鳞头犬牙南极鱼LeptinB基因在抵御低温应激中的作用  

Role of the Dissostichus mawsoni LeptinB Gene in Cellular Cold Resistance

作　　者：万金铭 王映[1] 陈良标[1,2] WAN Jinming;WANG Ying;CHEN Liangbiao(International Research Center for Marine Biosciences,Ministry of Science and Technology,Key Laboratory of Exploration and Utilization of Aquatic Genetic Resources,Ministry of Education,Shanghai Ocean University,Shanghai 201306,China;National Demonstration Center for Experimental Fisheries Science Education,Shanghai Ocean University,Shanghai 201306,China)

机构地区：[1]上海海洋大学国家海洋生物科学国际联合研究中心水产种质资源发掘与利用教育部重点实验室,上海201306 [2]上海海洋大学水产科学国家级实验教学示范中心,上海201306

出　　处：《渔业科学进展》2023年第6期97-106,共10页Progress in Fishery Sciences

基　　金：国家重点研发计划项目(2018YFD0900600);上海市科委“一带一路”国际联合实验室项目(19590750500);上海市教委研创新计划项目(2017-01-07-00-10-E00060)共同资助。

摘　　要：为了探究瘦素基因(LeptinB,LepB)在斑马鱼肝脏细胞系(ZFL)低温应激中的作用,本研究根据鳞头犬牙南极鱼(Dissostichusmawsoni)LepB基因(DM-LepB)编码的氨基酸序列,克隆到构建的pTOL2-actin-EGFP质粒中(启动子为斑马鱼的β-actin)。选取Eco RⅠ和Bam HⅠ为限制酶,设计构建了pTOL2-LepB+HIS-EGFP表达载体,并转染至ZFL细胞中。选择致死温度10℃进行实验,采用流式细胞术检测了低温胁迫下细胞的存活率,使用增强型CCK-8试剂盒检测了低温条件下细胞的生长增殖情况,使用荧光探针DHE检测了细胞活性氧(ROS)含量。结果显示,Lep B基因的过表达能有效减少细胞ROS的产生,减轻细胞凋亡以应对低温胁迫。分析还显示,DM-LepB的过表达也有利于维持低温刺激下的胞内ATP水平与线粒体状态,有效减轻低温刺激对细胞的凋亡和坏死作用,对细胞冷应激起到保护作用。采用油红O染色和甘油三酯(TG)实验检测发现,DM-LepB基因能减缓低温刺激时细胞脂质的消耗,较多的脂质保留可减弱低温刺激对细胞的伤害,使得细胞能更好地抵抗低温损害。研究结果为揭示南极鱼类的低温适应分子机制提供了基础资料。。Variation in fish temperature is determined by changes in the external aquatic environment,and temperature serves as a crucial regulatory factor for the behavior,physiology,and molecular processes of fish.A rapid decline in water temperature can elicit a strong stress response in fish,potentially inhibiting growth and even leading to mortality.Low temperature stress significantly impacts various life processes of organisms,including growth and development,energy metabolism,and reproductive development.In recent years,there has been increasing research on the effects of low-temperature stress on organisms,with a greater focus at the molecular level.Low temperatures induce a decrease in cellular activity and even growth arrest,resulting in apoptosis.Apoptosis is a form of programmed cell death ubiquitous in the biological world,and it plays a vital role in maintaining the normal physiological function of an organism.Once apoptosis occurs,it is irreversible and cannot be stopped.Temperature regulation is crucial for fish,and current knowledge of the role of leptin in fish primarily focuses on the regulation of feeding,lipid energy metabolism,and reproduction,with limited reports on its role in temperature regulation.Leptin,a protein hormone produced by adipocytes,has long been recognized as a product of the obese gene,which regulates organismal metabolism,neuroendocrine function,and other physiological processes.In animals,it exerts diverse physiological functions related to cell growth,proliferation,apoptosis,and metabolism.The homology of the leptin gene in fish is relatively low compared to that in mammals,and multiple leptin proteins encoded by multiple leptin genes can be produced due to genomic duplication events.Regarding the role of leptin in temperature regulation,more research has been conducted on mammals,mainly related to the regulation of energy metabolism and the influence on temperature adaptation evolution.There has been limited research on the regulatory mechanism of leptin in fish during cold stres

关 键 词：鳞头犬牙南极鱼 LeptinB 低温应激 细胞凋亡 脂质代谢 

分 类 号：Q786[生物学—分子生物学] S917.4[农业科学—水产科学]