Deletion of the R78 G Protein-Coupled Receptor Gene from Rat Cytomegalovirus Results in an Attenuated, Syncytium-Inducing Mutant Strain

Abstract

The rat cytomegalovirus (RCMV) R78 gene belongs to an uncharacterized class of viral G protein-coupled receptor (GCR) genes. The predicted amino acid sequence of the R78 open reading frame (ORF) shows 25 and 20% similarity with the gene products of murine cytomegalovirus M78 and human cytomegalovirus UL78, respectively. The R78 gene is transcribed throughout the early and late phases of infection in rat embryo fibroblasts (REF) in vitro. Transcription of R78 was found to result in three different mRNAs: (i) a 1.8-kb mRNA containing the R78 sequence, (ii) a 3.7-kb mRNA containing both R77 and R78 sequences, and (iii) a 5.7-kb mRNA containing at least ORF R77 and ORF R78 sequences. To investigate the function of the R78 gene, we generated two different recombinant virus strains: an RCMV R78 null mutant (RCMVΔR78a) and an RCMV mutant encoding a GCR from which the putative intracellular C terminus has been deleted (RCMVΔR78c). These recombinant viruses replicated with a 10- to 100-fold-lower efficiency than wild-type (wt) virus in vitro. Interestingly, unlike wt virus-infected REF, REF infected with the recombinants develop a syncytium-like appearance. A striking difference between wt and recombinant viruses was also seen in vivo: a considerably higher survival was seen among recombinant virus-infected rats than among RCMV-infected rats. We conclude that the RCMV R78 gene encodes a novel GCR-like polypeptide that plays an important role in both RCMV replication in vitro and the pathogenesis of viral infection in vivo.

Rat Cytomegalovirus-Accelerated Transplant Vascular Sclerosis Is Reduced with Mutation of the Chemokine-Receptor R33

“…From the above results, the prediction would be that RCMV contains a US28 GPCR homologue. However, unlike HCMV that encodes four GPCRs, RCMV does not encode a genetic homologue to HCMV US28 but does contain a CC (R33) and a CXC (r78)‐chemokine receptor in the viral genome (30,36). Since RCMV R33 is expressed at early intervals after infection, similar to US28, we examined the contribution of this viral gene product in the induction of SMC migration (30,37).…”

The human cytomegalovirus UL78 gene is highly conserved among clinical isolates, but is dispensable for replication in fibroblasts and a renal artery organ-culture system

“…Interestingly, both the UL33and UL78-like genes exhibit conservation of position, as well as orientation, within the genomes of b-herpesviruses. Until now, six UL78-like genes had been identified: HCMV UL78, MCMV M78, RCMV R78 and the U51 ORFs of HHV-6A, HHV-6B and HHV-7 (Gompels et al, 1995;Nicholas, 1996;Rawlinson et al, 1996;Beisser et al, 1999). HHV-6 U51 is known to be expressed on the surface of infected T cells and has been found to bind RANTES/ CCL5 with nanomolar affinity, as well as to downregulate the transcription of this chemokine (Menotti et al, 1999;Milne et al, 2000).…”

Mutational analysis of the R33-encoded G protein-coupled receptor of rat cytomegalovirus: identification of amino acid residues critical for cellular localization and ligand-independent signalling

“…Human CMV (HCMV) carries four such genes: US27, US28, UL33 and UL78 (Chee et al, 1990a, b). Only two of these, UL33 and UL78, have been found to have counterparts in rodent CMVs: rat CMV (RCMV; R33 and R78, respectively; Beisser et al, 1998Beisser et al, , 1999Vink et al, 1999Vink et al, , 2001) and murine CMV (MCMV; M33 and M78, respectively; Davis-Poynter et al, 1997;Rawlinson et al, 1996).…”