title: Single-chain intracellular antibodies inhibit influenza virus replication by disrupting interaction of proteins involved in viral replication and transcription.
abstract: Influenza A virus is responsible for influenza epidemics in avian and human populations and poses a great threat to human health. Many researches have been focused on the prevention and treatment of influenza A virus infection. The nucleoprotein (NP) of the virus is an important protein due to its ability to interact with a variety of viral and cellular factors and its role in the viral RNA synthesis. In this study, we have used the influenza A virus nucleoprotein as target for anti-viral therapy through a new approach. By screening a human single-chain intracellular antibody (intrabody) cDNA library using nucleoprotein as bait in a yeast antigen-antibody two-hybrid system, we have identified several intrabodies that specifically interact with the viral nucleoprotein. Interaction between nucleoprotein and anti-nucleoprotein intrabodies was further confirmed by mammalian two-hybrid analysis. Results showed that anti-nucleoprotein intrabodies changed their cellular distribution in association with the viral nucleoprotein. Further studies indicated that anti-nucleoprotein intrabodies abolished the accumulation of not only the complementary RNA and virion RNA but also messenger RNA of influenza virus. In addition, anti-nucleoprotein intrabodies significantly inhibited influenza A virus transcription and replication through blocking the interaction of nucleoprotein with the viral polymerase proteins, polymerase basic protein 1 and polymerase basic protein 2. Thus, this study not only provides a powerful tool for the study of viral protein's functions, but also opens a new potential avenue for the prevention and treatment of influenza virus infections.
authors.affiliation: State Key Laboratory of Virology, College of Life Sciences, Wuhan University, Wuhan 430072, PR China.
authors.forename: Muhammad Mahmood
authors.initials: MM
authors.lastname: Mukhtar
authors.forename: Shengfeng
authors.initials: S
authors.lastname: Li
authors.forename: Wei
authors.initials: W
authors.lastname: Li
authors.forename: Ting
authors.initials: T
authors.lastname: Wan
authors.forename: Yongxin
authors.initials: Y
authors.lastname: Mu
authors.forename: Wei
authors.initials: W
authors.lastname: Wei
authors.forename: Lei
authors.initials: L
authors.lastname: Kang
authors.forename: Sahibzada T
authors.initials: ST
authors.lastname: Rasool
authors.forename: Yibei
authors.initials: Y
authors.lastname: Xiao
authors.forename: Ying
authors.initials: Y
authors.lastname: Zhu
authors.forename: Jianguo
authors.initials: J
authors.lastname: Wu
mesh_terms: D000914:Antibodies, Viral / Q000276:immunology / Q000378:metabolism*; D000956:Antigens, Viral / Q000276:immunology; D015723:Gene Library; D006801:Humans; D009980:Influenza A virus / Q000235:genetics / Q000254:growth & development / Q000276:immunology*; D007251:Influenza, Human / Q000276:immunology* / Q000517:prevention & control; D009698:Nucleoproteins / Q000276:immunology; D011485:Protein Binding / Q000276:immunology; D021381:Protein Transport / Q000276:immunology; D012367:RNA, Viral / Q000032:analysis; D012324:RNA-Dependent RNA Polymerase / Q000037:antagonists & inhibitors; D014158:Transcription, Genetic; D020798:Two-Hybrid System Techniques; D014764:Viral Proteins / Q000037:antagonists & inhibitors; D014779:Virus Replication / Q000276:immunology*
chemical_list: D000914:Antibodies, Viral; D000956:Antigens, Viral; D009698:Nucleoproteins; C057027:PB2 protein, Influenzavirus A; D012367:RNA, Viral; D014764:Viral Proteins; C077346:influenza virus polymerase basic protein 1; D012324:RNA-Dependent RNA Polymerase