The gammaherpesvirus thymidine kinase

Lead Research Organisation: University of Cambridge
Department Name: Pathology

Abstract

Herpesviruses are ubiquitous infections and cause many clinical problems in immunosuppressed patients. The most effective treatment to date has been anti-viral drugs such as aciclovir, which exploit a viral enzyme called thymidine kinase (TK). The gamma-herpesviruses have this enzyme, but it is a much weaker enzyme than that of alpha-herpesviruses, against which aciclovir was developed. We suspect that the gamma-herpesvirus TK has different functions, and might therefore best be attacked through different types of drugs.
The human gamma-herpesviruses only infect primates, making many experiments impractical. We are therefore defining what the gamma-herpesvirus TK does using a related murine virus, called MHV-68. We know that the MHV-68 TK is important for viral replication in mice. We aim now to work out why, specifically whether it works in the same way as the alpha-herpesvirus TK, or whether it has very different functions that would be best inhibited in different ways. By understanding how the MHV-68 TK works, we should be able to provide a basis for effective TK-based therapies in gamma-herpesvirus infections.

Technical Summary

Herpesviruses are major causes of disease. Thymidine kinase (TK)-based therapies have proved very effective against alpha- and beta-herpesviruses. Analogous therapies, combined with lytic cycle induction, are therefore being developed against gamma-herpesviruses. However, it is unclear whether gamma-herpesvirus TKs are equivalent to those of alpha-herpesviruses. They are larger than alpha-herpesvirus TKs, differently distributed in cells, and much less active in nucleoside phosphorylation. Because gamma-herpesviruses do not establish latency in terminally differentiated cells, they may have less need to phosphorylate nucleotides directly for their replication, allowing TK to evolve other functions.
Viral escape mutants inevitably limit the efficacy of drug treatments. It is therefore important for TK-based therapies to understand what role TK plays in gamma-herpesvirus pathogenesis. We also need to improve our basic understanding of these ubiquitous and complex pathogens. Human gamma-herpesviruses lack good experimental lytic replication systems. We and others have therefore developed MHV-68 - a murid gamma-herpesvirus - as a model system for analyzing conserved gamma-herpesvirus lytic gene functions in infected cells and infected animals. TK-deficient MHV-68 is severely attenuated for host colonization. Preliminary data suggest that this reflects a role for TK not in DNA replication in quiescent cells but in viral capsid transport, especially on entry into myeloid cells. This finding has important implications both for how TK-directed therapies might work and for how gamma-herpesviruses normally colonize their hosts.
Our aim now is to define a molecular basis for gamma-herpesvirus TK function, comparing EBV, KSHV and MHV-68. We will focus mainly on virus-infected cells by generating BAC-based MHV-68, EBV and KSHV TK mutants. We will identify how the large N-terminal domain shared by all gamma-herpesvirus TKs contributes to its function, particularly what cellular proteins it interacts with and how its function relates to nucleoside phophorylation. Using MHV-68, we will establish how the molecularly defined TK function explains the dramatic in vivo attenuation of TK-deficient mutants. Our overall aim is to establish a general model of how gamma-herpesvirus TKs work, which should both inform an important area of anti-viral therapy and provide significant new insights into basic gamma-herpesvirus biology.

Publications

10 25 50
 
Description BELVIR collaborative grant
Amount € 100,000 (EUR)
Organisation National Fund for Scientific Research (NFSR) 
Sector Charity/Non Profit
Country Belgium
Start 03/2013 
End 03/2016
 
Description Comprehensive mapping of rhadinovirus dissemination and persistence
Amount £608,871 (GBP)
Funding ID BB/J014419/1 
Organisation Biotechnology and Biological Sciences Research Council (BBSRC) 
Sector Public
Country United Kingdom
Start 01/2013 
End 01/2016
 
Description Wellcome Trust project grant
Amount £350,000 (GBP)
Organisation Wellcome Trust 
Sector Charity/Non Profit
Country United Kingdom
Start 06/2010 
End 06/2013
 
Title generation of monoclonal antibodies 
Description monoclonal antibodies to KSHV and EBV thymidine kinases 
Type Of Material Antibody 
Year Produced 2008 
Provided To Others? Yes  
Impact unknown as yet 
 
Title knockout viruses 
Description generated knockout + fluorescent tagged viruses 
Type Of Material Technology assay or reagent 
Year Produced 2006 
Provided To Others? Yes  
Impact publications from other labs (we do not request joint authorship for providing published reagents) 
 
Description Australia 
Organisation The Walter and Eliza Hall Institute of Medical Research (WEHI)
Department Immunology Division
Country Australia 
Sector Academic/University 
PI Contribution Shared resources / projects leading to joint publications
Collaborator Contribution Shared resources / projects leading to joint publications
Impact See publications with G.T.Belz in main list. Also shared Wellcome Trust project grant.
 
Description Babraham 
Organisation Immunobiology Ltd, Babraham, UK
Country United Kingdom 
Sector Private 
PI Contribution shared PhD student supervision
Collaborator Contribution shared PhD student (BBSRC CASE)
Impact See joint publication with C.Colaco
Start Year 2007
 
Description Belgium 
Organisation University of Liege
Department Laboratory of Immunology - Vaccinology
Country Belgium 
Sector Academic/University 
PI Contribution Shared projects / resources leading to joint publications
Collaborator Contribution Shared projects / resources leading to joint publications.
Impact see publications with L.Gillet in main list
Start Year 2006
 
Description Cambridge 
Organisation University of Cambridge
Department Department of Medicine
Country United Kingdom 
Sector Academic/University 
PI Contribution worked on joint project
Collaborator Contribution joint project
Impact see joint publications with S.Efstathiou in main list
Start Year 2006
 
Description Portugal 
Organisation University of Lisbon
Country Portugal 
Sector Academic/University 
PI Contribution Shared resources / projects leading to joint publications.
Collaborator Contribution Shared resources / projects leading to joint publications
Impact See publications with J.P. Simas in main list
 
Description Department open day 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Primary Audience Public/other audiences
Results and Impact Engaged with public, explaining research in the department and its context. Also built some models of viruses.

Better public understanding of science.
Year(s) Of Engagement Activity 2008,2009
 
Description Interview for New Scientist 
Form Of Engagement Activity A magazine, newsletter or online publication
Part Of Official Scheme? No
Type Of Presentation Keynote/Invited Speaker
Geographic Reach International
Primary Audience Public/other audiences
Results and Impact Phone interview, transcribed by journalist to contribute to article.

Emails from members of public enquiring about herpesvirus vaccines.
Year(s) Of Engagement Activity 2012
 
Description interview 
Form Of Engagement Activity A magazine, newsletter or online publication
Part Of Official Scheme? No
Primary Audience Public/other audiences
Results and Impact telephone interview with journalist

web interest
Year(s) Of Engagement Activity 2010
 
Description naked scientist 
Form Of Engagement Activity A press release, press conference or response to a media enquiry/interview
Part Of Official Scheme? No
Primary Audience Public/other audiences
Results and Impact Question and answer session on radio.

audience showed interest
Year(s) Of Engagement Activity 2010