克氏原螯虾PcCRCN-L基因的克隆鉴定及在低氧胁迫下的表达响应  

Molecular Characterization and Expression Response Under Hypoxic-Reoxygenation Stress of a Crustacyanin-Like Gene in Procambarus clarkii

作　　者：韩一鸣 鲁苏皖 许志强 徐宇[2] 林海[2] 潘建林[1,2] 杨家新 李旭光[1,2] HAN Yiming;LU Suwan;XU Zhiqiang;XU Yu;LIN Hai;PAN Jianlin;YANG Jiaxin;LI Xuguang(College of Marine Science and Engineering,Nanjing Normal University,Nanjing 210023,China;Key Laboratory of Genetic Breeding and Cultivation for Freshwater Crustacean,Ministry of Agriculture and Rural Affairs,Freshwater Fisheries Research Institute of Jiangsu Province,Nanjing 210017,China;College of Marine Science and Fisheries,Jiangsu Ocean University,Lianyungang 222005,China)

机构地区：[1]南京师范大学海洋科学与工程学院,江苏南京210023 [2]江苏省淡水水产研究所农业农村部淡水虾蟹遗传育种与养殖重点实验室,江苏南京210017 [3]江苏海洋大学海洋科学与水产学院,江苏连云港222005

出　　处：《渔业科学进展》2024年第1期128-137,共10页Progress in Fishery Sciences

基　　金：江苏省种业振兴“揭榜挂帅”项目(JBGS(2021)119);国家重点研发计划创新专项(2020YFD0900303);江苏现代农业重点项目(BE2020348);江苏现代农业产业技术体系项目(JATS[2022]411);江苏省农业重大新品种创制项目(PZCZ201746)共同资助。

摘　　要：虾青蛋白(Crustacyanin,CRCN)在甲壳动物脂类代谢及低氧胁迫应激调控等方面具有重要的功能。为获取虾青蛋白在克氏原螯虾(Procambarus clarkii)性腺发育和低氧胁迫应答中的作用,本研究在克氏原螯虾肝胰腺组织中鉴定出1个类虾青蛋白PcCRCN-L基因的c DNA序列,分析了PcCRCN-L基因的结构特征和进化模式,研究了PcCRCN-L基因在不同组织与性腺不同发育时期的表达特征,探讨了Pc CRCN-L在低氧–复氧胁迫下的表达响应模式。结果显示,PcCRCN-L基因c DNA序列长2700 bp,其开放阅读框(ORF)长度为1587 bp,编码528个氨基酸残基;DNA序列长6130 bp,位于克氏原螯虾基因组的第12号染色体,包含5个外显子和4个内含子,内含子/外显子剪接方式符合GT-AG规则。Pc CRCN-L具有1个完整的lipocalin结构域,包含典型的序列保守区Ⅰ(SCR1)序列G-X-W、保守区Ⅱ(SCR2)序列T-D-Y和保守区Ⅲ(SCR3)序列精氨酸R。多序列比对与系统进化分析结果显示,PcCRCN-L独立于传统虾青蛋白A和C亚族,单独聚为一支。Pc CRCN-L在克氏原螯虾多个组织中均有表达,在肝胰腺中表达量最高;在性腺不同发育时期,肝胰腺以及卵巢组织中的Pc CRCN-L基因表达量显著降低;低氧胁迫下,肝胰腺组织中Pc CRCN-L表达量显著降低,复氧后显著升高。研究结果表明,PcCRCN-L与克氏原螯虾性腺发育密切相关,并参与了克氏原螯虾的低氧–复氧胁迫应激调控。Crustacyanin(CRCN),a family of lipocalin proteins specific to crustaceans,was primarily found in the exoskeleton of crustaceans.By binding with astaxanthin to form the astaxanthin-binding protein complex,it resets the proton at the end of astaxanthin and modifies the acidic site in astaxanthin,thereby regulating crustacean shell color.In addition to regulating shell colors,crustacyanin is involved in the transport of small molecule lipids such as steroid hormones and pheromones and has important biological functions in moult growth,gonadal and nervous system development,resistance to heavy metal lipid metabolism,and hypoxic stress.It was first discovered that the pigment in the blue shell of lobster could be extracted with ammonium chloride via a complex combination of organic bases and lipid pigments.It was later proposed that the pigment was a combination of astaxanthin and multimeric protein complex known asα-CRCN.This complex is composed of an octomer of dimericβ-CRCN subunits,with this dimer formed by two types of CRCN A and C in association with two astaxanthin molecules.Natural crustacyanins are allα-CRCN,comprising a total of 16 molecules.Procambarus clarkii,a member of the order Crustacea(Decapoda,Crayfish),is native to North America and is an important freshwater crustacean in China.It had been established that P.clarkii is susceptible to low oxygen stress during the culture process.The low oxygen environment inhibits the metabolic rate of P.clarkii,leading to increased susceptibility to pathogens;therefore,the ability of P.clarkii to tolerate and physiologically regulate stress due to other environmental factors was also affected,which can lead to irreversible damage and even death in severe cases.To understand the role of crustacyanin-like genes in gonadal development and hypoxic-reoxygenation stress in P.clarkii,a cDNA sequence of the PcCRCN-L gene was isolated from the hepatopancreas of this organism.The structural characteristics and evolutionary patterns of the PcCRCN-L gene were analyzed,and t

关 键 词：克氏原螯虾 类虾青蛋白 组织表达 性腺不同发育时期 低氧胁迫 

分 类 号：S917.4[农业科学—水产科学]