Uncovering the innate immune signatures and molecular processes driving influenza A virus hemagglutinin glycosylation-mediated immune pathology

Despite extensive research efforts, Influenza A virus (IAV) is still a major cause for animal and human mortality because of the absence of broadly neutralizing vaccines and viral resistance against approved antivirals. The envelope-glycoprotein hemagglutinin (HA) mediates entry into the host cell and is the primary determinant of antigenicity, virulence, and zoonotic potential. In response to the host immune reaction, it undergoes rapid adaptive mutational changes. Amino acid changes in the globular head domain of HA can lead to addition of glycosylation motifs that protect HAs antigenic sites from neutralizing antibodies and allow seasonal circulating H1 and H3 subtypes to partially escape the preexisting host antibody immune response. These glycosylation patterns were also shown to greatly influence the quality and breadth of the host antibody response and are therefore an important factor for vaccine design. Moreover, several studies suggest that glycans in the globular head domain also influence the pathogenicity of IAV strains. Severe disease during IAV infection is characterized by a hyper-induction of proinflammatory cyto- and chemokines and a massive infiltration of innate immune cells into the lung. However, so far it remains unclear how glycans in the globular head domain of HA influence the innate immune system and therefore pathogenicity. My preliminary data indicate that there is a glycosylation-dependent decrease of proinflammatory cyto- and chemokine secretion by primary human immune cells when infected ex vivo. Based on these findings, I propose to uncover the innate immune signatures and molecular processes driving influenza A virus hemagglutinin glycosylation-mediated immune pathology during my postdoctoral studies in the nameable influenza and innate immunity research laboratory of Prof. Ana Fernandez-Sesma at the Icahn School of Medicine at Mount Sinai (ISMMS), New York. Firstly, I aim to reveal the innate immune profile underlying HA glycosylation-mediated pathogenesis. I will do so by measuring the secretion of proinflammatory cyto- and chemokines and the surface expression of activation markers of primary human innate immune cells after ex vivo exposure to glycosylation mutant versions of pandemic H1N1 and H3N2 viruses, whose virulence has been previously assessed in vivo. These viruses will be provided by the Center for Research on Influenza Pathogenesis and Transmissibility (CRIPT), an international influenza virus surveillance network in which the laboratory of Prof. Ana Fernandez-Sesma plays an essential role. Secondly, I will generate an influenza virus like particle reporter system to identify HA glycosylation patterns in circulating avian IAV with the highest zoonotic potential, namely H5 and H7 viruses, that can induce this type of severe disease in humans. The result of this aim will refine pandemic preparedness and surveillance strategies. Lastly, I will investigate the signaling events responsible for the innate immune pathology mediated by HA glycosylation. To do so, I will benefit from the expertise in the laboratory of Prof. Ana Fernandez-Sesma in cytometry by time-of-flight (CyTOF) and combine it with my extensive knowledge in phosphorylation-mediated signaling. The obtained results will refine pandemic preparedness and broaden our understanding of the underlying molecular mechanism causing severe disease during IAV infection necessary to develop effective treatments.