Identification of a Novel Element Involved in Regulation of the Lytic Switch BZLF1 Gene Promoter of Epstein-Barr Virus

Kraus, Richard J.; Mirocha, Sarah; Stephany, Heather M.; Puchalski, Joel R.; Mertz, Janet E.

doi:10.1128/jvi.75.2.867-877.2001

Cited by 44 publications

(67 citation statements)

References 61 publications

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“…These include the ZI, ZII, and ZIII elements, which have been tested in both luciferase and chloramphenicol acetyltransferase reporter systems and with the complete BZLF1 gene (3,14). Mutation of an additional element known as ZV was found to cause a large increase in the response of the Zp promoter to either tetradecanoyl phorbol (3,18,19). This ZV mutation had the largest effect of any Zp mutation tested in luciferase reporter assays, giving an about 8-to 10-fold increase in Zp activity after anti-Ig induction but not making the promoter constitutively active.…”

Section: Resultsmentioning

confidence: 99%

Lytic Cycle Gene Regulation of Epstein-Barr Virus

Amon

Binné

Bryant

et al. 2004

J Virol

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Episomal reporter plasmids containing the Epstein-Barr virus (EBV) oriP sequence stably transfected into Akata Burkitt's lymphoma cells were used to analyze EBV lytic cycle gene regulation. First, we found that the Zp promoter of EBV, but not the Rp promoter, can be activated in the absence of protein synthesis in these oriP plasmids, casting doubt on the immediate early status of Rp. An additional level of regulation of Zp was implied by analysis of a mutation of the ZV element. Second, our analysis of late lytic cycle promoters revealed that the correct relative timing, dependence on ori lyt in cis, and sensitivity to inhibitors of DNA replication were reconstituted on the oriP plasmids. Late promoter luciferase activity from oriP plasmids also incorporating replication-competent ori lyt was phosphonoacetic acid sensitive, a hallmark of EBV late genes. A minimal ori lyt, which only replicates weakly, was sufficient to confer late timing of expression specifically on late promoters. Finally, deletion analysis of EBV late promoter sequences upstream of the transcription start site confirmed that sequences between ؊49 and ؉30 are sufficient for late gene expression, which is dependent on ori lyt in cis. However, the TATT version of the TATA box found in many late genes was not essential for late expression.We recently described a system for studying Epstein-Barr virus (EBV) lytic gene regulation with plasmids stably transfected into the Akata Burkitt's lymphoma cell line (3). The plasmids contain the promoter being studied linked to the luciferase reporter gene and also have the EBV oriP plasmid origin of replication and a selectable marker (hygromycin resistance). Cell lines stably maintaining the plasmids, typically at about 10 copies per cell, can readily be isolated. Akata cells containing EBV can be induced into the lytic cycle by treatment with anti-immunoglobulin (anti-Ig), which cross-links the surface B-cell receptor, providing a rapid and physiologically relevant method for studying the EBV lytic cycle in cell culture (28). This system accurately reconstituted the regulation of the Zp immediate early promoter of EBV and was used to study its regulation in detail (3,5,14). The stably transfected reporter plasmid has an average copy number similar to that of the endogenous EBV genome present in the same cell, the plasmids are assembled into chromatin like EBV, and the reactivation of endogenous virus provides viral lytic gene products that may act in trans at their normal concentrations. These features make the system a good model for studying viral gene regulation, avoiding artifacts associated with transient transfection and incorrect expression levels of regulatory factors. In this study, we extended the use of the system to investigate the Rp promoter and EBV delayed early and late gene promoters.Zp and Rp have been considered to be the immediate early promoters of EBV, but controversy has surrounded the status of Rp, partly because of technical difficulties in mapping the 5Ј end of the RNA from this ...

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Section: Resultsmentioning

confidence: 99%

Lytic Cycle Gene Regulation of Epstein-Barr Virus

Amon

Binné

Bryant

et al. 2004

J Virol

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“…To examine the effects of ER ligands, cells were maintained in stripped medium for 48 h before transfection and the addition of 17␤-estradiol (E 2 ) (Sigma) or the pure anti-estrogen ICI-182,780 (Astra Zeneca, London, UK) dissolved in ethanol and diluted in medium to obtain the indicated concentrations. Cells were harvested 48 h post-transfection, and lysates were assayed for luciferase activity normalized to protein concentration as previously described (36 The numbers refer to the amino acid residues from the amino terminus. The percentages indicate the amino acid sequence similarity between the corresponding domains of the two proteins, with ER␣ sequences set at 100%.…”

Section: Methodsmentioning

confidence: 99%

Estrogen-related Receptor α1 Actively Antagonizes Estrogen Receptor-regulated Transcription in MCF-7 Mammary Cells

Kraus

Ariazi

Farrell

et al. 2002

Journal of Biological Chemistry

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116

118

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The estrogen-related receptor ␣ (ERR␣) is an orphan member of the nuclear receptor superfamily. We show that the major isoform of the human ERR␣ gene, ERR␣1, can sequence-specifically bind a consensus palindromic estrogen response element (ERE) and directly compete with estrogen receptor ␣ (ER␣) for binding. ERR␣1 activates or represses ERE-regulated transcription in a cell type-dependent manner, repressing in ER-positive MCF-7 cells while activating in ER-negative HeLa cells. Thus, ERR␣1 can function both as a modulator of estrogen responsiveness and as an estrogen-independent activator. Repression likely occurs in the absence of exogenous ligand since charcoal treatment of the serum had no effect on silencing activity. Mutational analysis revealed that repression is not simply the result of competition between ER␣ and ERR␣1 for binding to the DNA. Rather, it also requires the presence of sequences within the carboxyl-terminal E/F domain of ERR␣1. Thus, ERR␣1 can function as either an active repressor or a constitutive activator of ERE-dependent transcription. We hypothesize that ERR␣1 can play a critical role in the etiology of some breast cancers, thereby providing a novel therapeutic target in their treatment. The nuclear receptor (NR)1 superfamily is comprised of hundreds of transcription factors that regulate a vast array of genes and physiological responses (1-9). Most nuclear receptors share a similar structural organization (Fig. 1A). The amino-terminal A/B domain can function as a hormone-independent activator of transcription. The highly conserved C domain contains the DNA binding domain (DBD) that confers sequence-specific DNA binding activity. A hinge region, called the D domain, bridges the C domain with the carboxyl-terminal E/F domain that includes the receptor-specific ligand binding domain (LBD) of the protein. The binding of appropriate ligands results in conformation changes leading to alterations in the transcriptional properties of the receptor, including the exposure of a transcriptional activation region within the carboxyl end. Although many nuclear receptor superfamily members bind known ligands (e.g., steroids, retinoids, thyroid hormones), some, termed orphan receptors, share significant sequence similarity in their LBDs with their ligand binding family members but lack as-yet known naturally occurring ligands (7-10).Among the first orphan receptors identified were the estrogen-related receptors ERR␣ and ERR␤ (officially named NR3B1 and NR3B2, respectively) (10). They were cloned by low stringency screening of cDNA libraries with probes corresponding to the DBD of estrogen receptor ␣ (ER␣) (10). Subsequently, ERR␣1 was identified as the major isoform present in HeLa cells (Fig. 1A) (11, 12). The DBD of human ERR␣1 shares 70% amino acid similarity with the DBD of human ER␣; the LBD shares 35% amino acid identity. A third member of the ERR family, ERR␥ (NR3B3), has also been identified (13-15). These three ERRs are closely related by sequence similarity but encoded by different genes.De...

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“…Activation of the BRLF1 (Rp) promoter requires EGR-1 and Sp1 binding sites (Zalani et al, 1992(Zalani et al, , 1995. Both IE promoters are negatively regulated by binding sites for the cellular YY-1 transcription factor (Montalvo et al, 1995;Zalani et al, 1997), and BZLF1 transcription is also inhibited by binding of the zincfinger protein, ZEB (Kraus et al, 2001), as well.…”

Section: Induction Of Lytic Ebv Infectionmentioning

confidence: 99%