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Title: Characterisation of the murine gammaherpesvirus-68 G protein-coupled receptor
Author: Wakeling, Madelaine N.
Awarding Body: University of Edinburgh
Current Institution: University of Edinburgh
Date of Award: 2001
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Murine gammaherpesvirus 68 (MHV-68) is a naturally occurring virus found in wild rodents. It productively infects a range of cells and can be grown to high titres. MHV-68 also infects laboratory mice, thus making it an excellent small animal model for studying gammaherpesvirus infection. Herpesvirus genomes contain a variety of homologues of cellular genes involved in immune regulation and/or cell growth and proliferation. Cellular homologues encoded by MHV-68 include Bcl-2, cyclin D and a G protein-coupled receptor (GPCR) that shares greatest amino acid identity with the mammalian chemokine receptor, CXCR2. GPCRs are a superfamily of seven-transmembrane signalling molecules, and homologues occur in several herpesviruses. The KSHV GPCR is a functional, constitutively active oncoprotein that induces vascular endothelial growth factor. The aim of this project was to characterise the MHV-68 GPCR (ORF74) and investigate its role in viral pathogenesis. This was approached in several ways. Firstly, the transcription pattern of the gene was determined using Northern analysis: GPCR expression was detected at early and late time-points during lytic infection on multiple rare transcripts. The size of the transcripts suggested they might be polycistronic and subsequent RT-PCR analysis demonstrated that the GPCR was co-expressed with v-Bcl-2, both in vitro and in vivo. The subcellular localisation of the GPCR protein was investigated using an "epitope-tagging" strategy and immunofluorescence experiments revealed an expression pattern consistent with localisation to the cell surface. The transforming activity of the GPCR was examined using conventional tissue culture-based assays. NIH3T3 cells expressing the GPCR exhibited focus formation and anchorage independent growth in soft agar. Lastly, a recombinant virus lacking the GPCR was generated by homologous recombination. However, this recombinant virus could not be purified from the parental wild type virus. In conclusion, the MHV-68 GPCR is a viral oncogene that is likely to contribute to the long-term persistence of MHV-68.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available