Influenza A viruses (IAVs) are aggressive pathogens that cause acute respiratory diseases and annual epidemics in humans. Host defense against IAV infection is initiated by macrophages, which are the principal effector cells of the innate immune system. We have previously shown that IAV infection of human macrophages is associated with robust secretion of proteins via conventional and unconventional protein release pathways. Here we have characterized unconventional, extracellular vesicle (EV)-mediated protein secretion in human macrophages during IAV infection using proteomics, bioinformatics, and functional studies. We demonstrate that at 9 h postinfection a robust EV-mediated protein secretion takes place. We identified 2359 human proteins from EVs of IAV-infected macrophages compared with 1448 proteins identified from EVs of control cells. Bioinformatic analysis shows that many proteins involved in translation, like components of spliceosome machinery and the ribosome, are secreted in EVs in response to IAV infection. Our data also shows that EVs derived from IAV-infected macrophages contain fatty acid-binding proteins, antiviral cytokines, copper metabolism Murr-1 domain proteins, and autophagy-related proteins. In addition, our data suggest that secretory autophagy plays a role in activating EV-mediated protein secretion during IAV infection.
Keywords: extracellular vesicles; influenza A virus; innate immunity; macrophages; proteomics.