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DNA甲基化在调节昆虫的生长发育、环境适应和免疫应答等生物学过程中发挥重要作用。但目前东方蜜蜂(Apis cerana)的DNA甲基转移酶(DNA methyltransferase, DNMT)的研究较为滞后，相关信息有限。本研究解析东方蜜蜂的DNA甲基转移酶1a(AcDNMT1a)和DNA甲基转移酶1b(AcDNMT1b)的理化性质和分子特征，并检测AcDnmt1a和AcDnmt1b在工蜂幼虫肠道发育及应答蜜蜂球囊菌(Ascosphaera apis)侵染过程中的表达模式，为深入开展AcDNMT1a和AcDNMT1b的分子功能和表观调控机制研究提供参考和依据。通过生物信息学预测AcDNMT1a和AcDNMT1b的理化性质、分子特征及系统进化，并比较东方蜜蜂与其他昆虫DNMT1a和DNMT1b所含的结构域和保守基序。采用RT-qPCR检测东方蜜蜂工蜂幼虫肠道发育过程及应答蜜蜂球囊菌侵染过程中AcDnmt1a和AcDnmt1b的相对表达量。AcDNMT1a和AcDNMT1b的蛋白质相对分子质量分别约为127.20 kDa和164.48 kDa,亲水性系数分别为-0.584和-0.544,均不含跨膜结构域与信号肽；AcDNMT1a可同时定位于细胞核、细胞质和线粒体，AcDNMT1b可同时定位于细胞核与细胞质。AcDNMT1a和AcDNMT1b的三级结构模板均为4wxx.1.A。AcDNMT1a和AcDNMT1b均含有5个功能结构域，包括DNMT1-RFD、 zf-CXXC、 BAH＿Dnmt1-I、 BAH和DNA＿methylase。东方蜜蜂与黑大蜜蜂(Apis laboriosa)和大蜜蜂(Apis dorsata)的DNMT1a在进化树上聚为一支；东方蜜蜂与黑大蜜蜂的DNMT1b在进化树上聚为一支。AcDNMT1a和AcDNMT1b具有相似的蛋白质-蛋白质相互作用(protein-protein interaction, PPI)网络，与A0A2A3E789等10个蛋白形成互作网络。在幼虫发育过程中，AcDnmt1a和AcDnmt1b的表达量随日龄增长而降低，在6日龄幼虫中AcDnmt1a和AcDnmt1b的表达量较4日龄幼虫极显著下调。蜜蜂球囊菌接种后，AcDnmt1a的表达量在4～6日龄幼虫肠道中均下调表达；AcDnmt1b的表达量也在4～6日龄幼虫肠道中下调表达。AcDNMT1a和AcDNMT1b均为潜在的亲水性蛋白、非跨膜蛋白和胞内蛋白，主要分布在细胞核；东方蜜蜂和其他蜂的DNMT1具有较高的保守性；AcDnmt1a和AcDnmt1b在东方蜜蜂工蜂幼虫肠道发育过程中差异表达，并参与对蜜蜂球囊菌侵染的免疫应答。
Abstract：DNA methylation plays a crucial role in regulating biological processes such as insect growth and development, environmental adaptation and immune responses. However, studies on DNA methyltransferases(DNMTs) in Apis cerana are quite limited and related information is scarce. This study aimed to analyze the physicochemical properties and molecular characteristics of DNA methyltransferases 1a(AcDNMT1a) and DNA methyltransferases 1b(AcDNMT1b) in Apis cerana, and to detect the expression patterns of AcDnmt1a and AcDnmt1b during worker larvae gut development and in response to Ascosphaera apis infection, to provide a reference for further research on the molecular function and epigenetic regulation mechanisms of AcDNMT1a and AcDNMT1b.Bioinformatics was used to predict the physicochemical properties, molecular characteristics, and phylogenetic analysis of AcDNMT1a and AcDNMT1b, and to compare the domains and conserved motifs of DNMT1a and DNMT1b in Apis cerana and other insects. RT-qPCR was used to detect the relative expression levels of AcDnmt1a and AcDnmt1b during the gut development of Apis cerana worker larvae and in response to Ascosphaera apis infection. The relative molecular mass of AcDNMT1a and AcDNMT1b were approximately 127.20 kDa and 164.48 kDa, respectively, with hydrophilicity coefficients of-0.584 and-0.544, respectively. Both proteins lacked transmembrane domains and signal peptides. AcDNMT1a was localized in the nucleus, cytoplasm, and mitochondria, while AcDNMT1b was localized in the nucleus and cytoplasm. The tertiary structure templates of AcDNMT1a and AcDNMT1b were both 4wxx.1.A. AcDNMT1a and AcDNMT1b contained five functional domains, including DNMT1-RFD, zf-CXXC, BAH＿Dnmt1-I, BAH, and DNA＿methylase. DNMT1a of Apis cerana, Apis laboriosa and Apis dorsata clustered into one branch on the phylogenetic tree, and DNMT1b of Apis cerana and Apis laboriosa clustered into one branch. AcDNMT1a and AcDNMT1b had similar protein interaction networks, forming interaction networks with 10 proteins such as A0A2A3E789. During larval development, the expression levels of AcDnmt1a and AcDnmt1b decreased with increasing age, and the expression levels of AcDnmt1a and AcDnmt1b in 6-day-old larvae were significantly downregulated compared to 4-day-old larvae. After Ascosphaera apis inoculation, the expression level of AcDnmt1a was downregulated in the gut of 4-6 day old larvae; the expression level of AcDnmt1b was also downregulated in the gut of 4-6 day old larvae.AcDNMT1a and AcDNMT1b are potential hydrophilic proteins, non-transmembrane proteins, and intracellular proteins, mainly distributed in the nucleus; DNMT1 of Apis cerana and other bees had high conservation; AcDnmt1a and AcDnmt1b were differentially expressed during the gut development of Apis cerana worker larvae and were involved in the immune response to Ascosphaera apis infection.
杜宇，冯睿蓉，王杰，等，2021.中华蜜蜂6日龄幼虫响应蜜蜂球囊菌侵染的长链非编码RNA应答研究.微生物学报，61(5):1338-1358.[DU Y,FENG R R,WANG J,et al.,2021.Long non-coding RNA response of 6-day-old Apis cerana cerana larvae to Ascosphaera apis infection.Acta Microbiologica Sinica,61(5):1338-1358.]
刘亭亭，刘俊峰，王文祥，等，2012.中华蜜蜂DNA甲基化转移酶Dnmt3基因克隆及表达谱分析.昆虫学报，55(3):284-290.[LIU T T,LIU J F,WANG W X,et al.,2012.Cloning and expression profiling of the DNA methyltransferase Dnmt3 gene in the Chinese honeybee,Apis cerana cerana(Hymenoptera:Apidae).Acta Entomologica Sinica,55(3):284-290.]
张巧伶，苏立德，吕志强，2015.细菌感染对家蚕DNA甲基化与组蛋白乙酰化相关基因表达的影响.昆虫学报，58(9):941-949.[ZHANG Q L,SU L D,LYU Z Q,2015.Expression of DNA methylation and histone acetylation rela-ted genes in response to bacterial infection in the silkworm,Bombyx mori.Acta Entomologica Sinica,58(9):941-949.]
ATINBAYEVA N,VALENT I,ZENK F,et al.,2024.Inheri-tance of H3K9 methylation regulates genome architecture in Drosophila early embryos.EMBO J.,43(13):2685-2714.
BARADARAN E,MOHARRAMIPOUR S,ASGARI S,et al.,2019.Induction of DNA methyltransferase genes in Heli-coverpa armigera following injection of pathogenic bacteria modulates expression of antimicrobial peptides and affects bacterial proliferation.J.Insect Physiol.,118:103939.
BAYLIN S B,OHM J E,2006.Epigenetic gene silencing in can-cer:a mechanism for early oncogenic pathway addiction?.Nat.Rev.Cancer,6(2):107-116.
CARDOSO-JúNIOR C A M,EYER M,DAINAT B,et al.,2018a.Social context influences the expression of DNA methyltransferase genes in the honeybee.Sci.Rep.,8:11076.
CARDOSO-JúNIOR C A M,GUIDUGLI-LAZZARINI K R,HARTFELDER K,2018b.DNA methylation affects the lifespan of honey bee (Apis mellifera L.) workers—evidence for a regulatory module that involves vitellogenin expression but is independent of juvenile hormone function.Insect Biochem.Mol.Biol.,92:21-29.
CHEN W F,WANG Y,ZHANG W X,et al.,2021.Methionine as a methyl donor regulates caste differentiation in the European honey bee (Apis mellifera).Insect Sci.,28(3):746-756.
DIVITO EVANS A,FAIRBANKS R A,SCHMIDT P,et al.,2023.Histone methylation regulates reproductive diapause in Drosophila melanogaster.PLoS Genet.,19(9):e1010906.
EGGER G,LIANG G N,APARICIO A,et al.,2004.Epigene-tics in human disease and prospects for epigenetic therapy.Nature,429(6990):457-463.
ESTEKIZADEH A,LANDáZUR N,BARTEK J Jr,et al.,2018.Increased cytomegalovirus replication by 5-Azacytidine and viral-induced cytoplasmic expression of DNMT-1 in medulloblastoma and endothelial cells.Int.J.Oncol.,52(4):1317-1327.
GUO R,CHEN D F,DIAO Q Y,et al.,2019.Transcriptomic investigation of immune responses of the Apis cerana cerana larval gut infected by Ascosphaera apis.J.Invertebr.Pathol.,166:107210.
HORII T,HATADA I,2016.Regulation of CpG methylation by Dnmt and Tet in pluripotent stem cells.J.Reprod.Dev.,62(4):331-335.
HUANG H L,WU P,ZHANG S L,et al.,2019.DNA methy-lomes and transcriptomes analysis reveal implication of host DNA methylation machinery in BmNPV proliferation in Bombyx mori.BMC Genomics,20(1):736.
IVASYK I,OLIVOS-CISNEROS L,VALDéS-RODRíGUEZ S,et al.,2023.DNMT1 mutant ants develop normally but have disrupted oogenesis.Nat.Commun.,14(1):2201.
KAY S,SKOWRONSKI D,HUNT B G,2018.Developmental DNA methyltransferase expression in the fire ant Solenopsis invicta.Insect Sci.,25(1):57-65.
KIM D J,2025.The role of the DNA methyltransferase family and the therapeutic potential of DNMT inhibitors in tumor treatment.Curr.Oncol.,32(2):88.
LAW J A,JACOBSEN S E,2010.Establishing,maintaining and modifying DNA methylation patterns in plants and animals.Nat.Rev.Genet.,11(3):204-220.
LI Y Y,2021.Modern epigenetics methods in biological research.Methods,187:104-113.
LYKO F,MALESZKA R,2011.Insects as innovative models for functional studies of DNA methylation.Trends Genet.,27(4):127-131.
MALESZKA R,KUCHARSKI R,2022.Without mechanisms,theories and models in insect epigenetics remain a black box.Trends Genet.,38(11):1108-1111.
MOORE L D,LE T,FAN G P,2013.DNA methylation and its basic function.Neuropsychopharmacology,38(1):23-38.
NISHIYAMA A,NAKANISHI M,2021.Navigating the DNA methylation landscape of cancer.Trends Genet.,37(11):1012-1027.
SCHMITZ R J,LEWIS Z A,GOLL M G,2019.DNA methy-lation:shared and divergent features across eukaryotes.Trends Genet.,35(11):818-827.
TAKAYAMA S,DHAHBI J,ROBERTS A,et al.,2014.Genome methylation in D.Melanogaster is found at specific short motifs and is independent of DNMT2 activity.Genome Res,24(5):821-830.
TAMURA K,STECHER G,KUMAR S,2021.MEGA11:molecular evolutionary genetics analysis version 11.Mol.Biol.Evol.,38(7):3022-3027.
UNNIKRISHNAN A,FREEMAN W M,JACKSON J,et al.,2019.The role of DNA methylation in epigenetics of aging.Pharmacol.Ther.,195:172-185.
VIJAY A,JHA P K,GARG I,et al.,2019.micro-RNAs dependent regulation of DNMT and HIF1α gene expression in thrombotic disorders.Sci.Rep.,9:4815.
WANG Y,JORDA M,JONES P L,et al.,2006.Functional CpG methylation system in a social insect.Science,314(5799):645-647.
WEDD L,KUCHARSKI R,MALESZKA R,2022.DNA methyla-tion in honey bees and the unresolved questions in insect methylomics.Adv.Exp.Med.Biol.,1389:159-176.
WU P Y,GAO J X,LAN G M,et al.,2024.The role of RNA m5C modification in central nervous system diseases.Discov.Med.,36(187):1555-1571.
XU G F,ZHANG J,LYU H,et al.,2018.DNA methylation mediates BmDeaf1-regulated tissue- and stage-specific expression of BmCHSA-2b in the silkworm,Bombyx mori.Epigenetics Chromatin,11(1):32.
YIN L,CHUNG W O,2011.Epigenetic regulation of human β-defensin 2 and CC chemokine ligand 20 expression in gingival epithelial cells in response to oral bacteria.Mucosal Immunol.,4(4):409-419.
YOUNESIAN S,YOUSEFI A M,MOMENY M,et al.,2022.The DNA methylation in neurological diseases.Cells,11(21):3439.
ZENG Y,REN R,KAUR G,et al.,2020.The inactive Dnmt3b3 isoform preferentially enhances Dnmt3b-mediated DNA methylation.Genes Dev.,34(21-22):1546-1558.
基本信息:
DOI：10.13417/j.gab.044.001110
中图分类号:S891
引用信息:
[1]吴陶,樊念,杨佳润,等.东方蜜蜂DNMT1a和DNMT1b蛋白的分子特征、系统进化及基因表达模式[J].基因组学与应用生物学,2025,44(11):1110-1120.DOI:10.13417/j.gab.044.001110.
基金信息:
国家自然科学基金面上项目(32172792,32372943); 国家现代农业产业技术体系专项资金(CARS-44-KXJ7); 福建省自然科学基金面上项目(2025J01616)共同资助
2025-09-23
2025-09-23
2025-09-23