The role of LC3-associated phagocytosis during virus infection

LC3-associated phagocytosis (LAP) is a recently-characterised phagocytic pathway that plays important roles during immunity to infectious agents including viruses. LAP shares many features with the well-characterised autophagy pathway but differs in signalling events upstream of LC3 recruitment to membranes. Thus, the precise role played by each pathway 'in vivo' has been very difficult to dissect. The unique aspect of this grant proposal is based on our generation of mice that are deficient in LAP, but have normal autophagy. This allows us to focus specifically on the role played by LAP in the control of viral infection. Our preliminary data using Influenza A virus (IAV) infection of LAP deficient mice reveal that there are higher virus titres, more inflammation and more clinical signs in the absence of LAP. We therefore believe that LAP plays an important role in host defence in the respiratory tract.
The aims of this project are to:
1. Discover the precise way that LAP acts in the defence mechanisms to IAV infection. This will involve comparing the progress of viral infection in genetically modified mice deficient in LAP with normal mice. We will also study whether LAP influences inflammatory responses, various types of immune responses (antibody and cytotoxic T cell) as well as other anti-viral responses such as interferon.
2. Discover whether LAP specifically in macrophages or epithelial cells plays a role in the defence against IAV. To do this, we generate conditional knockout mice that are lacking LAP in macrophages but not in all other tissues. These mice will be infected with IAV and then investigated as in Aim 1.

The results will significantly enhance our understanding of fundamental aspects of defence to virus infection as well as aspects of respiratory biology.
The research will be carried out at the Universities of Liverpool and UEA by a multi-disciplinary team comprising members of the Medical and Veterinary Faculties using well-equipped facilities currently situated at these sites. Defects in autophagy are associated with aberrant host defence, inflammatory disease and age-related disorders. It is possible that a defect in LAP, rather than autophagy, accounts for some of these pathological conditions. Characterisation LAP separate from autophagy, will allow examination of pathologies linked to autophagic machinery in a new light. In turn this may lead to novel approaches to improving antiviral therapies or therapeutic interventions against the consequences of viral infection

LC3-associated phagocytosis (LAP) is a recently characterised phagocytic pathway that plays important roles during innate defence and immunity to pathogens. LAP shares many components with the well-characterised autophagy pathway but differs in signalling events upstream of LC3 recruitment to membranes. Thus, the precise role played by each pathway 'in vivo' has been very difficult to dissect. The unique aspect of this grant proposal is based on our generation of mice that are deficient in LAP, but have normal autophagy. This allows us to focus specifically on the role played by LAP in the control of viral infection. Our preliminary data using Influenza A virus (IAV) infection of LAP deficient mice reveal that there are higher virus titres, more inflammation and more clinical signs in the absence of LAP. We therefore believe that LAP plays an important role in host defence in the respiratory tract.

We will use mice deficient in LAP, and mice where LAP is specifically removed from macrophages or epithelial cells. The transgenic models will be infected with IAV and the pathogenesis of infection analysed for virological, inflammatory and immunological parameters. IAV is relevant to disease in animals and has good sets of specific reagents that will allow the study of the host response in mice. This will allow us to determine the role played by LAP during an acute, cytopathic infection in the respiratory tract that is normally cleared rapidly generating immunity to re-infection.

The results will significantly enhance our understanding of fundamental aspects of host responses to virus infection in the lung as well as respiratory biology. In turn this may lead to novel approaches to improving antiviral therapies or therapeutic interventions against the consequences of viral infection.

Respiratory virus infections are of global significance to both the human and animal populations. The zoonotic potential of influenza and the implications of emerging new strains are well recognised by both the scientific community and the general public. Improved understanding of the pathogenesis of infection and the host response to these pathogens is critical in improving treatment and management of respiratory disease and the associated morbidity and mortality. This work aims to further this understanding by looking at the role of LC3-associate phagocytosis (LAP) in the innate and adaptive responses of the host respiratory tract following infection with influenza virus.

The academic impact of this work therefore will be to further the knowledge of the host response to viral infection, providing the foundations of academic knowledge and understanding on which future advancements in treatment and disease control can be built. The use of the multi-disciplinary approach as proposed here brings together expertise in cell biology, molecular virology, immunology and veterinary pathology, thereby maximising the potential outputs of the research. This is also advantageous to the RA in providing the experience and training involving multiple fields fostering a multi-disciplinary approach and its advantages for enhanced, productive science as well as allowing them to develop a range of transferable skills that will enhance their career development. This is achieved by collaboration between the Universities of East Anglia and Liverpool.

The societal impact of furthering the understanding of the host response to infection with Influenza virus on the health of the individual and the wider population is significant. Contribution to the improved treatment of individual high risk patients where infection with Influenza virus has a higher morbidity and mortality than the general population, or increased understanding of the risk factors associated with epidemic strains, both are important in the future of prevention and control of Influenza infection, in both veterinary species and the human population.