Interferon-induced transmembrane protein 3 blocks fusion of sensitive but not resistant viruses by partitioning into virus-carrying endosomes

Suddala, Krishna C.; Lee, Christine C.; Meraner, Paul; Marin, Mariana; Markosyan, Ruben M.; Desai, Tanay M.; Cohen, Fredric S.; Brass, Abraham L.; Melikyan, Gregory B.

We have previously shown that IFITM3 does not restrict the lipid-mixing (hemifusion) stage of viral fusion, but rather inhibits the formation of a fusion pore [30] . However, the inability to directly visualize IFITM3 in the context of virus entry into live cells precluded us from assessing whether this factor blocks fusion through a proximity-based mechanism. Here, we overcame this limitation by constructing a functional fluorescent IFITM3 protein and imaging virus co-trafficking and fusion with endosomal compartments in cells expressing this protein. Comparison of entry and fusion of IFITM3-sensitive (IAV) and-resistant (LASV) viruses by 3-color live cell imaging revealed that IAV enters into and remains trapped within endosomes enriched in fluorescent IFITM3, where viruses underwent hemifusion but failed to complete the fusion reaction. In contrast, LASV particles entered and fused with endosomes devoid of IFITM3, implying that LASV escapes restriction by utilizing an endocytic pathway distinct from that employed by IAV. Collectively, our results provide strong support to a proximity model by which presence of IFITM3 at the preferred sites of virus entry restricts viral fusion.