Abstract
Interferon inducible transmembrane proteins (IFITMs) are broad‐spectrum antiviral factors. In cell culture the entry of many enveloped viruses, including orthomyxo‐, flavi‐, and filoviruses, is inhibited by IFITMs, though the mechanism(s) involved remain unclear and may vary between viruses. We demonstrate that Sindbis and Semliki Forest virus (SFV), which both use endocytosis and acid‐induced membrane fusion in early endosomes to infect cells, are restricted by the early endosomal IFITM3. The late endosomal IFITM2 is less restrictive and the plasma membrane IFITM1 does not inhibit normal infection by either virus. IFITM3 inhibits release of the SFV capsid into the cytosol, without inhibiting binding, internalization, trafficking to endosomes or low pH‐induced conformational changes in the envelope glycoprotein. Infection by SFV fusion at the cell surface was inhibited by IFITM1, but was equally inhibited by IFITM3. Furthermore, an IFITM3 mutant (Y20A) that is localized to the plasma membrane inhibited infection by cell surface fusion more potently than IFITM1. Together, these results indicate that IFITMs, in particular IFITM3, can restrict alphavirus infection by inhibiting viral fusion with cellular membranes. That IFITM3 can restrict SFV infection by fusion at the cell surface equivalently to IFITM1 suggests that IFITM3 has greater antiviral potency against SFV.
Highlights
Human interferon inducible transmembrane proteins (IFITMs) are a family of five, 15–17 kDa membrane-associated proteins, of which three (IFITM1, 2 and 3) appear to function as broad-spectrum inhibitors of viral replication
We show that normal infection by both Semliki Forest virus (SFV) and Sindbis virus (SINV) is restricted by IFITM3 and, to a lesser extent, by IFITM2, but not by IFITM1
IFITMs can restrict alphavirus infection To investigate whether IFITMs can restrict infection by alphaviruses, we used A549 cells stably expressing human C-terminally HA-tagged IFITM1, 2 or 3 [19,20]
Summary
Human interferon inducible transmembrane proteins (IFITMs) are a family of five, 15–17 kDa membrane-associated proteins, of which three (IFITM1, 2 and 3) appear to function as broad-spectrum inhibitors of viral replication. Detailed studies are lacking for most viruses, work on influenza A virus (IAV) has suggested that IFITM3, in particular, inhibits viral entry by interfering with endosomal, low pH-induced fusion [1,2,3,4]. Alphaviruses, especially Semliki Forest virus (SFV) and Sindbis virus (SINV), have been used extensively to study viral entry into cells [8,9,10,11]. These small (∼75 nm diameter), positive sense, single-stranded RNA viruses were amongst the first to be shown to use clathrin-mediated endocytosis and endosomal low pH-dependent fusion to enter cells [8,10]. Release of the viral capsid protein into the cytosol is inhibited in IFITM3-expressing cells
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