This Article Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Reprints and Permissions Copyright Information Books from ASM Press MicrobeWorld Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Sarisky, R. T. Articles by Hayward, G. S. Search for Related Content PubMed PubMed Citation Articles by Sarisky, R. T. Articles by Hayward, G. S.

 Previous Article  |  Next Article 

J. Virol., 11 1996, 7398-7413, Vol 70, No. 11
Copyright © 1996, American Society for Microbiology

Evidence that the UL84 gene product of human cytomegalovirus is essential for promoting oriLyt-dependent DNA replication and formation of replication compartments in cotransfection assays

RT Sarisky and GS Hayward
Molecular Virology Laboratories, Department of Pharmacology and Molecular Sciences, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205-2185, USA.

The protein products of 11 viral genomic loci cooperate in a transient cotransfection assay to mediate lytic-phase DNA replication of oriLyt, the human cytomegalovirus (HCMV) origin of replication. Six of these genes have homology with the well-characterized herpes simplex virus replication genes and encode core replication machinery proteins that are typically essential for DNA synthesis. The remaining five HCMV gene loci, initially referred to as auxiliary components, include several known immediate-early (IE) transcriptional regulatory proteins as well as genes encoding functionally uncharacterized polypeptides. Some or all of the auxiliary components may be necessary in trans to replicate the HCMV oriLyt only because they are required for efficient expression or transactivation of the native early promoters and 3' processing elements included in the genomic clones. Therefore, we reassessed the requirements for the auxiliary components by adding constitutive heterologous promoters and control signals to the coding regions and carrying out transient DpnI replication assays in cotransfected Vero cells. The results revealed that in the presence of the UL69 posttranscriptional activator and the remaining auxiliary polypeptides, UL84 was the only auxiliary component that could not be omitted to obtain oriLyt-dependent DNA replication. Nevertheless, in human diploid fibroblasts, some additional auxiliary loci as well as UL84 were critical. There was also an obligatory requirement for UL84, in cooperation with two other auxiliary factors, UL112-113 and IE2, and the core machinery, to constitute the minimal HCMV proteins necessary to direct oriLyt-dependent DNA amplification. However, the Epstein-Barr virus core replication genes could substitute for the HCMV core genes, and in these circumstances, UL84 alone directed amplification of HCMV oriLyt. Moreover, there was also an absolute requirement for UL84 along with the core and other auxiliary factors for the formation of intranuclear replication compartments as assayed by immunofluorescence in transient DNA cotransfection assays. These compartments were typical of those associated with active viral DNA replication in HCMV-infected cells, they incorporated pulse-labeled bromodeoxyuridine, and their formation was both phosphonoacetic acid sensitive and oriLyt dependent. These results demonstrate that UL84 is obligatory for both intranuclear replication compartment formation and origin-dependent DNA amplification and suggest that it is a key viral component in promoting the initiation of HCMV oriLyt-directed DNA replication.

This article has been cited by other articles:

• Kalejta, R. F. (2008). Tegument Proteins of Human Cytomegalovirus. Microbiol. Mol. Biol. Rev. 72: 249-265 [Abstract] [Full Text]  
• Qian, Z., Xuan, B., Hong, T. T., Yu, D. (2008). The Full-Length Protein Encoded by Human Cytomegalovirus Gene UL117 Is Required for the Proper Maturation of Viral Replication Compartments. J. Virol. 82: 3452-3465 [Abstract] [Full Text]  
• Sourvinos, G., Tavalai, N., Berndt, A., Spandidos, D. A., Stamminger, T. (2007). Recruitment of Human Cytomegalovirus Immediate-Early 2 Protein onto Parental Viral Genomes in Association with ND10 in Live-Infected Cells. J. Virol. 81: 10123-10136 [Abstract] [Full Text]  
• Colletti, K. S., Smallenburg, K. E., Xu, Y., Pari, G. S. (2007). Human Cytomegalovirus UL84 Interacts with an RNA Stem-Loop Sequence Found within the RNA/DNA Hybrid Region of oriLyt. J. Virol. 81: 7077-7085 [Abstract] [Full Text]  
• White, E. A., Del Rosario, C. J., Sanders, R. L., Spector, D. H. (2007). The IE2 60-Kilodalton and 40-Kilodalton Proteins Are Dispensable for Human Cytomegalovirus Replication but Are Required for Efficient Delayed Early and Late Gene Expression and Production of Infectious Virus. J. Virol. 81: 2573-2583 [Abstract] [Full Text]  
• Wang, Y., Tang, Q., Maul, G. G., Yuan, Y. (2006). Kaposi's Sarcoma-Associated Herpesvirus ori-Lyt-Dependent DNA Replication: Dual Role of Replication and Transcription Activator. J. Virol. 80: 12171-12186 [Abstract] [Full Text]  
• Lischka, P., Rauh, C., Mueller, R., Stamminger, T. (2006). Human Cytomegalovirus UL84 Protein Contains Two Nuclear Export Signals and Shuttles between the Nucleus and the Cytoplasm.. J. Virol. 80: 10274-10280 [Abstract] [Full Text]  
• Park, M.-Y., Kim, Y.-E., Seo, M.-R., Lee, J.-R., Lee, C. H., Ahn, J.-H. (2006). Interactions among Four Proteins Encoded by the Human Cytomegalovirus UL112-113 Region Regulate Their Intranuclear Targeting and the Recruitment of UL44 to Prereplication Foci. J. Virol. 80: 2718-2727 [Abstract] [Full Text]  
• Davison, A. J., Stow, N. D. (2005). New Genes from Old: Redeployment of dUTPase by Herpesviruses. J. Virol. 79: 12880-12892 [Abstract] [Full Text]  
• Liao, G., Huang, J., Fixman, E. D., Hayward, S. D. (2005). The Epstein-Barr Virus Replication Protein BBLF2/3 Provides an Origin-Tethering Function through Interaction with the Zinc Finger DNA Binding Protein ZBRK1 and the KAP-1 Corepressor. J. Virol. 79: 245-256 [Abstract] [Full Text]  
• Xu, Y., Cei, S. A., Rodriguez Huete, A., Colletti, K. S., Pari, G. S. (2004). Human Cytomegalovirus DNA Replication Requires Transcriptional Activation via an IE2- and UL84-Responsive Bidirectional Promoter Element within oriLyt. J. Virol. 78: 11664-11677 [Abstract] [Full Text]  
• Xu, Y., Cei, S. A., Huete, A. R., Pari, G. S. (2004). Human Cytomegalovirus UL84 Insertion Mutant Defective for Viral DNA Synthesis and Growth. J. Virol. 78: 10360-10369 [Abstract] [Full Text]  
• Colletti, K. S., Xu, Y., Cei, S. A., Tarrant, M., Pari, G. S. (2004). Human Cytomegalovirus UL84 Oligomerization and Heterodimerization Domains Act as Transdominant Inhibitors of oriLyt-Dependent DNA Replication: Evidence that IE2-UL84 and UL84-UL84 Interactions Are Required for Lytic DNA Replication. J. Virol. 78: 9203-9214 [Abstract] [Full Text]  
• Nevels, M., Brune, W., Shenk, T. (2004). SUMOylation of the Human Cytomegalovirus 72-Kilodalton IE1 Protein Facilitates Expression of the 86-Kilodalton IE2 Protein and Promotes Viral Replication. J. Virol. 78: 7803-7812 [Abstract] [Full Text]  
• Rosenke, K., Fortunato, E. A. (2004). Bromodeoxyuridine-Labeled Viral Particles as a Tool for Visualization of the Immediate-Early Events of Human Cytomegalovirus Infection. J. Virol. 78: 7818-7822 [Abstract] [Full Text]  
• Lischka, P., Sorg, G., Kann, M., Winkler, M., Stamminger, T. (2003). A Nonconventional Nuclear Localization Signal within the UL84 Protein of Human Cytomegalovirus Mediates Nuclear Import via the Importin {alpha}/{beta} Pathway. J. Virol. 77: 3734-3748 [Abstract] [Full Text]  
• Ellsmore, V., Reid, G. G., Stow, N. D. (2003). Detection of human cytomegalovirus DNA replication in non-permissive Vero and 293 cells. J. Gen. Virol. 84: 639-645 [Abstract] [Full Text]  
• Xu, Y., Colletti, K. S., Pari, G. S. (2002). Human Cytomegalovirus UL84 Localizes to the Cell Nucleus via a Nuclear Localization Signal and Is a Component of Viral Replication Compartments. J. Virol. 76: 8931-8938 [Abstract] [Full Text]  
• Gawn, J. M., Greaves, R. F. (2002). Absence of IE1 p72 Protein Function during Low-Multiplicity Infection by Human Cytomegalovirus Results in a Broad Block to Viral Delayed-Early Gene Expression. J. Virol. 76: 4441-4455 [Abstract] [Full Text]  
• Shirakata, M., Terauchi, M., Ablikim, M., Imadome, K.-I., Hirai, K., Aso, T., Yamanashi, Y. (2002). Novel Immediate-Early Protein IE19 of Human Cytomegalovirus Activates the Origin Recognition Complex I Promoter in a Cooperative Manner with IE72. J. Virol. 76: 3158-3167 [Abstract] [Full Text]  
• Xu, Y., Ahn, J.-H., Cheng, M., apRhys, C. M., Chiou, C.-J., Zong, J., Matunis, M. J., Hayward, G. S. (2001). Proteasome-Independent Disruption of PML Oncogenic Domains (PODs), but Not Covalent Modification by SUMO-1, Is Required for Human Cytomegalovirus Immediate-Early Protein IE1 To Inhibit PML-Mediated Transcriptional Repression. J. Virol. 75: 10683-10695 [Abstract] [Full Text]  
• Liao, G., Wu, F. Y., Hayward, S. D. (2001). Interaction with the Epstein-Barr Virus Helicase Targets Zta to DNA Replication Compartments. J. Virol. 75: 8792-8802 [Abstract] [Full Text]  
• Huang, J., Levin, D. B. (2001). Expression, purification and characterization of the Spodoptera littoralis nucleopolyhedrovirus (SpliNPV) DNA polymerase and interaction with the SpliNPV non-hr origin of DNA replication. J. Gen. Virol. 82: 1767-1776 [Abstract] [Full Text]  
• Bahr, U., Darai, G. (2001). Analysis and Characterization of the Complete Genome of Tupaia (Tree Shrew) Herpesvirus. J. Virol. 75: 4854-4870 [Abstract] [Full Text]  
• Johnson, R. A., Ma, X.-L., Yurochko, A. D., Huang, E.-S. (2001). The role of MKK1/2 kinase activity in human cytomegalovirus infection. J. Gen. Virol. 82: 493-497 [Abstract] [Full Text]  
• Wu, F. Y., Ahn, J.-H., Alcendor, D. J., Jang, W.-J., Xiao, J., Hayward, S. D., Hayward, G. S. (2001). Origin-Independent Assembly of Kaposi's Sarcoma-Associated Herpesvirus DNA Replication Compartments in Transient Cotransfection Assays and Association with the ORF-K8 Protein and Cellular PML. J. Virol. 75: 1487-1506 [Abstract] [Full Text]  
• Greijer, A. E., Dekkers, C. A. J., Middeldorp, J. M. (2000). Human Cytomegalovirus Virions Differentially Incorporate Viral and Host Cell RNA during the Assembly Process. J. Virol. 74: 9078-9082 [Abstract] [Full Text]  
• Giesen, K., Radsak, K., Bogner, E. (2000). The potential terminase subunit of human cytomegalovirus, pUL56, is translocated into the nucleus by its own nuclear localization signal and interacts with importin {alpha}. J. Gen. Virol. 81: 2231-2244 [Abstract] [Full Text]  
• Winkler, M., aus dem Siepen, T., Stamminger, T. (2000). Functional Interaction between Pleiotropic Transactivator pUL69 of Human Cytomegalovirus and the Human Homolog of Yeast Chromatin Regulatory Protein SPT6. J. Virol. 74: 8053-8064 [Abstract] [Full Text]  
• Sinclair, J., Baillie, J., Bryant, L., Caswell, R. (2000). Human cytomegalovirus mediates cell cycle progression through G1 into early S phase in terminally differentiated cells. J. Gen. Virol. 81: 1553-1565 [Abstract] [Full Text]  
• Ahn, J.-H., Jang, W.-J., Hayward, G. S. (1999). The Human Cytomegalovirus IE2 and UL112-113 Proteins Accumulate in Viral DNA Replication Compartments That Initiate from the Periphery of Promyelocytic Leukemia Protein-Associated Nuclear Bodies (PODs or ND10). J. Virol. 73: 10458-10471 [Abstract] [Full Text]  
• Morello, C. S., Cranmer, L. D., Spector, D. H. (1999). In Vivo Replication, Latency, and Immunogenicity of Murine Cytomegalovirus Mutants with Deletions in the M83 and M84 Genes, the Putative Homologs of Human Cytomegalovirus pp65 (UL83). J. Virol. 73: 7678-7693 [Abstract] [Full Text]  
• Huang, J., Levin, D. B. (1999). Identification and functional analysis of a putative non-hr origin of DNA replication from the Spodoptera littoralis type B multinucleocapsid nucleopolyhedrovirus. J. Gen. Virol. 80: 2263-2274 [Abstract] [Full Text]  
• Gao, Z., Krithivas, A., Finan, J. E., Semmes, O. J., Zhou, S., Wang, Y., Hayward, S. D. (1998). The Epstein-Barr Virus Lytic Transactivator Zta Interacts with the Helicase-Primase Replication Proteins. J. Virol. 72: 8559-8567 [Abstract] [Full Text]  
• Goldstein, J. N., Weller, S. K. (1998). In Vitro Processing of Herpes Simplex Virus Type 1 DNA Replication Intermediates by the Viral Alkaline Nuclease, UL12. J. Virol. 72: 8772-8781 [Abstract] [Full Text]  
• Prichard, M. N., Jairath, S., Penfold, M. E. T., St. Jeor, S., Bohlman, M. C., Pari, G. S. (1998). Identification of Persistent RNA-DNA Hybrid Structures within the Origin of Replication of Human Cytomegalovirus. J. Virol. 72: 6997-7004 [Abstract] [Full Text]  
• Biegalke, B. J. (1998). Characterization of the Transcriptional Repressive Element of the Human Cytomegalovirus Immediate-Early US3 Gene. J. Virol. 72: 5457-5463 [Abstract] [Full Text]  
• Sanchez, V., Angeletti, P. C., Engler, J. A., Britt, W. J. (1998). Localization of Human Cytomegalovirus Structural Proteins to the Nuclear Matrix of Infected Human Fibroblasts. J. Virol. 72: 3321-3329 [Abstract] [Full Text]  
• Greaves, R. F., Mocarski, E. S. (1998). Defective Growth Correlates with Reduced Accumulation of a Viral DNA Replication Protein after Low-Multiplicity Infection by a Human Cytomegalovirus ie1 Mutant. J. Virol. 72: 366-379 [Abstract] [Full Text]