意大利蜜蜂lncRNA15837的调控作用和表达模式分析  

Analysis of the regulatory role and expression pattern of lncRNA15837 in Apis mellifera ligustica(Hymenoptera:Apidae)

作　　者：宋宇轩 李坤泽 臧贺 刘小玉 吴陶 徐细建 刘锋 骆群 邱剑丰 陈大福[1,2,3] 郭睿 SONG Yu-Xuan;LI Kun-Ze;ZANG He;LIU Xiao-Yu;WU Tao;XU Xi-Jian;LIU Feng;LUO Qun;QIU Jian-Feng;CHEN Da-Fu;GUO Rui(College of Bee Science and Biomedicine,Fujian Agriculture and Forestry University,Fuzhou 350002,China;National&Local United Engineering Laboratory of Natural Biotoxin,Fuzhou 350002,China;Apitherapy Research Institute of Fujian Province,Fuzhou 350002,China;Apicultural Research Institute of Jiangxi Province,Nanchang 330000,China)

机构地区：[1]福建农林大学蜂学与生物医药学院,福州350002 [2]天然生物毒素国家地方联合工程实验室,福州350002 [3]福建省蜂疗研究所,福州350002 [4]江西省养蜂研究所,南昌330000

出　　处：《昆虫学报》2025年第2期144-153,共10页Acta Entomologica Sinica

基　　金：国家自然科学基金面上项目(32372943,32172792);国家现代农业产业技术体系专项资金(蜜蜂)(CARS-44-KXJ7);福建省自然科学基金面上项目(2022J01133);福建农林大学科技创新专项基金项目(KFb22060XA);江西省重点研发计划项目(20212BBF63008);福建省大学生创新创业训练计划项目(X202310389077,X202310389073)。

摘　　要：【目的】本研究旨在通过解析意大利蜜蜂Apis mellifera ligustica lncRNA15837的调控方式和作用,并检测lncRNA15837在工蜂不同发育阶段和成虫不同组织及幼虫应答蜜蜂球囊菌Ascosphaera apis侵染中的表达模式,为深入探究lncRNA15837的调控功能与机制提供参考和依据。【方法】基于已获得的意大利蜜蜂7和10日龄工蜂成虫中肠高质量转录组数据,利用Miranda,RNAhybrid和TargetScan软件预测lncRNA15837靶向的miRNA及miRNA靶向的mRNA,再通过Cytoscape v3.7.1软件构建并可视化竞争性内源RNA(competing endogenous RNA,ceRNA)调控网络。使用Blast工具将靶mRNA比对至GO和KEGG数据库以获得相应的功能和通路注释。通过RT-PCR验证lncRNA15837在刚出房工蜂成虫不同组织(触角、咽下腺、脑、表皮、中肠、脂肪体和毒腺)中的表达。利用RT-qPCR检测lncRNA15837在工蜂的不同发育阶段(卵、3日龄幼虫、1和2日龄预蛹、4日龄蛹及1,2,6,12,15和18日龄工蜂成虫)、刚出房工蜂成虫不同组织(触角、咽下腺、脑、表皮、中肠、脂肪体和毒腺)和3日龄幼虫感染蜜蜂球囊菌后4,5和6日龄幼虫肠道中的表达量。【结果】LncRNA15837可靶向ame-miR-21-x和ame-miR-0046-3p等29个miRNA,进而靶向1559条mRNA,形成1个复杂的ceRNA网络;上述靶mRNA可注释到436个GO条目,其中114条靶mRNA涉及139个分子功能相关条目,115条靶mRNA涉及257个生物学进程相关条目,67条靶mRNA涉及40个细胞组分相关条目;可注释到224条KEGG通路,其中28条靶mRNA注释到内吞、吞噬细胞和溶酶体等细胞免疫通路,以及157条靶mRNA注释到JAK-STAT,Toll和Imd,NF-kappa B和Toll样受体信号通路等体液免疫通路;lncRNA15837在工蜂卵、幼虫、预蛹、蛹和成虫中差异表达,在1日龄预蛹中的表达量最低,而在3日龄幼虫中的表达量最高;lncRNA15837在刚出房工蜂成虫的触角、毒腺、脑、中肠、咽下腺、脂肪体和表皮中均有表达但表达量存在差【Aim】The objective of this study is to provide a reference and basis for further exploring the regulatory function and mechanism of lncRNA15837 in Apis mellifera ligustica by analyzing the regulatory mode and role of lncRNA15837 and detecting its expression patterns in different developmental stages and different adult tissues,and in larvae of A.mellifera ligustica in response to Ascosphaera apis infection.【Methods】Based on the obtained high-quality transcriptome data from the midguts of the 7-and 10-day-old adult workers of A.mellifera ligustica,Miranda,RNAhybrid and TargetScan software was employed to predict lncRNA15837-targeted miRNAs and miRNA-targeted mRNAs,and construction and visualization of the competing endogenous RNA(ceRNA)regulatory network were performed using Cytoscape v3.7.1 software.By using Blast tool,target mRNAs were aligned to GO and KEGG databases to gain their corresponding functional and pathway annotations,respectively.The expression of lncRNA15837 in different tissues(antennae,hypopharyngeal glands,brain,cuticle,midgut,fat body and venom gland)of the newly emerged adult workers was confirmed by RT-PCR.RT-qPCR was utilized to detect the expression levels of lncRNA15837 in different developmental stages of workers(eggs,3-day-old larvae,1-and 2-day-old prepupae,4-day-old pupae,and 1-,2-,6-,12-,15-and 18-day-old adult workers),in different tissues(antennae,pharyngeal glands,brain,cuticle,midgut,fat body and venom gland)of the newly emerged adult workers,and in the guts of the 4-,5-and 6-day-old larvae after infection of the 3-day-old larvae with Ascosphaera apis.【Results】LncRNA15837 could target 29 miRNAs such as ame-miR-21-x and ame-miR-0046-3p and further target 1559 mRNAs,forming a complex ceRNA network.The aforementioned target mRNAs could be annotated to 436 GO terms,among them 114 target mRNAs were involved in 139 molecular function-related terms,115 target mRNAs were involved in 257 biological process-related terms,67 target mRNAs were involved in 40 cellular component-rel

关 键 词：意大利蜜蜂 竞争性内源RNA 长链非编码RNA 时空表达谱 蜜蜂球囊菌 

分 类 号：S891[农业科学—特种经济动物饲养]