Methylation status of the Epstein-Barr virus (EBV) BamHI W latent cycle promoter and promoter activity: analysis with novel EBV-positive Burkitt and lymphoblastoid cell lines.
The Epstein-Barr virus (EBV) latent cycle promoter Wp, present in each tandemly arrayed copy of the BamHI W region in the EBV genome, drives expression of the EB viral nuclear antigens (EBNAs) at the initiation of virus-induced B-cell transformation. Thereafter, an alternative EBNA promoter, Cp, becomes dominant, Wp activity declines dramatically, and bisulfite sequencing of EBV-transformed lymphoblastoid cell lines (LCLs) shows extensive Wp methylation. Despite this, Wp is never completely silenced in LCLs. Here, using a combination of bisulfite sequencing and methylation-specific PCR, we show that in standard LCLs transformed with wild-type EBV isolates, some Wp copies always remain unmethylated, and in LCLs transformed with a recombinant EBV carrying just two BamHI W copies, Wp is completely unmethylated. Furthermore, we have analyzed rare LCLs, recently established using wild-type EBV isolates, and rare Burkitt lymphoma (BL) cell clones, recently established from tumors carrying EBNA2-deleted EBV genomes, which express EBNAs exclusively from Wp-initiated transcripts. Here, in sharp contrast to standard LCL and BL lines, all resident copies of Wp appear to be predominantly hypomethylated. Thus, studies of B cells with atypical patterns of Wp usage emphasize the strong correlation between the presence of unmethylated Wp sequences and promoter activity.
Hutchings
Isabel A
IA
Cancer Research UK Institute for Cancer Studies, The University of Birmingham, Edgbaston, Birmingham B15 2TT, United Kingdom. a.i.bell@bham.ac.uk.
Tierney
Rosemary J
RJ
Kelly
Gemma L
GL
Stylianou
Julianna
J
Rickinson
Alan B
AB
Bell
Andrew I
AI
D001402:B-Lymphocytes / Q000821:virology; D001483:Base Sequence; D002051:Burkitt Lymphoma / Q000821:virology; D002461:Cell Line, Transformed; D045744:Cell Line, Tumor; D002471:Cell Transformation, Neoplastic / Q000235:genetics; D002472:Cell Transformation, Viral / Q000235:genetics; D019175:DNA Methylation; D004279:DNA, Viral / Q000737:chemistry* / Q000235:genetics*; D019309:Epstein-Barr Virus Nuclear Antigens / Q000235:genetics; D005814:Genes, Viral; D004854:Herpesvirus 4, Human / Q000235:genetics* / Q000472:pathogenicity; D006801:Humans; D011401:Promoter Regions, Genetic
D004279:DNA, Viral; D019309:Epstein-Barr Virus Nuclear Antigens