SEQUENCE AND TRANSCRIPTIONAL ANALYSIS OF THE GUINEA-PIG CYTOMEGALOVIRUS UL97 HOMOLOG

Guinea pig cytomegalovirus (GPCMV) displays a similar pathogenesis to human cytomegalovirus (HCMV), and the guinea pig has been used as a mo del system for testing anti-CMV therapies, However, not all agents act ive against HCMV share antiviral activity against GPCMV. For example, GPCMV appears resistant to the nucleoside analog, ganciclovir. The mol ecular basis for this discrepancy in antiviral susceptibility is unkno wn because to date there has been little analysis of the GPCMV genome. For HCMV, the antiviral effect of ganciclovir depends upon phosphoryl ation of the drug to its active form. This effect is mediated by the v iral UL97 gene product. In order to begin to explore the molecular bas is of the resistance of GPCMV to ganciclovir, experiments were underta ken to test whether the GPCMV genome encoded a homolog of the HCMV UL9 7 gene. Based on the prediction of co-linearity of UL97 homologs withi n the respective viral genomes, the EcoR I S and F fragments of the GP CMV genome were cloned and partially sequenced, A 1815 base pair open reading frame (ORF) capable of encoding a 604 amino acid (aa) protein was identified spanning portions of the EcoR I S and adjacent EcoR I F genome fragments, Computer-assisted matrix analyses revealed identity between this ORF and the HCMV UL97 gene. ORFs upstream of the GPCMV U L97 gene were identified which shared homology with the HCMV UL95 and 96 genes. Northern blot analyses identified a UL97-specific mRNA of 3. 9 kb which was expressed at ''early'' times post-infection. RNA transc ripts of 6.0 and 4.6 kb were identified which corresponded to the UL95 and UL96 homolog coding sequences, respectively. Comparison of the GP CMV UL97 sequence to that of other herpesvirus homologs as well as tha t of ganciclovir-resistant clinical isolates of HCMV identified noncon servative aa substitutions in two domains involved in catalysis and su bstrate recognition.