Poster Presentation Lorne Infection and Immunity 2020

Entry strategies of respiratory viruses into cells and control of replication by IFITM antiviral molecules (#101)

Tina Meischel 1 , Fernando Villalon-Letelier 1 , Jeffrey Smith 1 , Andrew G Brooks 1 , Patrick C Reading 1 2 , Sarah L Londrigan 1
  1. Department of Microbiology and Immunology, University of Melbourne at the Peter Doherty Institute for Infection and Immunity, Melbourne, Victoria, Australia
  2. WHO Collaborating Centre for Reference and Research on Influenza, Victorian Infectious Diseases Reference Laboratory at the Peter Doherty Institute for Infection and Immunity, Melbourne, Victoria, Australia

Interferon-inducible transmembrane (IFITM) proteins, IFITM 1, 2 and 3, are known viral restriction factors. The antiviral role of IFITM3 against influenza A virus (IAV) is established, but the contribution of IFITM1 and IFITM2 in controlling IAV replication is less defined. Only limited studies have investigated restriction of other respiratory viruses, including paramyxoviruses, by IFITMs. Herein, we investigate the role of IFITM1/2/3 in their ability to restrict IAV and parainfluenza virus 3 (PIV3). A549 airway epithelial cells overexpressing IFITM 1, 2 and 3 were engineered to assess antiviral activity. We observed restriction of IAV infection by IFITM2 and IFITM3, whereas IFITM1 and IFITM3 potently blocked PIV3 infection. Using fluorescence microscopy, we showed that IFITM1 localizes to the plasma membrane while IFITM2 and IFITM3 are targeted to endocytic membranes in overexpressing A549 cells. This expression pattern corresponds to the localisation pattern reported for endogenous IFITMs. We hypothesised that the relative sensitivity of IAV and PIV3 to IFITM-mediated restriction may vary depending upon (i) the subcellular localisation of the IFITM molecule under investigation and (ii) the entry pathway used for infectious entry (i.e.: endocytosis vs direct fusion at the plasma membrane). Therefore, we used viral entry assays to confirm that IAV entered A549 cells through dynamin-dependent endocytosis, whereas PIV3 entry occurred independently of dynamin-mediated endocytosis. Collectively our data is consistent with IFITM1 restriction of PIV3 through inhibition of direct fusion with the plasma membrane in A549 cells. Furthermore, the antiviral activity of IFITM2 and IFITM3 during IAV endocytic entry is consistent with the reported mechanism of IFITM antiviral action. Further studies will investigate (i) the mechanism by which IFITM3 interferes with infectious entry of PIV3 into A549 cells and (ii) the relevance of IFITM1/2/3 in inhibition of other respiratory viruses.