IDENTIFICATION OF A PROMOTER MAPPING WITHIN THE REITERATED SEQUENCES THAT FLANK THE HERPES-SIMPLEX VIRUS TYPE-1 UL REGION

Analysis of the promoter for the herpes simplex virus (HSV) immediate-early (alpha) gene alpha0 in a short-term transient expression assay revealed that a SacI-to-NcoI fragment from -786 to +148 relative to the cap site directed the synthesis of chloramphenicol acetyltransferase when the fragment was present in either orientation. Although the constitutive levels of promoter activity were similar with either orientation, the reverse-orientation promoter was not induced in response to infection with HSV. Analysis of sequences composing the putative promoter in the opposite orientation revealed the presence of important regulatory elements associated with alpha promoters. These include an alpha-trans-inducing factor (alpha-TIF)-like response element, a high-affinity ICP4-binding site, numerous Sp1-binding sites, and a TATA box. Sequences contained within this region formed specific DNA-protein complexes in extracts from mock-infected and HSV-infected HeLa cells. Transient expression assays revealed that this sequence was positively regulated by the alpha0 and alpha-TIF genes but negatively regulated by alpha4. Finally, nuclear run-on transcription assays revealed that this promoter is active in its correct genomic context during the course of virus infection. We suggest that the promoter is a hybrid between an alpha and beta promoter because it exhibits maximal expression at 8 h postinfection and is expressed in the presence of cycloheximide.