ATP5J促进猪繁殖与呼吸综合征病毒复制研究  

ATP5J facilitates porcine reproductive and respiratory syndrome virus replication

作　　者：王飞飞 刘珂[2] 郇贝丽 欧安妮 魏建超[2] 李蓓蓓[2] 邵东华[2] 邱亚峰[2] 钱莺娟[1] JUNG Yong-Sam 马志永[2] WANG Feifei;LIU Ke;HUAN Beili;OU Anni;WEI Jianchao;LI Beibei;SHAO Donghua;QIU Yafeng;QIAN Yingjuan;JUNG Yong-Sam;MA Zhiyong(MOE Joint International Research Laboratory of Animal Health and Food Safety,College of Veterinary Medieine,Nanjing Agricultural University,Nanjing 210095,China;Shanghai Veterinary Researeh Institute,Chinese Aeademy of Agrieultural Seienees,Shanghai 200241,China)

机构地区：[1]南京农业大学动物医学院/教育部动物健康与食品安全国际合作联合实验室,江苏南京210095 [2]中国农业科学院上海兽医研究所,上海200241

出　　处：《畜牧与兽医》2018年第8期54-58,共5页Animal Husbandry & Veterinary Medicine

基　　金：973计划(2014CB542702)

摘　　要：为了研究猪繁殖与呼吸综合征病毒（porcine reproductive and respiratory syndrome virus,PRRSV）复制与细胞调控酶ATP5J的关系,本研究首先对猪源ATP5J基因进行了抗原表位分析,选取并合成了3段抗原小肽,制备ATP5J的多克隆的抗体。用制备的多抗检测了ATP5J质粒在293T细胞中过表达的情况。然后,用PRRSV感染猪肺泡巨噬细胞（PAMs）,利用qPCR和Western blot检测PRRSV感染对PAM细胞中ATP5J表达的影响。最后,我们通过小RNA干扰以及过表达ATP5J,使用qPCR和Western blot技术,在PAM和Marc-145细胞上检测ATP5J变化对PRRSV复制增殖的影响。结果显示：PRRSV感染能够导致宿主细胞ATP5J蛋白表达下调;ATP5J下调时,PRRSV复制减弱;ATP5J过表达时,PRRSV复制增强,说明ATP5J能够促进PRRSV复制增殖。进一步分析证明：ATP5J促进PRRSV复制功能没有剂量依赖性。综上,PRRSV感染能够导致宿主细胞ATP5J蛋白表达下调,而ATP5J具有促进PRRSV复制的作用。该研究为阐明PRRSV与宿主相互作用提供了新思路,为解析PRRSV免疫抑制和免疫逃逸机制奠定了基础。In order to study the relationship between porcine reproductive and respiratory syndrome virus （PRRSV） replication and cell regulatory enzyme ATP5J, the ATPSJ amino acid sequence of Sus scrofa species was analyzed and three small peptides were synthesized as antigens for generation of rabbit polyelonal antibodies against ATPSJ. The polyclonal antibodies were prepared to detect the overexpression of ATP5J in 293T cells. PRRSV was used to infect pig alveolar macrophages （PAMs）. Then, the expression of ATP5J in PRRSV-infected PAM cells was measured by real-time quantitative PCR （qPCR） and Western blot. Finally, qPCR and Western blot were performed to measure the effect of ATP5J on PRRSV replication in PAM and Marc-145 cells via either ATP5J silencing or overexpression. The results in-dicated that ATP5J mRNA and the protein levels were downregulated after infection with PRRSV. When ATP5J was downregulated, PRRSV replication was suppressed. In contrast, when ATPSJ was overexpressed, PRRSV replication was enhanced. These findings suggested that ATP5J promoted PRRSV replication. Further analysis showed that ATP5J promoted PRRSV replication in a dose-independent manner. In conclusion, PRRSV infection leads to downregulation of ATPSJ expression in host cells, but ATP5J promotes PRRSV replication. The present study provides new insights into the interaction between PRRSV and host cells and lays a foundation for exploring mechanisms of im-munosuppression and immune escape of PRRSV.

关 键 词：猪繁殖与呼吸综合征病毒 ATP5J 基因调节 病毒复制 

分 类 号：S852.6[农业科学—基础兽医学]