Abstract
Interferon-induced transmembrane proteins (IFITMs) are novel viral restriction factors which inhibit numerous virus infections by impeding viral entry into target cells. To investigate the roles of IFITMs during fish virus infection, we cloned and characterized an IFITM1 homolog from orange spotted grouper (Epinephelus coioides) (EcIFITM1) in this study. EcIFITM1 encodes a 131-amino-acid polypeptide, which shares 64 and 43% identity with Seriola dumerili and Homo sapiens, respectively. The multiple sequence alignment showed that EcIFITM1 contained five domains, including NTD (aa 1–45), IMD (aa 46–67), CIL (aa 68–93), TMD (aa 94–119), and CTD (aa 120–131). In vitro, the level of EcIFITM1 mRNA expression was significantly up-regulated in response to Singapore grouper iridovirus (SGIV), or red-spotted grouper nervous necrosis virus (RGNNV) infection. EcIFITM1 encoded a cytoplasmic protein, which was partly colocalized with early endosomes, late endosomes, and lysosomes. The ectopic expression of EcIFITM1 significantly inhibited the replication of SGIV or RGNNV, which was demonstrated by the reduced virus production, as well as the levels of viral gene transcription and protein expression. In contrast, knockdown of EcIFITM1 using small interfering RNAs (siRNAs) promoted the replication of both viruses. Notably, EcIFITM1 exerted its antiviral activity in the step of viral entry into the host cells. Furthermore, the results of non-targeted lipometabolomics showed that EcIFITM1 overexpression induced lipid metabolism remodeling in vitro. All of the detected ceramides were significantly increased following EcIFITM1 overexpression, suggesting that EcIFITM1 may suppress SGIV entry by regulating the level of ceramide in the lysosomal system. In addition, EcIFITM1 overexpression positively regulated both interferon-related molecules and ceramide synthesis-related genes. Taken together, our results demonstrated that EcIFITM1 exerted a bi-functional role, including immune regulation and lipid metabolism in response to fish virus infections.
Highlights
Interferon-stimulated genes (ISGs) are induced by interferon (IFN) through a series of signal transduction cascades, and exert virous antiviral effects at specific stages of the virus life cycle [1,2,3]
The amino acid alignment analysis indicated that EcIFITM1 contained five domains: [1] a variable hydrophobic N-terminal domain (NTD); [2] a conserved hydrophobic intramembrane domain (IMD); [3] a conserved intracellular loop (CIL); [4] a variable hydrophobic transmembrane domain (TMD); and [5] a highly variable C-terminal domain (CTD) (Figure 1A)
Bioinformatic analysis showed that EcIFITM1 contained all the 5 conserved domains, including NTD, IMD, CIL, TMD, and CTD
Summary
Interferon-stimulated genes (ISGs) are induced by interferon (IFN) through a series of signal transduction cascades, and exert virous antiviral effects at specific stages of the virus life cycle (e.g., inhibiting viral entry, gene transcription, and protein synthesis, assembly, and release) [1,2,3]. IFN-induced transmembrane proteins (IFITMs) are one of the earliest identified ISG families, and play a crucial role in virus infection. Studies on the mechanism of their antiviral actions demonstrated that IFITMs either restrict virus entry by suppressing viral fusion with the endosomal or lysosomal membrane [9, 19,20,21,22], or inhibit viral infection by regulating viral protein expression or interacting with viral proteins [23, 24]. The function of other members still remains largely unknown [25, 26]
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