Avian Interferon-Inducible Transmembrane Protein Family Effectively Restricts Avian Tembusu Virus Infection

Chen, Shilong; Wang, Long; Chen, Jieying; Zhang, Lanlan; Wang, Song; Goraya, Mohsan U.; Chi, Xiaojuan; Na, Yang; Shao, Wenhan; Yang, Zhou; Zeng, Xiancheng; Chen, Shaoying; Chen, Ji-Long

ATMUV is a newly emerging member of the Ntaya virus group within the genus of Flavivirus, causing severe egg-drop syndrome and neurological disease in domestic poultry Homonnay et al., 2014; Thontiravong et al., 2015) . On the basis of typical clinical symptoms, ATMUV was initially described as duck egg drop syndrome virus (DEDSV). However, this virus exhibits a wide range of pathogenicity to avian species including ducks, chicken, geese, house sparrows, and racing pigeon, and its genome is closely related to Tembusu virus strains. Flavivirus isolates with a more than 84% nucleotide sequence homology in the NS5 region are considered to be the same species (Kuno et al., 1998) . So we propose to name this novel flavivirus as "Avian Temubsu virus, ATMUV." Till now, there is no effective method for its prevention except vaccine. The previous studies have shown that ATMUV infection induced an effective antiviral immunity through MDA5 and TLR3-dependent signaling pathways Chen et al., 2016a; Fu et al., 2016) . However, our understanding of the molecular and cellular basis of interaction between the virus and host antiviral immune system is very limited. In this study, we examined the expression profile of key IFNs and IFITMs following ATMUV infection in vivo and in vitro. We found type I, type III IFNs, and immune-related IFITMs, including The viral titers in cell culture supernatants were determined by TCID 50 assay in DEF cells. (C) The mRNA expression of viral genes was examined by qRT-PCR at 36 hpi. The mRNA levels were normalized to the endogenous β-actin level and the expression in ATMUV infected DF-1 cells expressing empty vector was set to 1.0. The mRNA expression of viral genes in DF-1 cells expressing exogenous IFITMs was compared to that in control cells containing empty vector. Plotted are the average results from three independent experiments (means ± SD). Statistical significance was determined by one tail Student's t-test analysis. *P < 0.05, **P < 0.01. IFITM1, IFITM2, and IFITM3 were significantly upregulated in response to ATMUV infection. Strikingly, the spleen tissue had a high viral load and showed strong innate immune response as compared to other organs. IFN-α/β expression levels in spleen were increased significantly at 24 hpi and decreased after this time point, and the mRNA levels of IFITM1 and IFITM2 were also upregulated. Similar results were shown in kidney and bursa of fabricius tissues in infected animals. Although IFN expression in DEFs was triggered and reached the maximum value at 48 hpi, DEFs displayed an apparent CPE by ATMUV infection at this time point. These data suggest that host innate immune response is triggered by ATMUV, but the virus has developed multiple strategies to evade host antiviral immunity. In a recent study, Wang and his co-workers found that ATMUV NS1 could markedly suppress virus-induced IFN-β expression by inhibiting RLR receptor signaling . Most flaviviruses are potent in blocking the JAK-STAT pathway to evade the antiviral effects of the host innate immune system (Heim et al., 1999; Basu et al., 2001; Lin et al., 2004; Green et al., 2014) . Thus, further studies are needed to address the precise mechanisms by which ATMUV circumvents the host innate immune response.