Molecular biology of foot-and-mouth disease virus:entry replication and assembly

Lead Research Organisation: The Pirbright Institute
Department Name: UNLISTED

Abstract

The proposed research aims to investigate the following key stages of the FMDV life cycle: i) Membrane penetration by using model membranes, ii) Capsid assembly by using fluorescent and structural approaches, iii) Genome replication and packaging by using cell culture & cell-free replication systems. Work will build upon the research findings, hypotheses, collaborations and techniques developed during nine years of picornavirus research at the University of Leeds. This work has involved viruses such as poliovirus (PV), human rhinovirus (HRV) and of particular relevance to this application, the aphthovirus equine rhinitis A virus (ERAV), the latter having been developed specifically as a surrogate model for FMDV. Substantial preliminary data exists for several aspects of this proposal which will expedite rapid progress in i) experimental work, ii) publication in peer reviewed journals and iii) the procurement of external funding with which to expand the program.
 
Description Several key findings resulting from this research are summarized here:
1) The first demonstration that the picornavirus capsid protein VP4 induces membrane permeability by the formation of a size-selective multimeric pore.
2) The first demonstration that the picornavirus equine rhinitis A virus (ERAV) is dependent on endosome acidification during the process of cell entry.
3) The development of a novel plate-based high-throughput assay for virus stability and vaccine formulation.
4) Generating and reporting the crystal structure of equine rhinitis a virus in complex with its sialic acid receptor.
Exploitation Route The findings have contributed to further academic research into picornavirus particle alterations, vaccine stability and led to the identification of novel targets such as conserved epitopes suitable for disease control or diagnostics.
Sectors Agriculture, Food and Drink,Pharmaceuticals and Medical Biotechnology,Other

 
Description The findings have contributed to the development of FMD vaccine stability and led to the identification of novel targets such as conserved epitopes suitable for disease control or diagnostics.
First Year Of Impact 2014
Sector Agriculture, Food and Drink,Manufacturing, including Industrial Biotechology,Other
Impact Types Societal,Economic

 
Title Deep sequencing to identify RNA packaging signals 
Description A novel and simple approach to identify predicted RNA secondary structures involved in genome packaging in positive sense RNA viruses (e.g. the picornavirus foot-and-mouth disease virus [FMDV]). By interrogating deep sequencing data generated from both packaged and unpackaged populations of RNA, we have determined multiple regions of the genome with constrained variation in the packaged population. Predicted secondary structures of these regions revealed stem-loops with conservation of structure and a common motif at the loop. Disruption of these features resulted in attenuation of virus growth in cell culture due to a reduction in assembly of mature virions. This study provides evidence for the involvement of predicted RNA structures in picornavirus packaging and offers a readily transferable methodology for identifying packaging requirements in many other viruses. 
Type Of Material Technology assay or reagent 
Year Produced 2017 
Provided To Others? Yes  
Impact Impact so far is fundamental knowledge but with several potential avenues for future impact in for example improved vaccines. 
 
Description Dundee 
Organisation University of Dundee
Country United Kingdom 
Sector Academic/University 
PI Contribution Co-INvestigator on BBSRC sLoLa award
Collaborator Contribution Co-INvestigator on BBSRC sLoLa award
Impact BBSRC sLoLa award ongoing
Start Year 2012
 
Description Edinburgh 
Organisation University of Edinburgh
Country United Kingdom 
Sector Academic/University 
PI Contribution Co-INvestigator on BBSRC sLoLa award
Collaborator Contribution Co-INvestigator on BBSRC sLoLa award
Impact BBSRC sLoLa award ongoing
Start Year 2012
 
Description HMS 
Organisation Harvard University
Department Harvard Medical School
Country United States 
Sector Academic/University 
PI Contribution Collaboration
Collaborator Contribution Collaboration
Impact Collaborative research
 
Description Imperial 
Organisation Imperial College London
Country United Kingdom 
Sector Academic/University 
PI Contribution Joint supervision of PhD student
Collaborator Contribution Joint supervision of PhD student
Impact One successful completed PhD student. Papers published or in preparation. Another PhD student in progress.
Start Year 2010
 
Description JN - Preparation of capped mesoporpus silica nanoparticles 
Organisation Polytechnic University of Valencia
Country Spain 
Sector Academic/University 
PI Contribution We were involved with testing the nanoparticles once they had been made. We also contributed purified antibody and shipped it to our collaborators for the generation of one type of nanoparticle, which we subsequently tested. We were involved with the intellectual design of experiments
Collaborator Contribution Our partners generated the nanoparticles and shipped them to us for testing. Our partners were also involved with the intellectual design of experiments.
Impact Experimental work performed to investigate the use of mesoporous silica nanoparticles to detect FMDV proteases in clinical samples
Start Year 2016
 
Description JS - Production of monoclonal antibodies that recognise FMDV VP4 from mouse spleens 
Organisation University of Edinburgh
Department The Roslin Institute
Country United Kingdom 
Sector Academic/University 
PI Contribution VLPs displaying the N-terminal 15 amino acids of FMDV were designed and produced at The Pirbright Institute and then mice were immunised with them. The spleens of the immunised mice were harvested and splenocytes stored. The response to the VP4 sequence was checked by ELISA and the mouse with the best response was selected to send the splenocytes for monoclonal production.
Collaborator Contribution The collaborators took the splenocytes obtained from the mouse experiments and carried out the fusions. They then screened the supernatants for positive wells that detected the VP4 sequence.
Impact Positive wells have been identified indicating some monoclonals that are specific for VP4 have been generated. These will be further screened for ability to recognise virus and to neutralise infection.
Start Year 2017
 
Description KU Leuven 
Organisation University of Leuven
Country Belgium 
Sector Academic/University 
PI Contribution collaboration
Collaborator Contribution collaboration
Impact collaboration
Start Year 2012
 
Description Leeds FBS 
Organisation University of Leeds
Department Leeds Institute of Molecular Medicine
Country United Kingdom 
Sector Academic/University 
PI Contribution Research
Collaborator Contribution Research and student supervision
Impact Research
Start Year 2009
 
Description Oxford - Strubi 
Organisation University of Oxford
Country United Kingdom 
Sector Academic/University 
PI Contribution Collaboration on virus structural biology and joint PhD studentship
Collaborator Contribution Collaboration on virus structural biology and joint PhD studentship
Impact Collaboration on virus structural biology has produced a number of novel virus structures and new understanding of virus entry and packaging
Start Year 2008
 
Description St Andrews 
Organisation University of St Andrews
Country United Kingdom 
Sector Academic/University 
PI Contribution Co-investigator on BBSRC sLoLa
Collaborator Contribution Lead-investigator on BBSRC sLoLa
Impact BBSRC sLoLa ongoing
Start Year 2012
 
Description Lecture on foot-and-mouth disease virus to Merrist Wood College students 
Form Of Engagement Activity Participation in an open day or visit at my research institution
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Undergraduate students
Results and Impact Lecture on foot-and-mouth disease virus to Merrist Wood College students
Year(s) Of Engagement Activity 2013
 
Description Surrey Science And Technology Regional Organisation event at Brooklands 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? Yes
Geographic Reach Local
Primary Audience Schools
Results and Impact Hopefully inspired children to think about science
Year(s) Of Engagement Activity 2013