Studentship: Is PB1-F2 a virulence factor for avian influenza virus in poultry?

Lead Research Organisation: The Pirbright Institute
Department Name: UNLISTED

Abstract

Studentship; PB1-F2 is a 90 amino acid protein discovered in 2001 to be expressed from the second influenza genome segment in a +1 reading frame from the main protein PB1. Since then several different functions have been attributed to the protein including pro-apoptotic, interferon modulator, viral polymerase activity modulator and an inflammatory inducer. These functions have mostly been prescribed to the laboratory adapted influenza strain PR8 in mammalian cell lines. Recent work has shown that the PB1-F2 protein from other strains of influenza virus don’t behave in the same way. PB1-F2 is well conserved in avian influenza isolates in contrast to its frequent truncation or deletion in swine and human isolates. Therefore it stands that in the avian host PB1-F2 has an important function. Avian influenza makes frequent incursions into poultry populations resulting in economical and health burdens for farmers. In our fight to protect poultry populations from avian influenza it is important to understand any virulence factors in avian influenza strains so proper prevention and mitigation strategies can be developed. This PhD will look at several different aspects to do with PB1-F2 function in poultry to help shed some light on what it does and whether it is a virulence factor. The impact of PB1-F2 on viral replication kinetics and innate immune evasion will be investigated as well as trying to identify potentially new interactions that it may be involved in. Whilst investigating the question of PB1-F2 the PhD student will develop and use biologically relevant culture systems including immortalised chicken cell lines, primary mono-cultures, well differentiated respiratory and enteric tract cultures and ex vivo explants.
 
Description We have determined that the small accessory protein PB1-F2 of avian influenza viruses have an avian host specific role. Infection of chickens with avian influenza that contains a full length PB1-F2 protein results in prolonged infectious virus shedding from both ends of the bird and subsequently allows a longer transmission window to naïve birds. A full length PB1-F2 protein in an avian influenza virus reduces pathogenicity of infection in chickens but not in mice (used as a mammalian model). In chicken hosts the presence of a full length PB1-F2 protein suppresses Interferon stimulated genes and pro-inflammatory cytokines. We have mapped those functions to the C-terminus of the PB1-F2 protein and that function, cellular location and stability of the protein in a chicken cells are all related through the C-terminal sequence of the protein. We have demonstrated that there is a unique pressure to retain a full length PB1-F2 protein in avian host isolates of influenza viruses in contrast to virus recovered from mammalian hosts.
Exploitation Route This fundamental knowledge about PB1-F2 role could be utilized to understand it PB1-F2 can be used as a target for anti-viral therapy or vaccine component.
Sectors Agriculture, Food and Drink

 
Description BBSRC East Bio DTP Studentship
Amount £100,000 (GBP)
Organisation Biotechnology and Biological Sciences Research Council (BBSRC) 
Sector Public
Country United Kingdom
Start 09/2017 
End 08/2021
 
Description Paul Digard - The Roslin Insititute 
Organisation University of Edinburgh
Department The Roslin Institute
Country United Kingdom 
Sector Academic/University 
PI Contribution We generated constructs that express PB1-F2 from a variety of avian influenza strains in eukaryotic expression plasmids. We generated Chimera PB1-F2 expression plasmids where the C-terminus of the PB1-F2 are switched between strains. Using these plasmids we have determined the localization and antagonist activity toward the IFN beta signaling pathway and NF-kb signaling pathway in human and chicken cells. We have therefore provided reagents and intellectual input to the collaboration.
Collaborator Contribution Our collaborators have used these plasmids in there well defined stability assays to determine sequence specific effects on this characteristic. Intellectual input and expertise in a particular assay were made by these collaborators.
Impact We have successfully published some of this work; "The cellular localization of avian influenza virus PB1-F2 protein alters the magnitude of IFN2 promoter and NF?B-dependent promoter antagonism in chicken cells." doi: 10.1099/jgv.0.001220. We have an ongoing joint PhD studentship in this area. This collaboration remains current and active.
Start Year 2017
 
Description Steve Goodbourn - St Georges - chicken innate molecule interactions. 
Organisation Ulster University
Department Biomedical Sciences Research Centre
Country United Kingdom 
Sector Academic/University 
PI Contribution We have collaborated by producing yeast- 2-hybrid plasmids containing the influenza PB1-F2 gene from various avian influenza strains. The PhD student on this studentship then visited Professor Goodbourn's laboratory to carry out the yeast-2-hybrid screen for interaction innate chicken molecules.
Collaborator Contribution Professor Goodbourn's laboratory provided yeast-2-hybrid plasmids containing various innate chicken molecules to be screened for interaction with avian influenza virus PB1-F2 protein. They also offered help and guidance carrying out the interaction technique to understand if PB1-F2 directly associated with chicken innate components. They also provided reporter plasmids and guidance on innate reporter assays in chicken cells.
Impact Successful publication of work. doi: 10.1099/jgv.0.001220. The cellular localization of avian influenza virus PB1-F2 protein alters the magnitude of IFN2 promoter and NF?B-dependent promoter antagonism in chicken cells.
Start Year 2016
 
Description FLI - presentation 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Postgraduate students
Results and Impact Presentation of current research to the FLI - Germany to build collaborations 19/20 November 2018
Year(s) Of Engagement Activity 2018
 
Description Oral presentation at the Global Alliance for Research on Avian Disease meeting in Hanoi, Vietnam 15th -19th Jan 2018 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Other audiences
Results and Impact A talk was given on the PB1-F2 function in avian influenza infection of avian hosts at the GARAD meeting. This meeting was attended by approximately 120 researchers and poultry industry members from a wide international geographic region.
Year(s) Of Engagement Activity 2018