Role of a consensus AP-2 regulatory sequence within the Epstein-Barr virus LMP1 promoter in EBNA2 mediated transactivation.
The Epstein-Barr virus (EBV) tumor-associated latent membrane protein 1 (LMP1) gene expression is transactivated by EBV nuclear antigen 2 (EBNA2) in human B cells. We previously reported that an E-box element at the LMP1 regulatory sequence (LRS) represses transcription of the LMP1 gene through the recruitment of a Max-Mad1-mSin3A complex. In the present study, using deletion/mutation analysis, and electrophoretic mobility shift assays, we show that the promoter region adjacent to the E-box (-59/-67) is required for the full repression conferred by E-box binding proteins. The repressive effect of these factors was overcome by an inhibitor of histone deacetylation, Trichostatin A (TSA), concurring with the reports that histone deacetylation plays an important role in repression mediated by Max-Mad1-mSin3A complex. Furthermore, ChIP analyses showed that histones at the transcriptionally active LMP1 promoter were hyperacetylated, whereas in the absence of transcription they were hypoacetylated. EBNA2 activation of the promoter required a consensus AP-2 sequence in the -103/-95 LRS region. While EMSA results and the low level of AP-2 factors expression in B cells argue against known AP-2 factors binding to this site, several pieces of evidence point to a similar mechanism of promoter activation as seen by AP-2 factors. We conclude that an AP-2 site-binding factor and EBNA2 act in concert to overcome the repression of the LMP1 promoter via the consensus AP-2 site. This activation showed strong correlation with histone hyperacetylation at the promoter, indicating this to be a major mechanism for the EBNA2 mediated LMP1 transactivation.
Jansson
Ann
A
Department of Clinical Chemistry and Transfusion Medicine, The Sahlgrenska Academy at Göteborg University, Sahlgrenska University Hospital, 413 45 Gothenburg, Sweden.
Johansson
Pegah
P
Yang
Weiwen
W
Palmqvist
Lars
L
Sjöblom-Hallén
Anna
A
Rymo
Lars
L
D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D002460:Cell Line; D016384:Consensus Sequence; D019309:Epstein-Barr Virus Nuclear Antigens / Q000502:physiology*; D015967:Gene Expression Regulation, Viral / Q000502:physiology*; D004854:Herpesvirus 4, Human / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D008969:Molecular Sequence Data; D012045:Regulatory Sequences, Nucleic Acid; D050656:Transcription Factor AP-2 / Q000502:physiology*; D015533:Transcriptional Activation; D014763:Viral Matrix Proteins / Q000235:genetics*; D014764:Viral Proteins / Q000502:physiology*
C402641:EBNA-2 protein, Human herpesvirus 4; C037662:EBV-associated membrane antigen, Epstein-Barr virus; D019309:Epstein-Barr Virus Nuclear Antigens; D050656:Transcription Factor AP-2; D014763:Viral Matrix Proteins; D014764:Viral Proteins