摘要
脑缺血缺氧会引起长链非编码RNA(long non-coding RNA,lncRNA)表达变化。为了探究lncRNA BDNF-AS在氧糖剥夺/复氧复糖条件下的表达、定位及互作蛋白质,将SH-SY5Y细胞缺氧缺糖8 h、复氧复糖24 h,构建氧糖剥夺/复氧复糖细胞模型;采用CCK-8(Cell Counting Kit-8)法检测细胞活力;采用qRT-PCR检测核质中lncRNA BDNF-AS表达水平;利用pull-down和质谱技术对lncRNA BDNF-AS互作蛋白质进行鉴定;利用基因本体论(Gene Ontology,GO)、京都基因和基因组数据库(Kyoto Encyclopedia of Genes and Genomes,KEGG)分析互作蛋白质的功能及参与的通路;利用STRING数据库分析蛋白质-蛋白质相互作用网络。结果显示:氧糖剥夺/复氧复糖条件下,lncRNA BDNF-AS表达量显著升高,且胞质表达量显著高于胞核;氧糖剥夺/复氧复糖组有120种蛋白质可能与lncRNA BDNF-AS存在潜在的相互作用。进一步的生物信息学分析表明,lncRNA-BDNF-AS可能与UBA52、NKAP、TBK1、RAB1A、RPL38等蛋白质存在互作关系。综上可知,lncRNA BDNF-AS可能通过绑定自噬和凋亡相关蛋白质影响神经细胞功能。
Cerebral ischemia and hypoxia can cause the change of long non-coding RNA(lncRNA)expression.To investigate the expression and localization of lncRNA BDNF-AS and its interacting proteins under the condition of oxygen and glucose deprivation/reoxygenation(OGD/R),SH-SY5Y cells were treated with hypoxia for 8 h and reoxygenation(R)for 24 h to establish a OGD/R cell model.The cell viability was measured by Cell Counting Kit-8(CCK-8)assay.The expression level of lncRNA-BDNF-AS was detected by qRT-PCR in both the nucleus and cytoplasm.The interacting proteins of lncRNA BDNF-AS were analyzed by using pull-down method and mass spectrometry.Functional analysis of interacting proteins was performed using Gene Ontology(GO)and Kyoto Encyclopedia of Genes and Genomes(KEGG)enrichment analysis.The protein-protein interaction network was analyzed using STRING database.Under OGD/R condition,the expression of lncRNA BDNFAS was significantly increased,and was significantly higher in the cytoplasm than in the nucleus.In the OGD/R group,there were 120 kinds of specific proteins expressed in SH-SY5Y cells.Bioinformatic analysis showed that lncRNABDNF-AS might interact with proteins including UBA52,NKAP,TBK1,RAB1A and RPL38.Therefore,lncRNABDNF-AS mayaffect neuronal functionbybindingautophagy-and apoptosis-associatedproteins.
作者
李建明
唐亮
向勤
杨大为
项炬
LI Jianming;TANG Liang;XIANG Qin;YANG Dawei;XIANG Ju(School of Basic Medical Sciences,Changsha Medical University,Changsha 410219,Hunan,China;School of Computer Science and Engineering,Central South University,Changsha 410083,Hunan,China)
出处
《生命科学研究》
CAS
2023年第1期1-8,共8页
Life Science Research
基金
国家自然科学基金资助项目(81873780)
湖南省教育厅科学研究项目(21A0586,22A0662)
湖南省卫计委科学研究项目(2022-02082739)
长沙市杰出创新青年培养计划项目(kq2206058,kq2206056)
湖南省应用特色学科资助项目。
