title: Effective inhibition of mRNA accumulation and protein expression of H5N1 avian influenza virus NS1 gene in vitro by small interfering RNAs.
abstract: Avian influenza has emerged as a devastating disease and may cross species barrier and adapt to a new host, causing enormous economic loss and great public health threats, and non-structural protein 1 (NS1) is a multifunctional non-structural protein of avian influenza virus (AIV) that counters cellular antiviral activities and is a virulence factor. RNA interference (RNAi) provides a powerful promising approach to inhibit viral infection specifically. To explore the possibility of using RNAi as a strategy against AIV infection, after the fusion protein expression plasmids pNS1-enhanced green fluorescent protein (EGFP), which contain the EGFP reporter gene and AIV NS1 as silencing target, were constructed and NS1-EGFP fusion protein expressing HEK293 cell lines were established, four small interfering RNAs (siRNAs) targeting NS1 gene were designed, synthesized, and used to transfect the stable cell lines. Flow cytometry, real-time quantitative polymerase chain reaction, and Western blot were performed to assess the expression level of NS1. The results suggested that sequence-dependent specific siRNAs effectively inhibited mRNA accumulation and protein expression of AIV NS1 in vitro. These findings provide useful information for the development of RNAi-based prophylaxis and therapy for AIV infection.
authors.affiliation: College of Agriculture, Hainan Key Laboratory of Tropical Animal Reproduction & Breeding and Epidemic Disease Research, Animal Genetic Engineering Key Laboratory of Haikou, Hainan University, Haidian Island, Haikou, People's Republic of China.
authors.forename: Hanwei
authors.initials: H
authors.lastname: Jiao
authors.forename: Li
authors.initials: L
authors.lastname: Du
authors.forename: Yongchang
authors.initials: Y
authors.lastname: Hao
authors.forename: Ying
authors.initials: Y
authors.lastname: Cheng
authors.forename: Jing
authors.initials: J
authors.lastname: Luo
authors.forename: Wenhua
authors.initials: W
authors.lastname: Kuang
authors.forename: Donglin
authors.initials: D
authors.lastname: Zhang
authors.forename: Ming
authors.initials: M
authors.lastname: Lei
authors.forename: Xiaoxiao
authors.initials: X
authors.lastname: Jia
authors.forename: Xiaoru
authors.initials: X
authors.lastname: Zhang
authors.forename: Chao
authors.initials: C
authors.lastname: Qi
authors.forename: Hongxuan
authors.initials: H
authors.lastname: He
authors.forename: Fengyang
authors.initials: F
authors.lastname: Wang
mesh_terms: D019677:Artificial Gene Fusion; D015153:Blotting, Western; D002460:Cell Line; D005434:Flow Cytometry; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins / Q000032:analysis / Q000235:genetics; D006801:Humans; D053124:Influenza A Virus, H5N1 Subtype / Q000235:genetics / Q000254:growth & development*; D034741:RNA, Small Interfering; D060888:Real-Time Polymerase Chain Reaction; D011993:Recombinant Fusion Proteins / Q000032:analysis / Q000235:genetics; D017361:Viral Nonstructural Proteins / Q000037:antagonists & inhibitors* / Q000235:genetics; D037521:Virulence Factors / Q000037:antagonists & inhibitors* / Q000235:genetics
chemical_list: C085820:INS1 protein, influenza virus; D034741:RNA, Small Interfering; D011993:Recombinant Fusion Proteins; D017361:Viral Nonstructural Proteins; D037521:Virulence Factors; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins