PMID:23135711

Citation	Wille, CK, Nawandar, DM, Panfil, AR, Ko, MM, Hagemeier, SR and Kenney, SC (2013) Viral genome methylation differentially affects the ability of BZLF1 versus BRLF1 to activate Epstein-Barr virus lytic gene expression and viral replication. J. Virol. 87:935-50
Abstract	The Epstein-Barr virus (EBV) immediate-early proteins BZLF1 and BRLF1 can both induce lytic EBV reactivation when overexpressed in latently infected cells. Although EBV genome methylation is required for BZLF1-mediated activation of lytic gene expression, the effect of viral genome methylation on BRLF1-mediated viral reactivation has not been well studied. Here, we have compared the effect of viral DNA methylation on BZLF1- versus BRLF1-mediated activation of lytic EBV gene transcription and viral genome replication. We show that most early lytic viral promoters are preferentially activated by BZLF1 in the methylated form, while methylation decreases the ability of BRLF1 to activate most early lytic promoters, as well as the BLRF2 late viral promoter. Moreover, methylation of bacmid constructs containing the EBV genome enhances BZLF1-mediated, but decreases BRLF1-mediated, early lytic gene expression. Methylation of viral promoter DNA does not affect BRLF1 binding to a variety of different CpG-containing BRLF1 binding motifs (RREs) in vitro or in vivo. However, BRLF1 preferentially induces H3K9 histone acetylation of unmethylated promoters in vivo. The methylated and unmethylated forms of an oriLyt-containing plasmid replicate with similar efficiency when transfected into EBV-positive cells that express the essential viral replication proteins in trans. Most importantly, we demonstrate that lytic viral gene expression and replication can be induced by BRLF1, but not BZLF1, expression in an EBV-positive telomerase-immortalized epithelial cell line (NOKs-Akata) in which lytic viral gene promoters remain largely unmethylated. These results suggest that the unmethylated form of the EBV genome can undergo viral reactivation and replication in a BRLF1-dependent manner.
Links	PubMed PMC3554042 Online version:10.1128/JVI.01790-12
Keywords	DNA Methylation; DNA, Viral/metabolism; Gene Expression Regulation, Viral; Genome, Viral; Herpesvirus 4, Human/genetics; Herpesvirus 4, Human/physiology; Humans; Immediate-Early Proteins/metabolism; Trans-Activators/metabolism; Virus Activation; Virus Replication
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Annotations

Gene product	Qualifier	GO Term	Evidence Code	with/from	Aspect	Extension	Notes	Status
EBVB9:BZLF1	involved_in	GO:0045893: positive regulation of transcription, DNA-templated	ECO:0000314: direct assay evidence used in manual assertion		P		Seeded From UniProt	complete
EBVB9:BZLF1		GO:0045893: positive regulation of transcription, DNA-templated	ECO:0000314:		P		Fig 1: DNA methylation enhances BZLF1 transactivation of most early lytic EBV promoters. This proves BZLF1 has a major affect on EBV entering the lytic cycle.	complete CACAO 9645
EBVB9:RTA		GO:0010628: positive regulation of gene expression	ECO:0000314:		P		Figure 5A & 5C. BRLF1 can bind to both methylated and unmethylated promoters to activate lytic gene expression.	complete CACAO 7440

References

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