Functional Analysis of the Promoter of Epstein-Barr Virus BBLF2/3
Date Issued
2011
Date
2011
Author(s)
Lin, Yi-Ting
Abstract
Rta and Zta, two transcription factors expressed during the immediate-early stage of the Epstein-Barr virus (EBV) lytic cycle, are required for lytic activation. Rta activates many EBV early genes, including BMRF1, BHRF1, BHLF1 and BMLF1, through direct binding to Rta response elements (RRE). Rta also enhances SP1-mediated transcription through the interaction with MBD1-containing chromatin-associated factor 1 (MCAF1) and Sp1. In addition, the binding of Rta-MCAF1-Zta complex to Zta response elements (ZRE) activates BHLF1, BMRF1 and BRLF1 synergistically. The EBV BBLF2/3 gene encodes a primase-associated factor that is indispensable for EBV lytic DNA replication. However, the mechanism that activates the transcription of BBLF2/3 remains unclear. The purpose of this study is to elucidate how Rta and Zta activate BBLF2/3 transcription. This study finds that the BBLF2/3 promoter is activated by Rta, but not by Zta. Moreover, Rta and Zta activated the BBLF2/3 promoter synergistically. Sequence analysis from the TESS website predicts that BBLF2/3 promoter contains a putative AP-1-binding sequence, TGACACG, which is located at -74 to -80 in the BBLF2/3 promoter. Reporter assay showed that mutating the site lowers the transcription activity. Meanwhile, Rta and ATF2 activate a promoter in a reporter plasmid, pBBLF2/3-3AP1, which contains three copies of AP-1 site from the BBLF2/3 promoter, in a dose-dependent manner. In addition, ATF2 binds to the AP-1 site as demonstrated by DNA-affinity precipitation assay. Rta also interacts with the AP-1 site as shown by electrophoretic mobility shift assay. Taken together, this study demonstrates that the BBLF2/3 promoter contains an AP-1 binding site from -74 to -80. Rta activates the BBLF2/3 transcription via an indirect binding to the AP-1 site.
Subjects
EBV
AP-1 binding site
Type
thesis
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