Cellular Repressor Inhibits Human Cytomegalovirus Transcription from the UL127 Promoter

Abstract

The region of the human cytomegalovirus (HCMV) genome between the UL127 promoter and the major immediate-early (MIE) enhancer is referred to as the unique region. The role of this region during a viral infection is not known. In wild-type HCMV-infected permissive fibroblasts, there is no transcription from the UL127 promoter at any time during productive infection. Our investigators previously reported that the region upstream of the UL127 TATA box repressed expression from the UL127 promoter (C. A. Lundquist et al., J. Virol. 73:9039-9052, 1999). The region was reported to contain functional NF1 DNA binding sites (L. Hennighausen and B. Fleckenstein, EMBO J. 5:1367-1371, 1986). Sequence analysis of this region detected additional consensus binding sites for three transcriptional regulatory proteins, FoxA (HNF-3), suppressor of Hairy wing, and CAAT displacement protein. The cis -acting elements in the unique region prevented activation of the early UL127 promoter by the HCMV MIE proteins. In contrast, deletion of the region permitted very high activation of the UL127 promoter by the viral MIE proteins. Mutation of the NF1 sites had no effect on the basal activity of the promoter. To determine the role of the other sites in the context of the viral genome, recombinant viruses were generated in which each putative repressor site was mutated and the effect on the UL127 promoter was analyzed. Mutation of the putative Fox-like site resulted in a significant increase in expression from the viral early UL127 promoter. Insertion of wild-type Fox-like sites between the HCMV immediate-early (IE) US3 TATA box and the upstream NF-κB-responsive enhancer (R2) also significantly decreased gene expression, but mutated Fox-like sites did not. The wild-type Fox-like site inhibits activation of a viral IE enhancer-containing promoter. Cellular protein, which is present in uninfected or infected permissive cell nuclear extracts, binds to the wild-type Fox-like site but not to mutated sites. Reasons for repression of UL127 gene transcription during productive infection are discussed.