草鱼foxo4的基因克隆及其对饲料蛋白质营养调控的响应  

Molecular cloning and dietary protein regulation of foxo4 in grass carp(Ctenopharyngodon idella)

作　　者：徐哲华 易成琳 莫郁坚 杨立权 左安丽 杨瑞瑶 张鸣宇 瞿符发 唐建洲[1] 刘臻[1,2] 曹申平 XU Zhehua;YI Chenglin;MO Yujian;YANG Liquan;ZUO Anli;YANG Ruiyao;ZHANG Mingyu;QU Fufa;TANG Jianzhou;LIU Zhen;CAO Shenping(Hunan Provincial Key Laboratory of Nutrition and Quality Control of Aquatic Animals,Department of Biological and Chemical Engineering,Changsha University,Changsha 410022,China;State Key Laboratory of Freshwater Fish Developmental Biology,School of Life Sciences,Hunan Normal University,Changsha 410081,China)

机构地区：[1]长沙学院生物与化学工程学院,水生动物营养与品质调控湖南省重点实验室,湖南长沙410022 [2]湖南师范大学生命科学学院,淡水鱼类发育生物学国家重点实验室,湖南长沙410081

出　　处：《中国水产科学》2024年第1期14-28,共15页Journal of Fishery Sciences of China

基　　金：国家自然科学基金项目(32102813);湖南省自然科学基金项目(2021JJ40627);淡水生态与生物技术国家重点实验室开放课题项目(2022FB08)。

摘　　要：为探讨草鱼(Ctenopharyngodonidella)foxo4的分子特征及其对饲料蛋白营养调控响应的关系,通过同源克隆获得草鱼foxo4基因序列,其开放阅读框为1875 bp,编码624个氨基酸;系统进化树分析表明草鱼foxo4基因与黑头呆鱼(Pimephalespromelas)亲缘关系最近。对foxo4进行组织表达分析,发现foxo4基因mRNA在草鱼肌肉中表达水平最高(P<0.05),其次是心脏和肝,最后是肠、脑、脾和肾。此外,昼夜节律分析显示,foxo4在18:00表达量最高,而在24:00表达量最低。本研究进一步探究了菜粕、鱼粉和豆粕作为饲料蛋白对foxo4表达的调控,实验结果表明,在养殖14 d、21 d和35 d后,草鱼肠道中foxo4基因在菜粕组和鱼粉组相对于豆粕组均出现显著的上调(P<0.05),这表明草鱼foxo4基因的表达与饲料中蛋白源种类密切相关。不同比例L-丙氨酰-L-谷氨酰胺二肽(丙谷二肽)养殖实验结果显示,在0.5%~2%添加量范围内,随着饲料中丙谷二肽添加比例的增加,草鱼肠道中foxo4基因的表达呈逐渐下降趋势,其中在对照组表达量最高,在1.5%丙谷二肽添加组表达量最低。综上所述,草鱼foxo4基因表达具有组织特异性,且受到饲料蛋白源和二肽水平的调控影响。本研究为揭示foxo4基因在鱼类中的分子特征及其对蛋白质营养调控的响应提供了基础数据。The Forkhead box(Fox)protein family is a family of transcription factors with wing-like helical structured DNA-binding region and is widely distributed in biological groups ranging from yeast to mammals.The Fox protein family is divided into 19 subfamilies,A to S,among which the forkhead box O(FoxO)subfamily is the most intensively studied.Forkhead box O4(FoxO4)is a member of the FoxO subfamily,which includes FoxO1,FoxO3,and FoxO6 in mammals.These members share structural and functional similarities as well as regulations.In certain fish species,seven highly homologous members of the FoxO subfamily exist:FoxO1a,FoxO1b,FoxO3a,FoxO3b,FoxO4,FoxO6a,and Fox06b.The involvement of FoxO in the regulation of various physiological functions,including the cell cycle,apoptosis,DNA damage repair,oxidative stress,cell differentiation,and glucose metabolism,has been demonstrated in multiple studies.Furthermore,its activity is modulated through diverse mechanisms such as phosphorylation,acetylation,and ubiquitination.To explore the molecular characteristics of foxo4 in grass carp(Ctenopharyngodon idella)and its relationship with the nutritional regulation of dietary proteins,we obtained the grass carp foxo4 sequence through homologous cloning.Its open reading frame is 1875 bp,encoding 624 amino acids,and it consists of three functional domains:FH,Pfam(FOXO_KIX_bdg),and Pfam(FOXO_TAD).Amino acid sequence alignment analysis showed that the foxo4 gene in C.idella is highly homologous to that in Danio rerio,Homo sapiens,Mus musculus,and Electrophorus electricus.Analysis of codon usage bias revealed that foxo4 exhibited a strong preference for CUG and AGC codons across all examined species,whereas C.idella and Pimephales promelas shared certain similarities in their codon preferences,suggesting a close evolutionary relationship between them.Phylogenetic tree analysis showed that the foxo4 gene of grass carp has the highest homology to that of Pimephales promelas.Tissue expression analysis of foxo4 revealed that the mRNA expression le

关 键 词：草鱼 foxo4 基因克隆 组织分布 蛋白源 丙谷二肽 

分 类 号：S917[农业科学—水产科学]