Characterising the determinants of influenza A virus vRNP trafficking and packaging in swine

According to the WHO influenza A viruses (IAVs) are responsible for a billion cases of seasonal influenza annually, and with the continual concern for the next pandemic IAV, there is a perpetual need for further research to understand how these viruses replicate and manipulate the host cell environment.  IAVs have a remarkable ability to infect a wide range of host species, with pigs representing a species with a high risk of generating a novel IAV capable of pandemic disease.  This was indeed the case with the 2009 H1N1 pandemic resulting from the reassortment of swine, human and avian IAVs within a swine host, generating a novel strain.  With the mechanisms driving IAV reassortment remaining poorly understood, I predict that as-yet unidentified host cell factors play a role in the trafficking and packaging of newly produced viral ribonucleoproteins (vRNPs) during egress from the host cell, the stage of the life cycle when reassortment is presumed to take place.  Understanding fundamental aspects of vRNP regulation in pigs, which likely affect reassortment dynamics, may enable the development of broadly acting antivirals against these important host cell factors. Our previous work has identified novel host proteins that interact with vRNPs during intracellular trafficking in human cells.  We found that myoferlin (MYOF), a transmembrane protein involved in membrane dynamics and receptor recycling, co-localises with vRNPs in Rab11a-positive trafficking vesicles.  The remodelling of Rab11a vesicles by IAVs is well-described by others, and we have recently demonstrated that MYOF is an essential co-factor for this process.  However, I wish to move beyond this work and investigate the host species differences in vRNP regulation as little is currently known on the mechanisms of how vRNPs are trafficked and packaged into virions in porcine cells.  With the pig being an ideal 'mixing vessel' for human adapted IAVs, a better understanding of these essential processes in viral replication is critical as we aim to predict and avert the next influenza pandemic. During this project I will expand the understanding of IAV replication in the context of porcine cells, with three research aims: Determine the IAV vRNP-interacting host factors in porcine cells.  Validate the pro-viral functions of identified swine factors in the context of IAV vRNP trafficking.  Verify whether pathways utilised for vRNP egress are conserved between porcine and human cells.  Upon completion of this project, we will have a better understanding of the trafficking and packaging of vRNPs in porcine cells, identifying which host cell factors are key players at this stage of viral replication and budding from cells.  Identifying host co-factors that are conserved between porcine and human cells, and how IAV proteins interact with them will aid our understanding of how swine viruses need to mutate and evolve to cross species barriers and infect human cells.  Additionally, we will gain a clearer understanding of how the eight IAV gene segments are packaged together in newly formed virions and what stages of this process can be targeted with novel antivirals to block IAV replication and transmission.