白来航鸡在感染马立克氏病病毒(MDV)后引发的内脏淋巴瘤为样本,通过实时荧光定量PCR检测gga-miR-29b-3p的表达情况;利用在线生物软件对gga-miR-29b-3p潜在的靶基因进行功能分析.以该miRNA为研究对象,MDV转化细胞系MDCC-MSB1为试验材料,分别转染miRNA激动剂或阴性对照,检测细胞侵袭相关基因MM P2和MM P9以及MDV原癌基因Meq的表达情况.结果:Gga-miR-29b-3p在感染MDV白来航鸡的肿瘤化脾脏和肝脏淋巴瘤中均显著低表达.信号通路方面miRNA的预测靶基因分别参与FoxO信号通路、mTOR信号通路、Jak-STAT信号通路、Toll样受体信号通路、ErbB信号通路、VEGF信号通路和细胞凋亡等,这些信号通路可参与肿瘤发生进程.转染miRNA激动剂后,MMP2和MMP9基因的表达量在48 h显著下调(P<0.05);Meq基因在48 h表达显著下调(P<0.05).结论:Gga-miR-29b-3p抑制细胞侵袭和病毒原癌基因的表达,其潜在靶基因能够介导肿瘤发生,提示该miRN A可能参与M D肿瘤转化过程.

Marek’s disease (MD) is one of the top priority diseases in chicken. The mechanisms through which immune cells react to virus infection and pathway signals represent the fundamental cell biological question to defeat MD. We hypothesized epigenetic status in immune organs and cells is directly associated with how the Marek’s disease virus (MDV) infection influences intrinsic transcriptional and regulatory networks in MD. To investigate the epigenetics in CD4+ T cells induced by MDV infection, we assembled a multifaceted approach with epigenetics, deep sequencing, and computational methods together to explore the roles of epigenetics in unique inbred lines of chickens. First, a genome-wide transcriptome analysis in the immune organs from resistant line 63 and susceptible line 72 chickens was performed to explore disease resistance mechanisms. MDV infection influences both cytokine-cytokine receptor interaction and cellular development in resistant and susceptible chickens. Second, we examined the epigenetic status of CD4+ T cells induced by MDV infection, including DNA methylations, histone modifications, chromatin accessibility, and 3D chromatin structures. Our results revealed more than 5,000 epigenetic modification changes (FDR< 0.01) and methylation changes (>15,000, FDR< 0.01) caused by MDV infection. Only resistant line 63 chickens could initiate robust adaptive immune responses at the transcription level (>200 genes, FDR< 0.05). The increase in chromatin accessibility (P < 0.001) and chromosome reorganization represented by A/B compartment flipping were related to up-regulated genes induced by MDV infection ten days post-infection in line 63 chickens. Our findings provided a deeper insight into the CD4+ T cell commitment and responses toward viral infection. In particular, the identifications of cis-acting and trans-acting regulations and lipid pathways will improve our understanding of the sequence, structural basis of RNA-protein, and RNA-DNA interactions and serve as the impetus for mechanistic studies to refine the genomic and epigenetic control of MD resistance in poultry.