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The human cytomegalovirus (HCMV) UL94 gene product is a herpesvirus-common virion protein that is expressed with true late kinetics. To identify the importantcis - andtrans -acting factors which contribute to UL94 transcriptional regulation, we have cloned, sequenced, and analyzed UL94 promoter function by transient transfection analysis. Transfection of UL94 promoter-reporter gene constructs into permissive human fibroblasts or U373(MG) cells indicated that promoter activity was detected following infection with HCMV. Point mutations within a TATA-like element located upstream of the RNA start site significantly reduced UL94 promoter activity. Deletion mutagenesis of the promoter indicated that a positive regulatory element (PRE) was likely to exist downstream of the UL94 mRNA start site, while a negative regulatory element (NRE) was present upstream of the TATA box. At late times of infection, the PRE appeared to have a dominant effect over the NRE to stimulate maximum levels of UL94 promoter activity, while at earlier times of infection, no activity associated with the PRE could be detected. The NRE, however, appeared to cause constitutive down-regulation of UL94 promoter activity. Binding sites for the cellular p53 protein located within the NRE appeared to contribute to NRE function, and NRE function could be recapitulated in cotransfection assays by concomitant expression of p53 and HCMV IE2-86 protein. Our results suggest a novel mechanism by which the cellular protein p53, which is involved in both transcriptional regulation and progression of cellular DNA synthesis, plays a central role in the regulation of a viral promoter which is not activated prior the onset of viral DNA replication.

journal of virology

Identification of Positive and Negative Regulatory Regions Involved in Regulating Expression of the Human Cytomegalovirus UL94 Late Promoter: Role of IE2-86 and Cellular p53 in Mediating Negative Regulatory Function