Oral Presentation 12th Australasian Virology Society Meeting 2024

Identification Of Novel Host Proteins That Are Associated With Macrophage Control of Influenza A Virus Replication. (#9)

Svenja Fritzlar 1 , Shirley Lin 1 , Nichollas Scott 1 , Clare V.L Oates 1 , Rebecca Johnson 1 , Rubaiyea Farrukee 1 , Andrew G Brooks 1 , Patrick C Reading 1 2 , Sarah L Londrigan 1
  1. Microbiology and Immunology, The University of Melbourne, at the Peter Doherty Institute for Infection and Immunity, Melbourne, Victoria, Australia
  2. WHO Collaborating Centre for Reference and Research on Influenza, Victorian Infectious Diseases Reference Laboratory, at the Peter Doherty Institute for Infection and Immunity, Melbourne, Victoria, Australia

Airway macrophages (MΦ) are an important component of the innate host-defence against influenza A virus (IAV). In contrast to airway epithelial cells, MΦ limit seasonal IAV infection through “abortive replication” such that infectious virus particles are not released from infected cells. Our ongoing research program aims to investigate the consequences of abortive IAV replication in MΦ in the context of contributing to effective innate host immunity. However, some highly pathogenic avian influenza virus (HPAI) strains replicate productively in MΦ, which has been mapped to the viral hemagglutinin (HA) protein. We used reverse genetics (RG) H5N1 viruses containing 6 genes from seasonal IAV plus HA/NA genes from HPAI H5N1 (but lacking the multibasic cleavage site (MBCS)) to identify strains that productively or abortively replicate in MΦ. We leveraged this key observation in an innovative proteomics approach to identify MΦ proteins that differentially bound to HA proteins associated with productive or abortive IAV replication. We expressed V5-tagged HA proteins associated with productive and abortive replication in MΦ and co-precipitated each HA and associated MΦ proteins. Mass spectrometry and proteomics analysis identified (i) proteins that bound both HA and (ii) proteins bound only HA associated with productive replication. We have now used in vitro protein overexpression and knockdown approaches, as well as CRISPR activation to modulate levels of endogenous host proteins, to validate the ability of these MΦ-specific host proteins to modulate the replication of influenza A virus. In addition, our ongoing studies also show that identification of macrophage-specific antiviral host responses that shut down virus replication can be redirected or enhanced in respiratory epithelium to induce antiviral activity as a novel “host-directed” antiviral strategy.