大黄鱼UBXN1基因鉴定及其过表达后的转录组分析  

Identification of UBXN1 and transcriptome analysis after its overexpression in large yellow croaker(Larimichthys crocea)

作　　者：张东玲[1] 唐欣 王志勇[1] ZHANG Dongling;TANG Xin;WANG Zhiyong(Key Laboratory of Healthy Mariculture for the East China Sea,Ministry of Agriculture and Rural Affairs,Jimei University,Xiamen 361021,China)

机构地区：[1]集美大学,农业农村部东海海水健康养殖重点实验室,福建厦门361021

出　　处：《水产学报》2024年第4期336-345,共10页Journal of Fisheries of China

基　　金：国家自然科学基金(32172964);福建省自然科学基金重点项目(2022J02042)。

摘　　要：为探究一种包含泛素调节性X结构域的蛋白(ubiquitin regulatory X domain-containing protein,UBXN1)在大黄鱼抗盾纤毛虫感染中的作用,以及可能涉及的免疫信号通路。本实验克隆鉴定了大黄鱼UBXN1基因,并利用在线软件对其序列特征进行生物信息学分析;采用实时荧光定量PCR(qRT-PCR)检测UBXN1在健康大黄鱼各组织中的表达,及盾纤毛虫感染后的诱导表达变化;并进行了UBXN1的亚细胞定位;转录组测序分析了UBXN1过表达前后的差异表达基因。结果显示,UBXN1基因cDNA全长为915 bp,编码304个氨基酸。蛋白多重序列比对和结构预测表明UBXN1是一个进化保守的蛋白,包含UBA和UBX结构域。qRT-PCR分析表明UBXN1在所检测的11种组织中均有表达,脑中表达量最高,其次是肝脏、心脏和肾脏,在肌肉中表达量最低;盾纤毛虫感染大黄鱼后,UBXN1在脾脏、脑、肝脏和肾脏中表达量早期显著升高,后期逐步恢复至正常水平。亚细胞定位分析表明,UBXN1在大黄鱼肾脏细胞质和细胞核中均有表达。在293T细胞过表达UBXN1,转录组差异表达分析筛选到12个上调基因,4个下调基因,其中RPL41/RPL39/XIST/RNA45SN4表达量显著增加,而ATP8/ND4L表达量显著减少。研究表明UBXN1在大黄鱼抗寄生虫免疫应答中发挥重要作用。本实验为进一步研究UBXN1的免疫信号通路奠定基础。Scuticociliatia disease is a major threat to large yellow croaker Larimichthys crocea at the fry stage.In the previous study,a ubiquitin regulatory X domain-containing protein(UBXN1)gene was found to be involved in Scuticociliatia disease resistance through genome-wide association analysis.To further confirm its gene function in resistance against Scuticociliatia,we identified UBXN1 gene sequence and investigated its expression characteristics,as well as signal pathway.The complete cDNA sequence of UBXN1 was 915 bp,encoding 304 amino acids.The putative UBXN1 protein was evolutionarily conserved and contained a UBA and a UBX domain.Moreover,UBXN1 had varying degrees of expressions in different tissues,among which the highest expression was observed in brain,moderate expression was in liver,heart and kidney,and the lowest expression was in muscle by qRT-PCR detection.It also showed that induced expression was found in spleen,brain,liver,and kidney during the early phase of infection,and the expression gradually recovered to the control level at the late phase.Further subcellular localization analysis showed that UBXN1 uniformly distributed in cytoplasm and nucleus.In addition,16 differentially expressed genes were found in a comparative transcriptome analysis between the UBXN1 over expression group and control vector group,including up-regulated genes RPL41/RPL39/XIST/RNA45SN4 and down-regulated genes ATP8/ND4L.These findings provide new insights into finfish UBXN1 and its potential regulatory signaling pathway.

关 键 词：大黄鱼 包含泛素调节性X结构域的蛋白(UBXN1) 盾纤毛虫 转录组 免疫 

分 类 号：Q786[生物学—分子生物学] S942.1[农业科学—水产养殖]