Brooklyn Osborne, University of Alberta

Brooklyn Osborne (Department of Biological Sciences, University of Alberta), Mirzabek Kazbekov (Department of Biological Sciences, University of Alberta), Katharine Magor (Department of Biological Sciences, University of Alberta; Li Ka Shing Institute of Virology, University of Alberta)

Suppressing & Conquering

Non-structural protein 1 (NS1), a primary virulence factor of influenza A virus (IAV), has been shown to disrupt the RIG-I signalling pathway through RIPLET in a species-specific manner. Due to the continued circulation and zoonotic spillover of clade 2.3.4.4b in North America, the H5N1 IAV subtype remains a significant concern. Recently collected data revealed that H5Nx NS1 protein variants have varying degrees of signalling inhibition in humans and ducks, a reservoir host of influenza. It is unknown whether these effects of NS1 in humans are mediated through direct protein-protein binding with RIPLET or what amino acids sites play a critical role in this interaction in both species. To explore this interaction and determine the amino acid residues involved, wildtype and point mutation variants of NS1 proteins will be assessed for their capacity to inhibit interferon signalling, using luciferase assays, and their ability to bind to RIPLET, using Western blots and mass spectroscopy. This work will also explore the broader interactome of H5Nx NS1 proteins by employing methods which enable detection of protein interactions under native conditions, such as Virotrap or BioID. This work aims to define the residues necessary for the interaction between RIPLET and NS1, providing a potential target site for antiviral therapeutics. Additionally, exploration of the NS1 interactome will provide novel insights into host-pathogen interactions in relevant influenza hosts.