Promoter activity of the 5'-flanking region of the pseudorabies virus (PRV) early protein 0 (EP0) gene was analysed by transient transfection assays employing chloramphenicol acetyl transferase (CAT) reporter constructs. We identified a 213 bp segment of the viral genome that was capable of efficiently driving expression of the EP0 gene and a linked reporter gene upon transient transfection into Vero cells. This segment lacked the typical TATA element, and possessed an initiator element and the putative binding sites for the transcription factor Sp1 and immediate-early protein IE180, a strong transactivator of PRV. By analysing 5'-deletion mutants of the segment, a 48 bp segment (from nucleotide positions -65 to - 17), which possessed three Sp1 binding sites, was identified to be critical for the promoter activity. Cotransfection of Vero cells with the mutant constructs and an IE180 expression plasmid resulted in transactivation of only those constructs in which the Sp1 sites were present. These results indicate that the EP0 gene may be transcribed from the TATA-less promoter that responds to Sp1.
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