摘要
Jun(Jun proto-oncogene,AP-1 transcription factor subunit)为组成转录因子AP-1的亚单位或亚家族成员,参与机体的免疫应答。为了进一步了解jun基因在鱼类抗病毒天然免疫方面的功能,以日本青鳉为研究对象,采用CRISPR/Cas9介导的基因编辑技术获得jun缺失的纯合子突变体品系,并对其在神经坏死病毒(nervous necrosis virus,NNV)感染过程中的特征展开研究。基因克隆结果显示,野生型日本青鳉的jun基因编码区全长921 bp,共编码306个氨基酸,含有N端的Jun超家族结构域和C端的bZIP超家族结构域,并在眼、脑、鳃、心脏、头肾、卵巢、精巢、肝脏、脾脏9个检测的组织中均有较高的表达。突变体jun-/-在基因组减少了124 bp,而蛋白只剩下59个氨基酸的N端序列,缺失了Jun超家族结构域和bZIP结构域。病毒感染实验结果显示,jun-/-仔鱼对于神经坏死病毒(nervous necrosis virus,NNV)的耐受能力相对野生型仔鱼有所提升;jun-/-成鱼眼、脑、鳃和肌肉组织内的病毒相对含量都显著低于野生型。进一步的qRT-PCR结果显示,在jun-/-青鳉中,fosl1的表达极显著下调,tbk1的表达降低,而irf3的表达极显著上调。以上结果表明,jun缺失能增强青鳉耐受NNV病毒的能力,jun基因可能是一个青鳉抗病毒信号通路的负调控因子。
Jun(Jun proto-oncogene,AP-1 transcription factor subunit)is a subunit or subfamily member of the transcription factor AP-1 and play an important role in host immune response.To further understand the function of jun gene in natural antiviral immunity of fish,the Japanese medaka(Oryzias latipes)was selected as the research fish model,and the homozygous mutant with jun deletion were obtained by CRISPR/CAS9 system and exposed to nervous necrosis virus(NNV)infection.Based on the gene cloning,the open reading frame of the medaka jun is 921 bp encoding 306 amino acids,containing N terminal jun super family structure domain and C terminal bZIP family structure domain.The RNA expression was highly detected in all examined tissues including the eyes,brain,gill,heart,kidney,ovary,testis,liver,and spleen.The mutant jun-/-was reduced by 124 bp in the genome,leaving only 59 amino acids N-terminal sequences,and missing the Jun superfamily domain and bZIP domain.Antiviral experiments were then carried out on larval and adult fish of the mutant strain by administrating NNV.The results showed that the disease resistance of larval mutants was enhanced,and the relative content of virus in adult tissues including the eye,brain,gill and muscle were significantly lower than in the wild type.The expression of jun and interferon-related genes fosl1,tbk1 and irf3 that was induced by NNV virus were detected by qRT-PCR.It was found that the expression of jun in the mutant was blocked compared to the wild type,the expression of fosl1 and tbk1 was differentially reduced,while the expression of irf3 was significantly up-regulated,indicating that the inhibition of jun expression may promote the release of irf3.These results suggest that the jun mutant has the ability of virus tolerance,and jun may be a negative regulator of the antiviral signaling pathway.
作者
王志
潘启华
陆可
罗君志
夏必琳
姜正正
申萍
刘红
陈天圣
WANG Zhi;PAN Qihua;LU Ke;LUO Junzhi;XIA Bilin;JIANG Zhengzheng;SHEN Ping;LIU Hong;CHEN Tiansheng(Key Laboratory of Freshwater Animal Breeding,Ministry of Agriculture and Rural Affairs/College of Fisheries,Huazhong Agricultural University,Wuhan 430070,China;Key Laboratory of Healthy Mariculture for the East China Sea,Ministry of Agriculture and Rural Affairs/College of Fisheries,Jimei University,Xiamen 361021,China)
出处
《华中农业大学学报》
CAS
CSCD
北大核心
2021年第6期152-160,共9页
Journal of Huazhong Agricultural University
基金
国家自然科学基金项目(31771648,31672653)
集美大学科研基金项目(ZQ2020003)
国家重点基础研究发展计划项目(2013CB967700)。
