Bipartite DNA-binding region of the Epstein-Barr virus BMRF1 product essential for DNA polymerase accessory function

Kiehl, Anita; Dorsky, David I.

doi:10.1128/jvi.69.3.1669-1677.1995

Cited by 35 publications

(16 citation statements)

References 37 publications

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“…In this report, we demonstrate that the BMRF1 gene prod-uct, in addition to its role as the polymerase processivity factor (6,10,11,14,38,39,42,(62)(63)(64), activates expression of an essential oriLyt promoter, BHLF1. We also find that the BMRF1 gene product interacts directly with the BZLF1 gene product in vitro as well as in vivo.…”

mentioning

confidence: 73%

Functional and physical interactions between the Epstein-Barr virus (EBV) proteins BZLF1 and BMRF1: Effects on EBV transcription and lytic replication

Zhang

Dorsky

et al. 1996

J Virol

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The Epstein-Barr virus (EBV) proteins BZLF1 and BMRF1 are both essential for lytic EBV replication. BZLF1 is a transcriptional activator which binds directly to the lytic origin of replication (oriLyt) and plays a critical role in the disruption of viral latency. The BMRF1 protein is required for viral polymerase processivity. Here we demonstrate that the BMRF1 gene product functions as a transcriptional activator and has direct (as well as indirect) interactions with the BZLF1 gene product. The BMRF1 gene product activates an essential oriLyt promoter, BHLF1, but does not activate two other early EBV promoters (BMRF1 and BHRF1). Direct interaction between the BMRF1 and BZLF1 gene products requires the first 45 amino acids of BMRF1 and the bZip domain of BZLF1. The effect of the BZLF1-BMRF1 interaction on early EBV transcription is complex and is promoter specific. The oriLyt BHLF1 promoter is activated by either the BZLF1 or BMRF1 gene product alone and is further activated by the combination of the BZLF1 and BMRF1 gene products. Enhanced activation of BHLF1 transcription by the BMRF1-BZLF1 combination does not require direct interaction between these proteins. In contrast, BZLF1-induced activation of the BMRF1 promoter is inhibited in the presence of the BMRF1 gene product. A point mutation in the BZLF1 protein (amino acid 200), which prevents in vitro interaction with the BMRF1 protein but which does not reduce BZLF1 transactivator function, allows the BZLF1 protein to activate the BMRF1 promoter equally well in the presence or absence of the BMRF1 gene product. Therefore, direct interaction between the BZLF1 and BMRF1 proteins may inhibit BZLF1-induced transcription of the BMRF1 promoter. BZLF1 mutated at amino acid 200 is as efficient as wild-type BZLF1 in promoting replication of an oriLyt plasmid. However, this mutation reduces the ability of BZLF1 to induce lytic replication of the endogenous viral genome in D98/HE-R-1 cells. Our results indicate that functional and physical interactions between the BMRF1 and BZLF1 proteins may modulate the efficiency of lytic EBV infection. The BMRF1 gene product clearly has a transcriptional, as well as replicative, role during lytic EBV infection.

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mentioning

confidence: 73%

Functional and physical interactions between the Epstein-Barr virus (EBV) proteins BZLF1 and BMRF1: Effects on EBV transcription and lytic replication

Zhang

Dorsky

et al. 1996

J Virol

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“…The polymerase and polymerase accessory protein (BMRF1) interact and together mediate processive DNA replication with strand displacement in model replication systems (33,36,43,68,70). BMRF1 binds DNA nonspecifically (32). Further, BMRF1 has been found to interact with the bZip domain of Zta (75) and to function independently as a transcriptional activator in transient expression assays (52,75).…”

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confidence: 99%

The Epstein-Barr Virus Lytic Transactivator Zta Interacts with the Helicase-Primase Replication Proteins

et al. 1998

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The Epstein-Barr virus transactivator Zta triggers lytic gene expression and is essential for replication of the lytic origin, oriLyt. Previous analysis indicated that the Zta activation domain contributed a replication-specific function. We now show that the Zta activation domain interacts with components of the EBV helicase-primase complex. The three helicase-primase proteins BBLF4 (helicase), BSLF1 (primase), and BBLF2/3 (primase-associated factor) were expressed fused to the Myc epitope. When expression plasmids for BBLF4 or BBLF2/3 plus BSLF1 (primase subcomplex) were separately transfected, the proteins localized to the cytoplasm. Interaction between Zta and the components of the helicase-primase complex was tested by examining the ability of Zta to alter the intracellular localization of these proteins. Cotransfection of Zta with Myc-BBLF4 resulted in nuclear translocation of Myc-BBLF4; similarly, cotransfection of Zta with the primase subcomplex led to nuclear translocation of the Myc-BSLF1 and Myc-BBLF2/3 proteins. This relocalization provides evidence for an interaction between Zta and the helicase and Zta and the primase subcomplex. An affinity assay using glutathioneS-transferase–Zta fusion proteins demonstrated that Myc-BBLF4 and Myc-BBLF2/3 plus BSLF1 bound to the Zta activation domain (amino acids 1 to 133). In the nuclear relocalization assay, the amino-terminal 25 amino acids of Zta were required for efficient interaction with the primase subcomplex but not for interaction with BBLF4. Evidence for interaction between oriLyt bound Zta and the helicase-primase complex was obtained in a superactivation assay using an oriLyt-chloramphenicol acetyltransferase (CAT) reporter. Zta activated expression from a CAT reporter containing the complete oriLyt region and regulated by the oriLyt BHLF1 promoter. Cotransfection of the helicase-primase proteins, one of which was fused to a heterologous activation domain, led to Zta-dependent superactivation of CAT expression. This assay also provided evidence for an interaction between the single-stranded DNA binding protein, BALF2, and the Zta-tethered helicase-primase complex. The helicase-primase interaction is consistent with a role for Zta in stabilizing the formation of an origin-bound replication complex.

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“…However, this does not preclude the possibility that BMRF2 is expressed from the BMRF1 promoter as well. It is interesting to note that the DNA binding and polymerase processivity domains of BMRF1 are located within the first 900 bp (2,15) BMRF1 are required for BHLF1 promoter activation and nuclear localization (38). Our data show that this 3Ј region (the last 360 bp of BMRF1) includes the functional promoter for BMRF2.…”

Section: Discussionmentioning

confidence: 66%

“…These include the BHLF1 (oriLyt) promoter, the BNLF1 (LMP1) promoter, and the promoter for LMP1 3.5-kb transcript (15,25,33,39). The LMP1 3.5-kb transcript is the major transcript in HL (15) and is also expressed in the epithelial cells of NPC (33). The promoter for the LMP1 3.5-kb transcript is located in the terminal repeat region and contains four Sp1 sites.…”

Section: Discussionmentioning

confidence: 99%

Regulation of Epstein-Barr Virus Promoters in Oral Epithelial Cells and Lymphocytes

Lagenaur

Palefsky

1999

J Virol

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Hairy leukoplakia (HL) is a proliferative lesion of the tongue that supports abundant Epstein-Barr virus (EBV) replication. Previous work showed high-level expression of the EBV BMRF2 gene in HL. To characterize the regulation of BMRF2 expression in HL, we mapped the 5′ ends of the BMRF1 and BMRF2 transcripts and showed that BMRF2 is expressed from a novel internal promoter within the BMRF1 coding region. Mechanisms of BMRF2 regulation were compared in oral epithelial cells and B lymphocytes, as were those of BMRF1 and BDLF3, early and late EBV transcripts, respectively, that are also known to be expressed in HL. Basal activity of the putative BMRF2 promoter was 10-fold higher in HSC-3 epithelial cells than in B lymphocytes. The BMRF2 and the BDLF3 promoters were responsive to induction by phorbol ester, but unlike the BMRF1 promoter, they were not responsive to BZLF1 transactivation. By mutational analysis, the major activity of the BMRF2 promoter mapped to a 50-bp region, which includes a TATA-like element and a GC box. The BMRF2 promoter may be regulated differentially from the BMRF1 promoter and more closely resembles that of BDLF3. This novel BMRF2 promoter likely belongs to a class of viral promoters that is more responsive to mechanisms known to induce epithelial cell differentiation, consistent with its high level of expression in HL.

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Bipartite DNA-binding region of the Epstein-Barr virus BMRF1 product essential for DNA polymerase accessory function

Cited by 35 publications

References 37 publications

Functional and physical interactions between the Epstein-Barr virus (EBV) proteins BZLF1 and BMRF1: Effects on EBV transcription and lytic replication

Functional and physical interactions between the Epstein-Barr virus (EBV) proteins BZLF1 and BMRF1: Effects on EBV transcription and lytic replication

The Epstein-Barr Virus Lytic Transactivator Zta Interacts with the Helicase-Primase Replication Proteins

Regulation of Epstein-Barr Virus Promoters in Oral Epithelial Cells and Lymphocytes

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