Uncovering the non-canonical MHC class II receptor-usage of a newly discovered influenza A virus subtype in wild diving ducks

Influenza A virus (IAV) is a seasonally recurring respiratory pathogen that causes epidemics in the human population with significant burden to health and economy. Despite having a considerably broad host range including humans, the main reservoir of IAV resides in wild waterfowl and shore birds. For this reason, zoonotic IAVs as emerging pathogens carry the potential to cause devastating pandemics.IAVs are classified by their immunogenicity of their surface glycoproteins hemagglutinin (HA) and neuraminidase (NA). Until today, there are 18 and 11 different HA and NA subtypes described, re-spectively. The HA, as a key determinant of the IAV host range, engages the host cell receptor and facilitates cell entry and infection, and is therefore a major focus of research. The most recently dis-covered IAV subtypes H17N10 and H18N11 in bats have not only expanded the IAV host range but also unveiled a novel mode of the IAV receptor-usage that is independent of the canonical IAV recep-tor sialic acid (SA), a sugar modification on host cell glycoproteins and glycolipids. During my PhD studies, I have discovered that bat IAVs exploit the major histocompatibility complex (MHC) class II from multiple species including humans, chicken, bats and pigs, as entry receptors. This finding has major implications for the host range of bat IAV and more importantly has revealed MHC class II as a non-canonical IAV receptor.The discovery of a novel IAV subtype in wild diving ducks in Northern California, provisionally termed HXNX, with properties reminiscent of bat IAV raised the question whether HXNX enters its host cells through MHC class II as well. My preliminary experiments support this hypothesis and indicate that HXNX does indeed not bind SA, but instead uses MHC class II as host cell receptor. These findings suggest a more widespread role for MHC class II as an IAV host cell receptor than initially thought.Based on these recent findings, I propose to uncover the non-canonical MHC class II receptor-usage of the novel influenza A virus subtype HXNX during my postdoctoral studies in the renowned influenza research laboratory of Prof. Adolfo García-Sastre at the Icahn School of Medicine at Mount Sinai (ISMMS), New York. First, I will examine the host range of HXNX by establishing entry assays using HX-pseudotyped virus-like particles (VLP) and test on cells expressing MHC class II from po-tential HXNX hosts, including different duck and bat species. Furthermore, I will work in collaboration with a global IAV surveillance center in the U.S. to determine the distribution of HXNX in duck and bat populations in Northern California. Second, I will generate and test chimeric MHC class II proteins of species that represent functional or non-functional HXNX receptors, which will enable the identification of the receptor-binding site of HX on MHC class II. Finally, I will establish the reverse genetics tools to rescue the recombinant HXNX virus, thereby generating the infectious HXNX virus for the very first time, which is essential to further investigate the biology of HXNX, including its pathogenicity and tissue tropism in vivo.Covering significant areas of influenza virus research, the proposed project aims at comprehensively studying a novel IAV subtype by assessing its host cell receptor-usage, a key determinant of the virus's host range, pathogenesis and tissue tropism. Moreover, the obtained results will underscore the role of MHC class II as a genuine IAV receptor used by multiple IAV subtypes, and thus will pro-vide profound insights into IAV biology.