C/EBPα通过结合鸡神经调节蛋白4基因外显子2正调控其表达  

C/EBPαPositively Regulates Chicken NRG 4 Gene Expression via Binding to Its Exon 2

作　　者：罗昊玉 黄佳新 高帆 闫晓红 牟芳 王宁 LUO Hao-Yu;HUANG Jia-Xin;GAO Fan;YAN Xiao-Hong;MU Fang;WANG Ning(Key Laboratory of Chicken Genetics and Breeding,Ministry of Agriculture and Rural Affairs,Harbin 150030,China;Key Laboratory of Animal Genetics,Breeding and Reproduction,Education Department of Heilongjiang Province,Harbin 150030,China;Department of Animal Genetics,Breeding and Reproduction,College of Animal Science and Technology,Northeast Agricultural University,Harbin 150030,China)

机构地区：[1]农业农村部鸡遗传育种重点实验室,哈尔滨150030 [2]黑龙江省普通高等学校动物遗传育种与繁殖重点实验室,哈尔滨150030 [3]东北农业大学动物科学技术学院动物遗传育种与繁殖系,哈尔滨150030

出　　处：《中国生物化学与分子生物学报》2024年第11期1608-1617,共10页Chinese Journal of Biochemistry and Molecular Biology

基　　金：国家自然科学基金项目(No.U23A20230);黑龙江省自然科学基金(优秀青年项目)(No.YQ2023C008)资助。

摘　　要：神经调节蛋白4(NRG4)是一个新发现的脂肪细胞因子,它在机体能量平衡、糖脂代谢等许多生理过程中发挥重要调控作用。目前,NRG 4基因的转录调控机制尚不清楚。我们前期的5′RACE分析显示,鸡NRG 4基因转录起始位点(TSS)弥散分布于一段约200 bp的区域,提示鸡NRG 4基因的启动子为分散型启动子(dispersed promoter)。本研究开展了鸡NRG 4基因近端外显子和内含子的启动子活性分析,报告基因分析显示,包含NRG 4基因外显子1、内含子1和外显子2的基因组片段(-122/+452)具有很强的双向启动子活性。生物信息学分析显示,鸡NRG 4基因外显子2上存在1个转录因子CCAAT增强子结合蛋白α(C/EBPα)的潜在结合位点;报告基因分析发现,删除或突变C/EBPα结合位点会导致C/EBPα丧失对NRG 4基因片段(-122/+452)启动子活性的调控作用。基因表达相关性分析显示,鸡脂肪组织中C/EBPα和NRG 4基因的mRNA表达存在显著正相关(P<0.01)。进一步的脂肪组织ChIP-qPCR分析显示,C/EBPα直接结合鸡NRG 4基因的外显子2。本研究发现,鸡NRG 4基因的近端外显子和内含子具有双向启动子活性,C/EBPα通过直接结合外显子2调控鸡脂肪组织NRG 4基因的表达。Neuregulin 4(NRG4)is a newly discovered adipokine,which plays crucial roles in many physiological processes such as energy balance,and glucose and lipid metabolism.The transcriptional regulation of NRG 4 gene remains largely unknown.Our previous 5′RACE analysis showed that the transcription start sites(TSS)of chicken NRG 4 gene were dispersed over a region of 200 bp,suggesting that chicken NRG 4 gene possesses a dispersed promoter.In the present study,the promoter activity of the proximal exon and intron of chicken NRG 4 gene was analyzed.Dual-luciferase reporter assay demonstrated that the genomic fragment(-122/+452)containing exon 1,intron 1,and exon 2 of chicken NRG 4 gene had strong bidirectional promoter activity.Bioinformatics analysis showed that there was a putative binding site of the transcription factor CCAAT enhancer-binding proteinα(C/EBPα)in exon 2 of chicken NRG 4 gene.Reporter gene assay showed that both deletion and site-directed mutagenesis of the C/EBPαbinding site abrogated the regulatory effect of C/EBPαon the promoter activity of the-122/+452 fragment of chicken NRG 4 gene.Gene expression correlation analysis showed that C/EBPαand NRG 4 gene expression were positively correlated in chicken adipose tissues.ChIP-PCR revealed that C/EBPαdirectly binds to the exon 2 of NRG 4 gene in chicken adipose.In conclusion,the proximal exon and intron of chicken NRG 4 gene has bidirectional promoter activity,and C/EBPαdirectly regulates NRG 4 gene expression in chicken adipose via binding to its exon 2.

关 键 词：神经调节蛋白4 近端外显子和内含子 启动子活性 转录调控 CCAAT增强子结合蛋白α(C/EBPα) 

分 类 号：Q75[生物学—分子生物学]