title: Gene expression profile after activation of RIG-I in 5'ppp-dsRNA challenged DF1.
abstract: Retinoic acid inducible gene I (RIG-I) can recognize influenza viruses and evoke the innate immune response. RIG-I is absent in the chicken genome, but is conserved in the genome of ducks. Lack of RIG-I renders chickens more susceptible to avian influenza infection, and the clinical symptoms are more prominent than in other poultry. It is unknown whether introduction of duck RIG-I into chicken cells can establish the immunity as is seen in ducks and the role of RIG-I in established immunity is unknown. In this study, a chicken cell strain with stable expression of duRIG-I was established by lentiviral infection, giving DF1/LV5-RIG-I, and a control strain DF1/LV5 was established in parallel. To verify stable, high level expression of duRIG-I in DF1 cells, the levels of duRIG-I mRNA and protein were determined by real-time RT-PCR and Western blot, respectively. Further, 5'triphosphate double stranded RNA (5'ppp-dsRNA) was used to mimic an RNA virus infection and the infected DF1/LV5-RIG-I and DF1/LV5 cells were subjected to high-throughput RNA-sequencing, which yielded 193.46 M reads and 39.07 G bases. A total of 278 differentially expressed genes (DEGs), i.e., duRIG-I-mediated responsive genes, were identified by RNA-seq. Among the 278 genes, 120 DEGs are annotated in the KEGG database, and the most reliable KEGG pathways are likely to be the signaling pathways of RIG-I like receptors. Functional analysis by Gene ontology (GO) indicates that the functions of these DEGs are primarily related to Type I interferon (IFN) signaling, IFN-β-mediated cellular responses and up-regulation of the RIG-I signaling pathway. Based on the shared genes among different pathways, a network representing crosstalk between RIG-I and other signaling pathways was constructed using Cytoscape software. The network suggests that RIG-mediated pathway may crosstalk with the Jak-STAT signaling pathway, Toll-like receptor signaling pathway, Wnt signaling pathway, ubiquitin-mediated proteolysis and MAPK signaling pathway during the transduction of antiviral signals. After screening, a group of key responsive genes in RIG-I-mediated signaling pathways, such as ISG12-2, Mx1, IFIT5, TRIM25, USP18, STAT1, STAT2, IRF1, IRF7 and IRF8, were tested for differential expression by real-time RT-PCR. In summary, by combining our results and the current literature, we propose a RIG-I-mediated signaling network in chickens.
authors.affiliation: The Key Laboratory of Animal Genetics and Breeding and Molecular Design of Jiangsu Province, Yangzhou University, Yangzhou 225009, PR China.
authors.forename: Yang
authors.initials: Y
authors.lastname: Chen
authors.affiliation: The Key Laboratory of Animal Genetics and Breeding and Molecular Design of Jiangsu Province, Yangzhou University, Yangzhou 225009, PR China.
authors.forename: Qi
authors.initials: Q
authors.lastname: Xu
authors.affiliation: The Key Laboratory of Animal Genetics and Breeding and Molecular Design of Jiangsu Province, Yangzhou University, Yangzhou 225009, PR China.
authors.forename: Yang
authors.initials: Y
authors.lastname: Li
authors.affiliation: The Key Laboratory of Animal Genetics and Breeding and Molecular Design of Jiangsu Province, Yangzhou University, Yangzhou 225009, PR China.
authors.forename: Ran
authors.initials: R
authors.lastname: Liu
authors.affiliation: The Key Laboratory of Animal Genetics and Breeding and Molecular Design of Jiangsu Province, Yangzhou University, Yangzhou 225009, PR China.
authors.forename: Zhengyang
authors.initials: Z
authors.lastname: Huang
authors.affiliation: The Key Laboratory of Animal Genetics and Breeding and Molecular Design of Jiangsu Province, Yangzhou University, Yangzhou 225009, PR China.
authors.forename: Bin
authors.initials: B
authors.lastname: Wang
authors.affiliation: The Key Laboratory of Animal Genetics and Breeding and Molecular Design of Jiangsu Province, Yangzhou University, Yangzhou 225009, PR China. Electronic address: ghchen@yzu.edu.cn.
authors.forename: Guohong
authors.initials: G
authors.lastname: Chen
mesh_terms: D000818:Animals; D005075:Biological Evolution; D002460:Cell Line; D002645:Chickens / Q000276:immunology; D000071457:DEAD Box Protein 58 / Q000235:genetics*; D004372:Ducks / Q000276:immunology*; D005786:Gene Expression Regulation; D063990:Gene Ontology; D059014:High-Throughput Nucleotide Sequencing; D007113:Immunity, Innate; D005585:Influenza in Birds / Q000276:immunology*; D007370:Interferon Type I / Q000235:genetics*; D012333:RNA, Messenger / Q000235:genetics; D012367:RNA, Viral / Q000276:immunology; D020239:Receptor Cross-Talk; D015398:Signal Transduction / Q000235:genetics; D059467:Transcriptome
chemical_list: C551867:5'PPP-RNA; D007370:Interferon Type I; D012333:RNA, Messenger; D012367:RNA, Viral; D000071457:DEAD Box Protein 58
keywords: Crosstalk; Duck; Expression profile; RIG-I; RNA-Seq