摘要
流感病毒核蛋白(NP)是由病毒RNA节段5编码的主要结构蛋白,与病毒基因组RNA及聚合酶(PB1、PB2、PA)构成核糖核蛋白(vRNP)复合体,是病毒基因组的转录和复制的基本功能单位。本实验室采用酵母双杂交技术从人细胞系cDNA文库中筛选与流感病毒NP蛋白相互作用的宿主蛋白U1 small nuclear ribonucleoprotein A(SNRPA)。本研究通过酵母回交验证、免疫共沉淀(Co-IP)和GST pull down试验进一步证实NP与SNRPA之间存在直接相互作用。利用siRNA干扰基因的表达后,流感病毒的复制滴度在24 h和48 h分别下降2.7倍和1.75倍,表明SNRPA蛋白表达对流感病毒的复制发挥正调控作用。本研究丰富了流感病毒蛋白与宿主蛋白之间的蛋白质调控网络,为进一步研究流感病毒复制调控的机制奠定了基础。
Influenza virus nucleoprotein （NP） is a major structural protein encoded by the 5th segment of viral genome, which associates with viral RNA and three polymerase proteins （PB1, PB2 and PA） to form viral ribonucleoprotein （vRNP） complex, the basic functional unit for transcription and replication of the viral genome. In this study, we identified host protein of U1 small nuclear ribonucleoprotein A （SNRPA） as an interacting partner of NP protein by using yeast two-hybrid screening from a cDNA library prepared from human cell lines. The interaction between NP and SNRPA was further validated by yeast backcross verification, co-immunoprecipitation （Co-IP） and GST pull-down assay. Moreover, knock down of SRNPA expression in A549 cell by siRNA was found to reduce virus titers of A/WSN/33 up to 2.7-fold at 24 hrs and 1.75-fold at 48 hrs compared to cells transfected with a non-targeting control siRNA. This study enriched the network formed between influenza viral proteins and host cellular proteins and laid a foundation for further studies of the regulation mechanisms of influenza virus life cycle.
出处
《中国预防兽医学报》
CAS
CSCD
北大核心
2017年第3期167-171,共5页
Chinese Journal of Preventive Veterinary Medicine
基金
国家自然科学基金(31402206
31672582)
