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Methylation-dependent binding of the epstein-barr virus BZLF1 protein to viral promoters.

Dickerson SJ, Xing Y, Robinson AR, Seaman WT, Gruffat H, Kenney SC - PLoS Pathog. (2009)

Bottom Line: Z serine residue 186, which was previously shown to be required for Z binding to methylated ZREs in Rp, but not for Z binding to the AP1 site, is required for Z binding to methylated Nap ZREs.The Z(S186A) mutant cannot activate methylated Nap in reporter gene assays and does not induce Na expression in cells with latent EBV infection.Molecular modeling studies of Z bound to the methylated Nap ZREs help to explain why methylation is required for Z binding, and the role of the Z Ser186 residue.

View Article: PubMed Central - PubMed

Affiliation: McArdle Laboratory, Departments of Oncology and Medicine, University of Wisconsin School of Medicine and Public Health, Madison, Wisconsin, USA.

ABSTRACT
The switch between latent and lytic Epstein-Barr virus (EBV) infection is mediated by the viral immediate-early (IE) protein, BZLF1 (Z). Z, a homologue of c-jun that binds to AP1-like motifs (ZREs), induces expression of the BRLF1 (R) and BRRF1 (Na) viral proteins, which cooperatively activate transcription of the Z promoter and thereby establish a positive autoregulatory loop. A unique feature of Z is its ability to preferentially bind to, and activate, the methylated form of the BRLF1 promoter (Rp). To date, however, Rp is the only EBV promoter known to be regulated in this unusual manner. We now demonstrate that the promoter driving transcription of the early BRRF1 gene (Nap) has two CpG-containing ZREs (ACGCTCA and TCGCCCG) that are only bound by Z in the methylated state. Both Nap ZREs are highly methylated in cells with latent EBV infection. Z efficiently activates the methylated, but not unmethylated, form of Nap in reporter gene assays, and both ZREs are required. Z serine residue 186, which was previously shown to be required for Z binding to methylated ZREs in Rp, but not for Z binding to the AP1 site, is required for Z binding to methylated Nap ZREs. The Z(S186A) mutant cannot activate methylated Nap in reporter gene assays and does not induce Na expression in cells with latent EBV infection. Molecular modeling studies of Z bound to the methylated Nap ZREs help to explain why methylation is required for Z binding, and the role of the Z Ser186 residue. Methylation-dependent Z binding to critical viral promoters may enhance lytic reactivation in latently infected cells, where the viral genome is heavily methylated. Conversely, since the incoming viral genome is initially unmethylated, methylation-dependent Z activation may also help the virus to establish latency following infection.

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Comparison of ZRE sites.Sequences of the consensus AP1 site, the three Rp ZRE sites and the two Nap ZREs are shown. Methylated cytosines are indicated with an asterisk.
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ppat-1000356-g009: Comparison of ZRE sites.Sequences of the consensus AP1 site, the three Rp ZRE sites and the two Nap ZREs are shown. Methylated cytosines are indicated with an asterisk.

Mentions: The sequences of the consensus AP1 site (from the BMRF1p), a ZRE which is bound by Z in the unmethylated form (from the Rp), and the four known methylation-sensitive ZREs (from the Rp and Nap) are compared in Fig. 9. Interestingly, the ZREs which are bound by Z in the methylated form all have a CpG motif in the left half-site at the same position (leading to methylated cytosines at −2 and +1').


Methylation-dependent binding of the epstein-barr virus BZLF1 protein to viral promoters.

Dickerson SJ, Xing Y, Robinson AR, Seaman WT, Gruffat H, Kenney SC - PLoS Pathog. (2009)

Comparison of ZRE sites.Sequences of the consensus AP1 site, the three Rp ZRE sites and the two Nap ZREs are shown. Methylated cytosines are indicated with an asterisk.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2654727&req=5

ppat-1000356-g009: Comparison of ZRE sites.Sequences of the consensus AP1 site, the three Rp ZRE sites and the two Nap ZREs are shown. Methylated cytosines are indicated with an asterisk.
Mentions: The sequences of the consensus AP1 site (from the BMRF1p), a ZRE which is bound by Z in the unmethylated form (from the Rp), and the four known methylation-sensitive ZREs (from the Rp and Nap) are compared in Fig. 9. Interestingly, the ZREs which are bound by Z in the methylated form all have a CpG motif in the left half-site at the same position (leading to methylated cytosines at −2 and +1').

Bottom Line: Z serine residue 186, which was previously shown to be required for Z binding to methylated ZREs in Rp, but not for Z binding to the AP1 site, is required for Z binding to methylated Nap ZREs.The Z(S186A) mutant cannot activate methylated Nap in reporter gene assays and does not induce Na expression in cells with latent EBV infection.Molecular modeling studies of Z bound to the methylated Nap ZREs help to explain why methylation is required for Z binding, and the role of the Z Ser186 residue.

View Article: PubMed Central - PubMed

Affiliation: McArdle Laboratory, Departments of Oncology and Medicine, University of Wisconsin School of Medicine and Public Health, Madison, Wisconsin, USA.

ABSTRACT
The switch between latent and lytic Epstein-Barr virus (EBV) infection is mediated by the viral immediate-early (IE) protein, BZLF1 (Z). Z, a homologue of c-jun that binds to AP1-like motifs (ZREs), induces expression of the BRLF1 (R) and BRRF1 (Na) viral proteins, which cooperatively activate transcription of the Z promoter and thereby establish a positive autoregulatory loop. A unique feature of Z is its ability to preferentially bind to, and activate, the methylated form of the BRLF1 promoter (Rp). To date, however, Rp is the only EBV promoter known to be regulated in this unusual manner. We now demonstrate that the promoter driving transcription of the early BRRF1 gene (Nap) has two CpG-containing ZREs (ACGCTCA and TCGCCCG) that are only bound by Z in the methylated state. Both Nap ZREs are highly methylated in cells with latent EBV infection. Z efficiently activates the methylated, but not unmethylated, form of Nap in reporter gene assays, and both ZREs are required. Z serine residue 186, which was previously shown to be required for Z binding to methylated ZREs in Rp, but not for Z binding to the AP1 site, is required for Z binding to methylated Nap ZREs. The Z(S186A) mutant cannot activate methylated Nap in reporter gene assays and does not induce Na expression in cells with latent EBV infection. Molecular modeling studies of Z bound to the methylated Nap ZREs help to explain why methylation is required for Z binding, and the role of the Z Ser186 residue. Methylation-dependent Z binding to critical viral promoters may enhance lytic reactivation in latently infected cells, where the viral genome is heavily methylated. Conversely, since the incoming viral genome is initially unmethylated, methylation-dependent Z activation may also help the virus to establish latency following infection.

Show MeSH
Related in: MedlinePlus