DNA Polymerase Processivity Factor of Human Cytomegalovirus May Be a Key Molecule for Molecular Coupling of Viral DNA Replication to Transcription

Abstract

Human cytomegalovirus (HCMV) is a member of the betaherpesvirus family. Like all herpesviruses, HCMV is an enveloped, double-stranded DNA virus. The genome of HCMV is 240,000 bp in size with at least 150 known open reading frames (ORFs) (Dunn, Chou et al. 2003). A majority of the ORFs are nonessential for viral replication in cell culture. These nonessential ORFs likely encode proteins with redundant functions or proteins that may be required for replication in the human host. In addition, several ORFs are beneficial but not required for replication. However, approximately one-quarter, or 41 ORFs, are absolutely required for viral replication (Yu, Silva et al. 2003). The UL44 gene is essential. During productive infection, HCMV genes are expressed in a temporal cascade, designated immediate early (IE), delayed early, and late. The major IE genes (MIE) UL123/122 (IE1/IE2) play a critical role in subsequent viral gene expression and the efficiency of viral replication (Meier and Stinski 1997; Meier and Pruessner 2000; Meier, Keller et al. 2002; Isomura and Stinski 2003; Isomura, Tsurumi et al. 2004; Isomura, Stinski et al. 2005). The IE72 protein, the predominant product of the IE1 transcript, is encoded by exons 2 and 3 spliced to exon 4. The IE86 protein, the predominant product of the IE2 transcript, is encoded by exons 2 and 3 spliced to exon 5. Translation of the IE1 and IE2 transcripts begins in exon 2. The IE72 protein is not essential for viral replication at high MOI, but the IE86 protein is essential (Marchini, Liu et al. 2001). The early viral genes encode proteins necessary for viral DNA replication (Pari and Anders 1993). Following viral DNA replication, delayed early and late viral genes are expressed which encode structural proteins for virion production. The UL 44 protein (pUL44), which binds double-stranded DNA, is an essential accessory protein for viral DNA replication and interacts specifically with the viral DNA polymerase encoded by UL54 (Pari, Kacica et al. 1993; Ripalti, Boccuni et al. 1995). pUL44 increases processivity of the polymerase along the viral DNA template (Ertl and Powell 1992; Weiland, Oien et al. 1994; Zuccola, Filman et al. 2000). The accumulation of the pUL44 is to strikingly high levels at late times after infection (Stinski 1978; Geballe, Leach et al. 1986). Its late kinetics of transcription and the high level of expression suggest an additional important role for viral replication. pUL44 is phosphorylated by the viral UL97 protein