与甲型流感病毒NP蛋白相互作用的宿主因子的筛选与分析  

Screening and Analyses of the Host Proteins Interacting with the Nucleoprotein of the Influenza-A Virus

作　　者：陈梦苹 崔晓兰[1] 郭姗姗[1] CHEN Mengping;CUI Xiaolan;GUO Shanshan(Institute of Chinese Materia Medica,China Academy of Chinese Medical Sciences,Beijing 100700,China)

机构地区：[1]中国中医科学院中药研究所,北京100700

出　　处：《病毒学报》2022年第2期385-393,共9页Chinese Journal of Virology

基　　金：国家自然科学基金项目(项目号:81773977),题目:与流感病毒NP蛋白发生相互作用的宿主靶点辨识及栀子环烯醚萜苷的干预作用研究。

摘　　要：筛选与甲型流感病毒(Influenza A virus,IAV)NP蛋白相互作用的人类宿主蛋白,并对其进行分析,为抗流感病毒新药的研发提供依据。利用蛋白质芯片技术,将两张HuProtTM 20K芯片分别与实验组(NP蛋白)和对照组(灭活生物素)样本杂交孵育,扫描芯片读取数据。计算各位点的原始信号强度I(I=F635 Median/B635 Media)用于数据分析,并进行Z-Score标准化处理,筛选出芯片上两重复位点均满足Z-Score≥3的蛋白,即认为该蛋白与样本相结合。对NP样本特异性结合蛋白进行GO、KEGG分析,计算这些蛋白在各组内两原始信号强度的均值IMean及组间比值IMean_Ratio(实验组_vs_对照组),筛选出IMean_Ratio≥1.4的NP显著特异结合蛋白,将实验组显著特异结合蛋白导入Human proteome map、VirHostNet3.0数据库,在malacards数据库中检索Influenza,将得到的流感相关蛋白与显著特异结合蛋白进行比较分析。筛选出NP宿主蛋白176种,GO分析结果显示,这些蛋白主要是作为结合蛋白参与剪接过程。KEGG富集分析结果显示NP宿主蛋白显著富集到剪接体通路、内吞通路、抗原加工与呈递通路(P<0.05)。有5个显著特异结合蛋白同时参与到与流感最相关的两条通路中,这5个蛋白分别是RASAL3、PTK2、PSMC4、PSME1和CD80,其中,除蛋白RASAL3、CD80外,蛋白PTK2、PSME1和PSMC4在成人肺组织中皆有表达。查阅相关资料,蛋白PTK2已被证实可与甲型流感病毒NP蛋白相互作用,参与调控病毒的复制过程。后续可进一步展开实验,验证蛋白PSMC4、PSME1与甲型流感病毒NP蛋白的相互作用,并探讨这两种蛋白在流感病毒感染过程中发挥的作用。Human host proteins interacting with the nucleoprotein(NP)of the influenza-A virus were screened by the protein chip and analyzed to further understand the pathogenic mechanism of the influenza virus. Two HuProtTM 20 K chips were incubated with experimental group(NP protein) and control group(inactivated biotin)samples respectively. The chips were scanned. The original signal strength I(I = F635 Median/B635 Median)on each spot was calculated for data analysis,and Z-Score standardization was performed on I. Only proteins having a Z-Score ≥3 on both spots were considered to be bound to the sample. Specific binding proteins in the experimental group were screened and analyzed using Gene Ontology(GO) and Kyoto Encyclopedia of Genes and Genomes(KEGG)databases. The IMean of the two repeat spots in each group and the IMean_Ratio between groups(experimental group _vs_ control group)were calculated. Proteins with an IMean_Ratio ≥1.4 were significantly specific binding proteins in the experimental group. These proteins underwent human proteome mapping in adult lung tissue and VirHostNet. Then,the retrieved influenza-related proteins were compared with the significantly specific binding proteins. A total of 176 NP-specific binding proteins were screened. Enrichment analyses using the GO database showed that the molecular function of these proteins was primarily“binding function”,and the main biological processes were“RNA and mRNA splicing”.Signaling-pathway analyses using the KEGG database showed that these proteins were significantly rich in“Spliceosome”,“Endocytosis”and“Antigen processing”and presented three signaling pathways(P<0.05).Two pathways and the proteins most closely associated with influenza were retrieved in the MalaCards database,and five significantly specific binding proteins were associated with both pathways,among which PPK2,PSME1 and PSMC4 were expressed in higher abundance in adult lung tissue. The protein PTK2 was shown to interact with NP and to participate in the regulation of

关 键 词：NP蛋白 人类蛋白质芯片 宿主因子 生物信息学分析 

分 类 号：R373.1[医药卫生—病原生物学] R51[医药卫生—基础医学]