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

Since silencing IFITM1 and IFITM3 could increase the susceptibility of DF-1 cell to ATMUV infection, we further investigated whether forced expression of IFITMs could inhibit ATMUV replication. Thus, we cloned chicken and duck IFITM1 and IFITM3, and their cDNA sequences were analyzed and deposited in the Genbank database. DF-1 cells were then transiently transfected with constructs expressing chicken or duck IFITM1 and IFITM3 or empty vector, respectively and challenged with ATMUV. Virus titers in cell culture supernatants were determined by TCID 50 assay at 24, 36, and 48 hpi. The forced expression of IFITMs in DF-1 cells was examined by Western blotting. As shown in Figure 6A , flag-tagged IFITM proteins were expressed well in DF-1 cells (Figure 6A) . We observed that overexpression of chicken or duck IFITM1 and IFITM3 obviously suppressed ATMUV replication in DF-1 cells, as indicated by lower TCID 50 titers in culture supernatants from the IFITM-overexpressing cells than those in control cells ( Figure 6B) . To further confirm this observation, the DF-1 cells were infected with ATMUV and harvested at 36 hpi, followed by qRT-PCR to examine the viral gene expression. As compared with control cells containing empty vector, overexpression of chicken or duck, IFITM1 or IFITM3 in DF-1 cells reduced mRNA expression of ATMUV envelope gene by average of 58, 43, 53, 62%, respectively ( Figure 6C) . These data suggest that avian IFITM1 or IFITM3 could strongly interfere with ATMUV infection.