The role of LC3-associated phagocytosis during virus infection

Lead Research Organisation: University of Liverpool
Department Name: Institute of Infection and Global Health

Abstract

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

Technical Summary

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.

Planned Impact

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.
 
Description Non-canonical autophagy is a cellular process that this conventionally thought of as being involved in the function of phagocytic cells and how they are able to mediate defence against invading pathogens. We have shown that during influenza infection non-canonical autophagy has an important role to play in the pathogenesis of influenza disease and that this process is important in the epithelial barrier not phagocytic cells.
Exploitation Route Modulation of non-canonical autophagy by drugs might be a means of improving outcomes in influenza (and other respiratory virus) infections
Sectors Agriculture, Food and Drink,Pharmaceuticals and Medical Biotechnology

 
Description Digard 
Organisation University of Edinburgh
Country United Kingdom 
Sector Academic/University 
PI Contribution Infection model of 3D tracheal epithelial cell cultures derived from wild type and BPIFA1 KO mice
Collaborator Contribution Reagents and expertise in terms of labelled IAV and highly concentrated preps of IAV
Impact Determination that BPIFA1 is involved in binding and entry of IAV into epithelial cells
Start Year 2015
 
Description SPLUNC1 Tripp 
Organisation University of Georgia
Country United States 
Sector Academic/University 
PI Contribution The Stewart group are investigating the host response to virus infection in mouse models
Collaborator Contribution The Tripp group have supplied expertise and access to reagents such as influenza strains as well a performed infections in mice with highly pathogenic influenza strains.
Impact 1. Leeming, G.H., Kipar, A., Hughes, D.J., Bingle, L., Bennett, E., Moyo, N., Tripp, R.A., Bigley, A., Bingle, C.D., Sample, J.T., Stewart, J.P. Gammaherpesvirus infection modulates the temporal and spatial expression of SCGB1A1 (CCSP) and BPIFA1 (SPLUNC1) in the respiratory tract. Laboratory Investigation, In Press.
Start Year 2011
 
Description Simon Carding vaccine 
Organisation Quadram Institute Bioscience
Country United Kingdom 
Sector Academic/University 
PI Contribution Design and preliminary testing of bacterial OMVs containing influenza proteins for use as a mucosal vaccine against avian influenza
Collaborator Contribution Construction and production of bacterial OMVs containing influenza proteins for use as a mucosal vaccine against avian influenz
Impact none as yet
Start Year 2018
 
Description Podcast interview 
Form Of Engagement Activity A broadcast e.g. TV/radio/film/podcast (other than news/press)
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Media (as a channel to the public)
Results and Impact Podacast for Guardian Science Weekly about how respiratory viruses cause disease focussed around the Covid-19 epidemic.
Year(s) Of Engagement Activity 2020