Identification of cellular proteins associated with human cytomegalovirus (HCMV) DNA replication suggests novel cellular and viral interactions

In the nuclei of permissive cells, human cytomegalovirus genomes form nucleosomal structures initially resembling heterochromatin but gradually switching to a euchromatin-like state. This switch is characterized by a decrease in histone H3 K9 methylation and a marked increase in H3 tail acetylation and H3 K4 methylation across the viral genome. We used ganciclovir and a mutant virus encoding a reversibly destabilized DNA polymerase to examine the impact of DNA replication on histone modification dynamics at the viral chromatin. The changes in H3 tail acetylation and H3 K9 methylation proceeded in a DNA replication-independent fashion. In contrast, the increase in H3 K4 methylation proved to depend widely on viral DNA synthesis. Consistently, labeling of nascent DNA using “click chemistry” revealed preferential incorporation of methylated H3 K4 into viral (but not cellular) chromatin during or following DNA replication. This study demonstrates largely selective epigenetic tagging of postreplicative human cytomegalovirus chromatin.

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Identification of cellular proteins required for simian virus 40 DNA replication

Journal of Biological Chemistry ◽

10.1016/s0021-9258(19)81684-8 ◽

1989 ◽

Vol 264 (5) ◽

pp. 2801-2809

Author(s):

M S Wold ◽

D H Weinberg ◽

D M Virshup ◽

J J Li ◽

T J Kelly

Keyword(s):

Dna Replication ◽

Simian Virus 40 ◽

Simian Virus ◽

Cellular Proteins

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Kaposi's Sarcoma-Associated Herpesvirus ori-Lyt-Dependent DNA Replication: Involvement of Host Cellular Factors

Journal of Virology ◽

10.1128/jvi.01319-07 ◽

2008 ◽

Vol 82 (6) ◽

pp. 2867-2882 ◽

Cited By ~ 57

Author(s):

Yan Wang ◽

Hong Li ◽

Qiyi Tang ◽

Gerd G. Maul ◽

Yan Yuan

Keyword(s):

Dna Replication ◽

Viral Replication ◽

Kaposi's Sarcoma ◽

Kaposi’S Sarcoma ◽

Cellular Proteins ◽

Cellular Factors ◽

Virally Encoded ◽

Kaposi’S Sarcoma Associated Herpesvirus ◽

Lytic Dna Replication ◽

Parp 1

ABSTRACT Herpesvirus lytic DNA replication requires both the cis-acting element, the origin, and trans-acting factors, including virally encoded origin-binding protein, DNA replication enzymes, and auxiliary factors. Two lytic DNA replication origins (ori-Lyt) of Kaposi's sarcoma-associated herpesvirus (KSHV) have been identified, and two virally encoded proteins, namely, RTA and K8, have been shown to bind to the origins. In this study, we sought to identify cellular factors that associate with ori-Lyt by using DNA affinity purification and mass spectrometry. This approach led to identification of several cellular proteins that bind to KSHV ori-Lyt. They include topoisomerases (Topo) I and II, MSH2/6, RecQL, poly(ADP-ribose) polymerase I (PARP-1), DNA-PK, Ku86/70 autoantigens, and scaffold attachment factor A (SAF-A). RecQL appears to associate with prereplication complexes and be recruited to ori-Lyt through RTA and K8. Topoisomerases, MSH2, PARP-1, DNA-PK, and Ku86 were not detected in prereplication complexes but were present in replication initiation complexes on ori-Lyt. All these cellular proteins accumulate in viral replication compartments in the nucleus, indicating that these proteins may have a role in viral replication. Topo I and II appear to be essential for viral DNA replication as inhibition of their activities with specific inhibitors (camptothecin and ellipticine) blocked ori-Lyt-dependent DNA replication. Furthermore, inhibition of PARP-1 with chemical inhibitors (3-aminobenzamide and niacinamide) resulted in decreased ori-Lyt-dependent DNA replication, whereas hydroxyurea, which raises PARP-1 activity, caused an increase in the DNA replication, suggesting a positive role for PARP-1 in KSHV lytic DNA replication.

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Interaction of the Putative Human Cytomegalovirus Portal Protein pUL104 with the Large Terminase Subunit pUL56 and Its Inhibition by Benzimidazole-d-Ribonucleosides

Journal of Virology ◽

10.1128/jvi.79.23.14660-14667.2005 ◽

2005 ◽

Vol 79 (23) ◽

pp. 14660-14667 ◽

Cited By ~ 48

Author(s):

Alexandra Dittmer ◽

John C. Drach ◽

Leroy B. Townsend ◽

Anke Fischer ◽

Elke Bogner

Keyword(s):

Dna Replication ◽

Human Cytomegalovirus ◽

Unit Length ◽

Large Subunit ◽

Intracellular Localization ◽

Direct Interaction ◽

Portal Protein ◽

Specific Association ◽

Carboxy Terminal

ABSTRACT Herpesvirus DNA replication leads to unit length genomes that are translocated into preformed procapsids through a unique portal vertex. The translocation is performed by the terminase that cleaves the DNA and powers the insertion by its ATPase activity. Recently, we demonstrated that the putative human cytomegalovirus (HCMV) portal protein, pUL104, also forms high-molecular-weight complexes. Analyses now have been performed to determine the intracellular localization and identification of interaction partners of pUL104. In infected cells, HCMV pUL104 was found to be predominantly localized throughout the nucleus as well as in cytoplasmic clusters at late times of infection. The latter localization was abolished by phosphonoacetic acid, an inhibitor of viral DNA replication. Immunofluorescence revealed that pUL104 colocalized with pUL56, the large subunit of the HCMV terminase. Specific association of in vitro translated pUL104 with the carboxy-terminal half of GST-UL56C was detected. By using coimmunoprecipitations a direct interaction with pUL56 was confirmed. In addition, this interaction was no longer detected when the benzimidazole-d-nucleosides BDCRB or Cl4RB were added, thus indicating that these HCMV inhibitors block the insertion of the DNA into the capsid by preventing a necessary interaction of pUL56 with the portal. Electron microscopy revealed that in the presence of Cl4RB DNA is not packaged into capsids and these capsids failed to egress from the nucleus. Furthermore, pulsed-field gel electrophoresis showed that DNA concatemers synthesized in the presence of the compound failed to be processed.

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Infection of Cells with Human Cytomegalovirus during S Phase Results in a Blockade to Immediate-Early Gene Expression That Can Be Overcome by Inhibition of the Proteasome

Journal of Virology ◽

10.1128/jvi.76.11.5369-5379.2002 ◽

2002 ◽

Vol 76 (11) ◽

pp. 5369-5379 ◽

Cited By ~ 54

Author(s):

Elizabeth A. Fortunato ◽

Veronica Sanchez ◽

Judy Y. Yen ◽

Deborah H. Spector

Keyword(s):

Gene Expression ◽

Dna Replication ◽

Human Cytomegalovirus ◽

Dna Content ◽

Laser Scanning ◽

Small Population ◽

S Phase ◽

Early Gene ◽

Immediate Early ◽

Laser Scanning Cytometry

ABSTRACT Cells infected with human cytomegalovirus (HCMV) after commencing DNA replication do not initiate viral immediate-early (IE) gene expression and divide before arresting. To determine the nature of this blockade, we examined cells that were infected 24 h after release from G0 using immunofluorescence, laser scanning cytometry, and fluorescence-activated cell sorting (FACS) analysis. Approximately 40 to 50% of the cells had 2N DNA content, became IE+ in the first 12 h, and arrested. Most but not all of the cells with >2N DNA content did not express IE antigens until after mitosis. To define the small population of IE+ cells that gradually accumulated within the S and G2/M compartments, cells were pulsed with bromodeoxyuridine (BrdU) just prior to S-phase infection and analyzed at 12 h postinfection for IE gene expression, BrdU positivity, and cell cycle position. Most of the BrdU+ cells were IE− and had progressed into G2/M or back to G1. The majority of the IE+ cells in S and G2/M were BrdU−. Only a few cells were IE+ BrdU+, and they resided in G2/M. Multipoint BrdU pulse-labeling revealed that, compared to cells actively synthesizing DNA at the beginning of the infection, a greater percentage of the cells that initiated DNA replication 4 h later could express IE antigens and proceed into S. Synchronization of the cells with aphidicolin also indicated that the blockade to the activation of IE gene expression was established in cells soon after initiation of DNA replication. It appears that a short-lived protein in S-phase cells may be required for IE gene expression, as it is partially restored by treatment with the proteasome inhibitor MG132.

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pUL34 binding near the human cytomegalovirus origin of lytic replication enhances DNA replication and viral growth

Virology ◽

10.1016/j.virol.2018.03.017 ◽

2018 ◽

Vol 518 ◽

pp. 414-422 ◽

Cited By ~ 2

Author(s):

Mark Slayton ◽

Tanvir Hossain ◽

Bonita J. Biegalke

Keyword(s):

Dna Replication ◽

Human Cytomegalovirus ◽

Lytic Replication ◽

Viral Growth

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Human cytomegalovirus UL141 protein interacts with CELF5 and affects viral DNA replication

Molecular Medicine Reports ◽

10.3892/mmr.2018.8419 ◽

2018 ◽

Cited By ~ 2

Author(s):

Fei Zou ◽

Zhi‑Tao Lu ◽

Shuang Wang ◽

Si Wu ◽

Ying‑Ying Wu ◽

...

Keyword(s):

Dna Replication ◽

Human Cytomegalovirus ◽

Viral Dna ◽

Viral Dna Replication

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Effects of Tat proteins and Tat mutants of different human immunodeficiency virus type 1 clades on glial JC virus early and late gene transcription

Journal of General Virology ◽

10.1099/vir.0.047902-0 ◽

2013 ◽

Vol 94 (3) ◽

pp. 514-523 ◽

Cited By ~ 4

Author(s):

Clayton A. Wright ◽

Jonas A. Nance ◽

Edward M. Johnson

Keyword(s):

Human Immunodeficiency Virus ◽

Dna Replication ◽

Gene Transcription ◽

Cellular Proteins ◽

Late Gene ◽

Clade B ◽

Immunodeficiency Virus ◽

The Brain ◽

Hiv 1

Polyomavirus JC (JCV) is the aetiological agent of progressive multifocal leukoencephalopathy (PML), a frequently fatal infection of the brain afflicting nearly 4 % of AIDS patients in the USA. Human immunodeficiency virus type 1 (HIV-1) Tat, acting together with cellular proteins at the JCV non-coding control region (NCCR), can stimulate JCV DNA transcription and replication. Tat in the brain is secreted by HIV-1-infected cells and incorporated by oligodendroglia, cells capable of infection by JCV. Thus far the effects of Tat on JCV have been studied primarily with protein encoded by the HIV-1 B clade most common in North America. Here, we determine the abilities of Tat from different HIV-1 clades to alter JCV early and late gene transcription and DNA replication initiated at the JCV origin. Tat from all clades tested stimulates both JCV early and late gene promoters, with clade B Tat being significantly most effective. Tat proteins from the HIV-1 clades display parallel patterns of differences in their effects on HIV-1 and JCV transcription, suggesting that Tat effects in both cases are mediated by the same cellular proteins. Clade B Tat is most effective at directing Smad mediators of tumour growth factor beta and cellular partner Purα to the NCCR. Tat proteins from all non-B clades inhibit initiation of JCV DNA replication. The effectiveness of HIV-1 clade B Tat at promoting JCV transcriptional and replicative processes highlights a need for further investigation to determine which molecular aspects of Tat from distinct HIV-1 substrains can contribute to the course of PML development in neuroAIDS.

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