Soniya Agarwal, University of Ottawa

Authors: Soniya Agarwal and Marceline Côté; Affiliations: Department of Biochemistry, Microbiology and Immunology, Ottawa Institute of Systems Biology (OISB), Centre for Infection, Immunity and Inflammation (CI3), Faculty of Medicine, University of Ottawa, Ottawa, ON, Canada.

Breaking & Entering

Ebola virus (EBOV) is a highly pathogenic hemorrhagic fever virus that primarily targets macrophages and dendritic cells. These innate immune cells play a dual role during infection: enabling viral replication and triggering a pathogenic inflammatory response. Various attachment factors are displayed on their cell surfaces driving tropism and susceptibility. DC-SIGN is an attachment factor well known to enhance viral entry into macrophages. While this enhancement has been attributed to attachment, whether DC-SIGN further contributes to entry remains unknown. To confirm the necessity of the highly conserved N257 and N563 glycoprotein (GP) glycans hypothesized to interact with DC-SIGN, we infected DC-SIGN-expressing HeLa cells using EBOV-like particles harboring mutant GPs that eliminate site-specific glycosylation potential. Interestingly, while entry remained elevated with the single mutants, double mutant entry was unchanged. Given that the filovirus GPs have one or both glycan sites, we infected DC-SIGN-expressing HeLa cells with various filoviruses. We saw increased entry with all filoviruses, except Bombali and Tai Forest virus, confirming the conserved nature of this interaction. Therefore, we sought to uncover whether DC-SIGN can modulate macropinocytosis, which EBOV hijacks. Using 70kDa dextran-FITC, we observed more FITC cells among DC-SIGN-expressing cells, suggesting that DC-SIGN triggers macropinocytosis via signaling. As a screen of DC-SIGN-associated signaling modulators, we treated THP-1 macrophages with inhibitors of Syk, a kinase shown to co-precipitate with DC-SIGN, and observed a significant reduction in EBOV entry. Taken together, our study suggests that DC-SIGN functions beyond EBOV attachment, and may actively promote internalization by activating signaling cascades that support macropinocytosis.