IFITM 3 requires an amphipathic helix for antiviral activity

Abstract

Interferon-induced transmembrane protein 3 (IFITM3) is a cellular factor that blocks virus fusion with cell membranes. IFITM3 has been suggested to alter membrane curvature and fluidity, though its exact mechanism of action is unclear. Using a bioinformatic approach, we predict IFITM3 secondary structures and identify a highly conserved, short amphipathic helix within a hydrophobic region of IFITM3 previously thought to be a transmembrane domain. Consistent with the known ability of amphipathic helices to alter membrane properties, we show that this helix and its amphipathicity are required for the IFITM3-dependent inhibition of influenza virus, Zika virus, vesicular stomatitis virus, Ebola virus, and human immunodeficiency virus infections. The homologous amphipathic helix within IFITM1 is also required for the inhibition of infection, indicating that IFITM proteins possess a conserved mechanism of antiviral action. We further demonstrate that the amphipathic helix of IFITM3 is required to block influenza virus hemagglutinin-mediated membrane fusion. Overall, our results provide evidence that IFITM proteins utilize an amphipathic helix for inhibiting virus fusion.

Functional Mapping of Regions Involved in the Negative Imprinting of Virion Particle Infectivity and in Target Cell Protection by Interferon-Induced Transmembrane Protein 3 against HIV-1

“…A number of studies have revealed domains that have important regulatory functions on the antiviral properties of the different IFITM members, influencing their ability to recruit multiple cellular partners and to mediate protection of target cells from viral challenge (19,33,43,(48)(49)(50)(51)(52)(53)(54)(55). However, little is known about domains that specify the negative imprinting properties of IFITMs.…”

Construction, expression and antiviral activity analysis of recombinant adenovirus expressing human IFITM3 in vitro

“…IFITM3 blocks virus fusion with cell membranes via acidic endosome [38,44]. According to the previous reports, the IFITM3 gene is conserved in chimpanzee, Rhesus monkey, mouse, and rat, and 14 organisms have orthologs with human IFITM3 gene (https://www.ncbi.nlm.nih.gov/ gene/10410), indicating a cross protection between species was exist.…”

“…IFITM3 is a virus restriction factor which primarily localizes to endosomes and lysosomes [38]. Thus, a suitable vector or strategy is needed to transfer the protein into the target cell.…”

IFITMs Inhibit Cell Fusion Mediated by Trophoblast Syncytins

“…In order to determine whether IFITMs are specifically capable of inhibiting fusion mediated individually by Syncytin-1 or -2, we employed a cell-to-cell fusion assay utilizing HEK293T cells, which naturally lack expression of endogenous fusion proteins and endogenous IFITMs, and that we previously validated for studying effects of IFITMs on cell-to-cell fusion mediated by viral fusion proteins (12,17,33). In short, one population of cells was transfected with vector control or Syncytin-1 or -2, while target cells were transfected with individual IFITM expression plasmids or vector control.…”

“…IFITM constructs cloned into the pCMV-HA vector (Clontech) were described previously (13,31) and myc-FLAG-tagged Syncytin-1 and 2 expression constructs in the pCMV6 vector were purchased from Origene. pFR-Luc and pBD-NFkB (Agilent) were co-transfected with plasmids as indicated in Fig 3A. Cell-to-cell fusion assays were performed as outlined and validated previously (12,17,34). In short, transfections were performed overnight using LipoJet transfection reagent (Signagen).…”