Identification, Tissue Expression And Immunological Characteristics Of Goose Type III Interferon And Its Receptor

Interferons (IFNs), as the first line of defense against the viral infection, belong to class II cytokines, play an important role in innate immune response. Now, according to the amino acid sequences, structure characteristics and their receptor characteristics, etc. Interferons were divided into three types, they are type ? interferon, type ? interferon and type ? interferon, respectively. Type ? interferons (IFN-?s) was found and named in 2003.The antiviral effect of IFN-?s is not directly kill the virus, but it binds to a heterodimeric cell surface receptors composed of two chains:IFNLR1 and IL-10R?. Activated the JAK-STAT signaling pathway which similar to type I interferon, result in directly hindering the viral replication in host cells.In this study, the goIFNL and goIFNLR1 cDNA from Sichuan white goose were firstly identified, and a related bioinformatics analysis were carried out. The full-length sequence of goIFNL was 823 bp, contain a complete ORF domain, encoding 189 amino acids (aa). The full-length sequence of goIFNLR1 was 1952 bp, contain a complete ORF domain, encoding 512 aa. Though NJ phylogenetic tree analyzed, shown that the two gene belong to birds branch has a closest relatives with poultry.To figure out the tissue distribution condition of goIFNL and goIFNLR1, quantitate RT-PCR was performed. We found that the mRNA expression level of goIFNL and goIFNLR1 was primary expressed in epithelial related tissues derived from respiratory tract tissue such as lung (LU), trachea (TR); gastrointestinal tissues such as small intestine (SI), gizzard (GI), skin (SK), pancreas (P) and Blood.To have a better understand of the immunological characteristics of goIFNL, several kinds of TLR agonists (Poly (I:C), ODN2006, LPS and R848) and poultry virus (GPV, H9N2 AIV and DTMUV) were used to treat goose PBMCs in this study, for set up the infection cell model in vitro. The result shown that the mRNA transcription level of goIFNL has a significant rise under the stimulate of agonists Poly (I:C) and ODN2006, which were TLR3 and TLR9 ligand, respectively. While the mRNA transcription level of goIFNL and goIFNLR1 compared with the control group has no significant difference under the stimulus of LPS and R848, which were TLR4 and TLR7 ligand, respectively. The mRNA transcription level of goIFNL also has a significant rise after infected with GPV, and there were also no significant difference of the mRNA transcription level of goIFNL and goIFNLR1 when infected with H9N2 AIV and DTMUV. At the same time, gosling were artificial infected with poultry virus (GPV, H9N2 AIV and DTMUV) in vivo. We found after the H9N2 AIV infection, the mRNA transcription level of goIFNL compared with the control group has a nearly 20 times rise in LU tissue and has a nearly 40 times rise in P tissue. It is suggested goIFNL plays a certain role in the antiviral innate immune of Chinese goose.In order to further understand the biological function of goIFNL, the recombinant pET-32a-goIFNL prokaryotic expression plasmid and pcDNA3.1-goIFNL eukaryotic expression plasmid were constracted to prokaryotic and eukaryotic expression of the goIFNL protein. The recombinant prokaryotic expression plasmid was successful expressed in E.col BL21 (DE3) cells induced by IPTG with a final concentration at 0.8 mM for 3h, the fusion protein expressed in a form of inclusions and purified by using Ni-NTA Agarose. The goIFNL fusion protein was used as the immunogen to prepare anit-goIFNL mouse polyclonal antibody. The recombinant pcDNA3.1-goIFNL eukaryotic expression plasmid also successful expressed in BHK21 cells, it's laid the foundation for the research of goIFNL antivial immunological function.In conclusion, this study has successful cloned goIFNL and goIFNLR1 genes, has a preliminary study for its tissue distribution in Chinese goose and the immunological characteristics, the successful prokaryotic and eukaryotic expression of the goIFNL protein lay a foundation for the subsequent biological activity research.