摘要
为了探究斑马鱼(Danio rerio)组织蛋白酶B(Cathepsin B,Ctsb)在脂肪细胞分化中的功能,本文利用基因过表达和敲降技术研究了Ctsb对小鼠脂肪前体细胞(3T3-L1)分化的影响。生物信息学分析显示,斑马鱼基因组中存在2个组织蛋白酶B基因(ctsb):cathepsin ba(ctsba)和cathepsin bb(ctsbb)。其中斑马鱼ctsba编码的氨基酸序列同哺乳动物CTSB的氨基酸序列相似性更高一些,它们的二级结构和三级结构也十分相似。在成体斑马鱼中,ctsba基因在卵巢、脂肪组织、精巢、肌肉、脾、肠、心、肾脏、鳃、脑和肝脏等不同组织中都有表达,其中在卵巢、脂肪组织和精巢中表达较高。通过慢病毒表达载体在3T3-L1细胞中过表达斑马鱼ctsba,可显著增加脂滴的形成;而敲降内源性的ctsb基因则显著减少了脂滴的形成,表明ctsba可促进3T3-L1脂肪前体细胞向脂肪细胞分化。另外,实时定量PCR技术检测显示,斑马鱼ctsba过表达可上调PPARγ、C/EBPα、FAS、ACC1和FABP4等成脂基因的表达,敲降ctsb基因则相反。总之,本研究揭示了斑马鱼Ctsba调控脂肪细胞分化的功能。
To explore the role of cathepsin B(Ctsb)in adipocyte differentiation in zebrafish(Danio rerio),we investigated the effects of Ctsb on adipocyte precursors(3T3-L1)differentiation in mice using gene overexpression and knockout techniques.Bioinformatics analysis showed that there were two cathepsin B genes(ctsb)in the zebrafish genome:cathepsin ba(ctsba)and cathepsin bb(ctsbb),in which the amino acid sequence encoded by zebrafish ctsba is more similar to that of mammalian CTSB,and their secondary and tertiary structures are also very similar.In adult zebrafish,ctsba gene is expressed in different tissues such as ovary,adipose tissue,testis,muscle,spleen,intestine,heart,kidney,gill,brain and liver,with higher expression in ovary,adipose tissue and testis.Overexpression of zebrafish ctsba in 3T3-L1 cells by lentiviral expression vector can significantly increase the formation of lipid droplets.However,knockdown of endogenous ctsb gene significantly reduced the formation of lipid droplets,suggesting that ctsba can promote the differentiation of 3T3-L1 adipocyte precursor cells into adipocytes.In addition,real-time quantitative PCR showed that overexpression of ctsba could up-regulate the expression of PPARγ,C/EBP-α,FAS,ACC1 and FABP4 lipogenic genes in zebrafish,while knockout of ctsb gene was opposite.In conclusion,this study revealed the function of Ctsba in regulating adipocyte differentiation in zebrafish.
作者
风英
邢泽宇
李欣怡
王鑫媛
刘振辉
Feng Ying;Xing Zeyu;Li Xinyi;Wang Xinyuan;Liu Zhenhui(Institute of Marine Biodiversity and Evolution,College of Marine Life Sciences,Ocean University of China,Qingdao 266003,China)
出处
《中国海洋大学学报（自然科学版）》
CAS
CSCD
北大核心
2024年第6期88-97,共10页
Periodical of Ocean University of China
基金
国家自然科学基金项目(31970429,32171139)资助。
