We propose to investigate the immediate early (IE) genes of human cytomegalovirus (CMV) to define their role in reactivation from latency and enhancement of viral gene expression resulting in persistent infection of the human host. Elements upstream of the major IE gene will be characterized to determine their role in transcription. An analysis of the structure of the IE genes as to exons, introns, DNA sequence, and protein sequence will continue in order to better understand function of the IE proteins. The role of the major IE and/or minor IE proteins on transcription of the CMV genome or transfected early or late viral genes will be investigated using SV-ori expression vectors that replicate in cells (COS) constitutively synthesizing T-antigen. Two genes highly transcribed at early times will be investigated as examples of regions turned-on after synthesis of the IE proteins. One gene is an early gene based on mRNA and protein product in the cytoplasm. This gene codes for a protein kinase presumably necessary for regulation and/or viral DNA replication. Although transcribed at early times, the other gene is a late gene according to the above criteria. This gene codes for a virion structural component. These two different viral genes will be investigated as an approach towards understanding posttranscriptional regulation of CMV gene expression. Human CMV replicates slowly in the infected cell. It is proposed that the major IE proteins of CMV enhance transcription of early and late viral genes as well as cellular genes. It is hypothesized that a viral protein functions at three to four hours postinfection to control the amount of transcription of the major IE gene. The viral RNAs of defined early and late genes are discriminated by a posttranscriptional regulation mechanism. These regulatory viral gene products and events play important roles in the replication of infectious virus in the human host.
