Identification and transcriptional analyses of the UL3 and UL4 genes of equine herpesvirus 1, homologs of the ICP27 and glycoprotein K genes of herpes simplex virus.

The DNA sequence of 3,240 nucleotides of the XbaI G fragment located in the unique long (UL) region of the equine herpesvirus 1 genome revealed two major open reading frames (ORFs) designated UL3 and UL4. The UL3 ORF of 470 amino acids (aa) maps at nucleotides (nt) 4450 to 3038 from the long terminus, and its predicted 51.4-kDa protein product exhibits significant homology to the ICP27 alpha regulatory protein of herpes simplex virus type 1 (HSV-1; 32% identity) and to the ORF4 protein of varicella-zoster virus (13% identity). Interestingly, a zinc finger motif is conserved in the C-terminal domains of both ICP27 of HSV-1 (aa 483 to 508) and UL3 of equine herpesvirus 1 (aa 441 to 466). The UL4 ORF of 343 aa maps at nt 5618 to 4587 and could encode a protein of 38.1 kDa which exhibits significant homology to the UL53 protein (cell fusion protein or glycoprotein K) of HSV-1 (26% identity) and to the ORF5 protein of varicella-zoster virus (33% identity). Analyses of the UL4 amino acid sequence revealed domains characteristic of a membrane-bound glycoprotein and included potential signature sequences for (i) a signal sequence, (ii) two N-linked glycosylation sites, and (iii) four transmembrane domains. Nucleotide sequence analyses also revealed potential TATA boxes located upstream of the UL3 and UL4 ORFs. However, only a single polyadenylation signal (nt 2988 to 2983) was detected downstream of the UL3 ORF. Northern (RNA) blot hybridization and S1 nuclease analyses were used to map and characterize the UL3 and UL4 mRNAs. Metabolic inhibitors were used to identify the kinetic class of these two genes. The data revealed that UL3 is an early gene that encodes a 1.6-kb mRNA, while UL4 is a late gene encoding a 3.8-kb mRNA that overlaps the UL3 transcript. Both transcripts were shown by S1 nuclease analyses to initiate 24 to 26 nt downstream of their respective TATA boxes and to have a common transcription termination signal as a pair of 3'-coterminal mRNAs.