IFITM3 Clusters on Virus Containing Endosomes and Lysosomes Early in the Influenza A Infection of Human Airway Epithelial Cells

Kummer, Susann; Avinoam, Ori; Kräusslich, Hans-Georg

A semi-automated image analysis of IAV A/HK/1/68 (pdmH3N2) and IAV A/R/D6/2009 (pdmH1N1) infected A549 cells revealed IFITM3 clustering early after infection in both cases, with few differences in kinetics, which may reflect strain specific replication kinetics [73] . The CD225 domain of IFITM3, which is composed of the intermembrane domain 1 and the intracellular loop, contains two phenylalanine residues mediating the physical association between IFITMs; these residues are strongly connected with the antiviral function [39] . Combining this finding with our observation of the increasing levels of larger IFITM3 clusters, we suggest a direct relation between cluster formation and IFITM3's antiviral activity on endosomal vesicles. The IFITM3 clustering on apparently vesicular structures started early after infection (3-5 h p.i.), with an initial co-localization of IFITM3 with an early endosomal marker and a later co-localization with a late endosomal marker, reflecting the early to late endosomal pathway of IAV. It appears likely, therefore, that IFITM3 clusters, initially recruited to early endosomes and then coating endosomal vesicles through their trafficking pathway, may mediate the antiviral activity of IFITM3 and block the release of vRNPs. This hypothesis is further supported by the identification of a critical sorting signal that is essential for the IFITM3 localization to endosomes and its anti-viral activity [74, 75] . Similar to IAV, arena-(including LASV, lymphocytic choriomeningitis virus, and MACV) and alphaviruses (including the chikungunya virus, Sindbis virus, and Venezuelan encephalitis virus) also fuse from late endosomes and lysosomes at a similar acidic pH, but are not restricted by IFITM3 [51] . It will be of interest, therefore, whether a similar IFITM3 clustering can be observed on endosomal vesicles carrying these viruses or whether they can escape from IFITM3 clustering.