Biomaterial Database

The Rad6/18 ubiquitin complex interacts with the Epstein-Barr virus deubiquitinating enzyme, BPLF1, and contributes to virus infectivity. 2014

Ravindra Kumar, and Christopher B Whitehurst, and Joseph S Pagano

Lineberger Comprehensive Cancer Center, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, USA.

Epstein-Barr virus (EBV) encodes BPLF1, a lytic cycle protein with deubiquitinating activity that is contained in its N-terminal domain and conserved across the Herpesviridae. EBV replication is associated with cellular DNA replication and repair factors, and initiation of EBV lytic replication induces a DNA damage response, which can be regulated at least in part by BPLF1. The cellular DNA repair pathway, translesion synthesis (TLS), is disrupted by BPLF1, which deubiquitinates the DNA processivity factor, PCNA, and inhibits the recruitment of the TLS polymerase, polymerase eta (Pol eta), after damage to DNA by UV irradiation. Here we showed that the E3 ubiquitin ligase, which activates TLS repair by monoubiquitination of PCNA, is also affected by BPLF1 deubiquitinating activity. First, BPLF1 interacts directly with Rad18, and overexpression of BPLF1 results in increased levels of the Rad18 protein, suggesting that it stabilizes Rad18. Next, expression of functionally active BPLF1 caused relocalization of Rad18 into nuclear foci, which is consistent with sites of cellular DNA replication that occur during S phase. Also, levels of Rad18 remain constant during lytic reactivation of wild-type virus, but reactivation of BPLF1 knockout virus resulted in decreased levels of Rad18. Finally, the contribution of Rad18 levels to infectious virus production was examined with small interfering RNA (siRNA) targeting Rad18. Results demonstrated that reducing levels of Rad18 decreased production of infectious virus, and infectious titers of BPLF1 knockout virus were partially restored by overexpression of Rad18. Thus, BPLF1 interacts with and maintains Rad18 at high levels during lytic replication, which assists in production of infectious virus. OBJECTIVE Characterization of EBV BPLF1's deubiquitinating activity and identification of its targets and subsequent functional effects remain little studied. All members of the Herpesviridae contain BPLF1 homologs with conserved enzymatic activity, and findings discovered with EBV BPLF1 are likely applicable to other members of the family. Discovery of new targets of BPLF1 will point to cellular pathways and viral processes regulated by the enzymatic activity of the EBV-encoded deubiquitinating enzyme. Here we determined the importance of the cellular ubiquitin ligase Rad18 in these processes and how it is affected by BPLF1. Our findings demonstrate that EBV can co-opt Rad18 as a novel accessory factor in the production of infectious virus.

UI	MeSH Term	Description	Entries
D004268	DNA-Binding Proteins	Proteins which bind to DNA. The family includes proteins which bind to both double- and single-stranded DNA and also includes specific DNA binding proteins in serum which can be used as markers for malignant diseases.	DNA Helix Destabilizing Proteins,DNA-Binding Protein,Single-Stranded DNA Binding Proteins,DNA Binding Protein,DNA Single-Stranded Binding Protein,SS DNA BP,Single-Stranded DNA-Binding Protein,Binding Protein, DNA,DNA Binding Proteins,DNA Single Stranded Binding Protein,DNA-Binding Protein, Single-Stranded,Protein, DNA-Binding,Single Stranded DNA Binding Protein,Single Stranded DNA Binding Proteins
D004854	Herpesvirus 4, Human	The type species of LYMPHOCRYPTOVIRUS, subfamily GAMMAHERPESVIRINAE, infecting B-cells in humans. It is thought to be the causative agent of INFECTIOUS MONONUCLEOSIS and is strongly associated with oral hairy leukoplakia (LEUKOPLAKIA, HAIRY;), BURKITT LYMPHOMA; and other malignancies.	Burkitt Herpesvirus,Burkitt Lymphoma Virus,E-B Virus,EBV,Epstein-Barr Virus,Human Herpesvirus 4,Infectious Mononucleosis Virus,Burkitt's Lymphoma Virus,HHV-4,Herpesvirus 4 (gamma), Human,Burkitts Lymphoma Virus,E B Virus,E-B Viruses,Epstein Barr Virus,Herpesvirus, Burkitt,Infectious Mononucleosis Viruses,Lymphoma Virus, Burkitt,Mononucleosis Virus, Infectious,Mononucleosis Viruses, Infectious
D004926	Escherichia coli	A species of gram-negative, facultatively anaerobic, rod-shaped bacteria (GRAM-NEGATIVE FACULTATIVELY ANAEROBIC RODS) commonly found in the lower part of the intestine of warm-blooded animals. It is usually nonpathogenic, but some strains are known to produce DIARRHEA and pyogenic infections. Pathogenic strains (virotypes) are classified by their specific pathogenic mechanisms such as toxins (ENTEROTOXIGENIC ESCHERICHIA COLI), etc.	Alkalescens-Dispar Group,Bacillus coli,Bacterium coli,Bacterium coli commune,Diffusely Adherent Escherichia coli,E coli,EAggEC,Enteroaggregative Escherichia coli,Enterococcus coli,Diffusely Adherent E. coli,Enteroaggregative E. coli,Enteroinvasive E. coli,Enteroinvasive Escherichia coli
D005455	Fluorescent Antibody Technique	Test for tissue antigen using either a direct method, by conjugation of antibody with fluorescent dye (FLUORESCENT ANTIBODY TECHNIQUE, DIRECT) or an indirect method, by formation of antigen-antibody complex which is then labeled with fluorescein-conjugated anti-immunoglobulin antibody (FLUORESCENT ANTIBODY TECHNIQUE, INDIRECT). The tissue is then examined by fluorescence microscopy.	Antinuclear Antibody Test, Fluorescent,Coon's Technique,Fluorescent Antinuclear Antibody Test,Fluorescent Protein Tracing,Immunofluorescence Technique,Coon's Technic,Fluorescent Antibody Technic,Immunofluorescence,Immunofluorescence Technic,Antibody Technic, Fluorescent,Antibody Technics, Fluorescent,Antibody Technique, Fluorescent,Antibody Techniques, Fluorescent,Coon Technic,Coon Technique,Coons Technic,Coons Technique,Fluorescent Antibody Technics,Fluorescent Antibody Techniques,Fluorescent Protein Tracings,Immunofluorescence Technics,Immunofluorescence Techniques,Protein Tracing, Fluorescent,Protein Tracings, Fluorescent,Technic, Coon's,Technic, Fluorescent Antibody,Technic, Immunofluorescence,Technics, Fluorescent Antibody,Technics, Immunofluorescence,Technique, Coon's,Technique, Fluorescent Antibody,Technique, Immunofluorescence,Techniques, Fluorescent Antibody,Techniques, Immunofluorescence,Tracing, Fluorescent Protein,Tracings, Fluorescent Protein
D005786	Gene Expression Regulation	Any of the processes by which nuclear, cytoplasmic, or intercellular factors influence the differential control (induction or repression) of gene action at the level of transcription or translation.	Gene Action Regulation,Regulation of Gene Expression,Expression Regulation, Gene,Regulation, Gene Action,Regulation, Gene Expression
D006801	Humans	Members of the species Homo sapiens.	Homo sapiens,Man (Taxonomy),Human,Man, Modern,Modern Man
D014779	Virus Replication	The process of intracellular viral multiplication, consisting of the synthesis of PROTEINS; NUCLEIC ACIDS; and sometimes LIPIDS, and their assembly into a new infectious particle.	Viral Replication,Replication, Viral,Replication, Virus,Replications, Viral,Replications, Virus,Viral Replications,Virus Replications
D015151	Immunoblotting	Immunologic method used for detecting or quantifying immunoreactive substances. The substance is identified by first immobilizing it by blotting onto a membrane and then tagging it with labeled antibodies.	Dot Immunoblotting,Electroimmunoblotting,Immunoelectroblotting,Reverse Immunoblotting,Immunoblotting, Dot,Immunoblotting, Reverse,Dot Immunoblottings,Electroimmunoblottings,Immunoblottings,Immunoblottings, Dot,Immunoblottings, Reverse,Immunoelectroblottings,Reverse Immunoblottings
D044763	Ubiquitin-Conjugating Enzymes	A class of enzymes that form a thioester bond to UBIQUITIN with the assistance of UBIQUITIN-ACTIVATING ENZYMES. They transfer ubiquitin to the LYSINE of a substrate protein with the assistance of UBIQUITIN-PROTEIN LIGASES.	Ubiquitin-Conjugating Enzyme,HHR6 Protein,Ubiquitin-Conjugating Enzyme E2,E2, Ubiquitin-Conjugating Enzyme,Enzyme E2, Ubiquitin-Conjugating,Enzyme, Ubiquitin-Conjugating,Enzymes, Ubiquitin-Conjugating,Ubiquitin Conjugating Enzyme,Ubiquitin Conjugating Enzyme E2,Ubiquitin Conjugating Enzymes
D044767	Ubiquitin-Protein Ligases	A diverse class of enzymes that interact with UBIQUITIN-CONJUGATING ENZYMES and ubiquitination-specific protein substrates. Each member of this enzyme group has its own distinct specificity for a substrate and ubiquitin-conjugating enzyme. Ubiquitin-protein ligases exist as both monomeric proteins multiprotein complexes.	Ubiquitin-Protein Ligase,E3 Ligase,E3 Ubiquitin Ligase,Ubiquitin Ligase E3,Ubiquitin-Protein Ligase E3,Ligase E3, Ubiquitin,Ligase E3, Ubiquitin-Protein,Ligase, E3,Ligase, E3 Ubiquitin,Ligase, Ubiquitin-Protein,Ligases, Ubiquitin-Protein,Ubiquitin Ligase, E3,Ubiquitin Protein Ligase,Ubiquitin Protein Ligase E3,Ubiquitin Protein Ligases