EBV nuclear antigen EBNALP dismisses transcription repressors NCoR and RBPJ from enhancers and EBNA2 increases NCoR-deficient RBPJ DNA binding

Abstract

EBV nuclear antigen 2 (EBNA2) and EBV nuclear antigen LP (EBNALP) are critical for B-lymphocyte transformation to lymphoblastoid cell lines (LCLs). EBNA2 activates transcription through recombination signal-binding immunoglobulin κJ region (RBPJ), a transcription factor associated with NCoR repressive complexes, and EBNALP is implicated in repressor relocalization. EBNALP coactivation with EBNA2 was found to dominate over NCoR repression. EBNALP associated with NCoR and dismissed NCoR, NCoR and RBPJ, or NCoR, RBPJ, and EBNA2 from matrix-associated deacetylase (MAD) bodies. In non-EBV-infected BJAB B lymphoma cells that stably express EBNA2, EBNALP, or EBNA2 and EBNALP, EBNALP was associated with hairy and enhancer of split 1 (hes1), cd21, cd23, and arginine and glutamate-rich 1 (arglu1) enhancer or promoter DNA and was associated minimally with coding DNA. With the exception of RBPJ at the arglu1 enhancer, NCoR and RBPJ were significantly decreased at enhancer and promoter sites in EBNALP or EBNA2 and EBNALP BJAB cells. EBNA2 DNA association was unaffected by EBNALP, and EBNALP was unaffected by EBNA2. EBNA2 markedly increased RBPJ at enhancer sites without increasing NCoR. EBNALP further increased hes1 and arglu1 RNA levels with EBNA2 but did not further increase cd21 or cd23 RNA levels. EBNALP in which the 45 C-terminal residues critical for transformation and transcriptional activation were deleted associated with NCoR but was deficient in dismissing NCoR from MAD bodies and from enhancer and promoter sites. These data strongly support a model in which EBNA2 association with NCoR-deficient RBPJ enhances transcription and EBNALP dismisses NCoR and RBPJ repressive complexes from enhancers to coactivate hes1 and arglu1 but not cd21 or cd23.lymphoma | notch E BV causes posttransplantation and AIDS-related lymphomas and is a cofactor in African Burkitt lymphoma (BL), and Hodgkin disease. In vitro, EBV latency III infection converts B lymphocytes to immortal lymphoblast cell lines (LCLs) (1).EBV nuclear antigen 2 (EBNA2) and EBNALP are the first proteins expressed in latency III infection (2, 3). EBNA2 is essential for B-cell transformation (4, 5) and for up-regulation of EBV latency III and cell RNA expression through CSL/CBF1/ recombination signal-binding protein for immunoglobulin κJ region (RBPJ), a DNA sequence-specific cell transcription factor (6-8). EBNA2 up-regulated cell genes include hairy and enhancer of split 1 (hes1), myc, cd21, and cd23 (2, 9, 10). The EBNA2 acidic activation domain recruits basal and activated transcription factors (TF) including TFIIB, TFIIE, TFIIH, histone acetyl transferases p300, CBP, and PCAF, and RNA polymerase II (11-16).RBPJ is in repressive complexes containing NCoR1 and -2, SKIP, and SHARP (17-23). NCoR recruits class I and II histone deacetylases (HDAC), including HDAC3, which repress transcription and associate with matrix-associated deacetylase (MAD) bodies, subnuclear sites of repressed transcription (24, 25). NCoR overexpression represses EBNA2 up-regulation of promot...

Epstein-Barr virus exploits intrinsic B-lymphocyte transcription programs to achieve immortal cell growth

“…S2). Overall, EBNA2/RBPJ had more RBPJ signals than RBPJ-only sites genome-wide, consistent with the previous finding that EBNA2 significantly increased RBPJ association with a limited number of enhancer or promoter DNA sites (25). Motifs.…”

The Human Papillomavirus Type 8 E6 Protein Interferes with NOTCH Activation during Keratinocyte Differentiation

“…This suggested that HPV8 E6 can bind to NOTCH-responsive RBPJ target sites. MAML binding to RBPJ requires a composite ICN/RBPJ binding groove, and thus MAML cannot bind either of the factors independent of the other (18). It has been reported that HPV8 E6 binds to MAML directly and subsequently indirectly to RBPJ and ICN (9).…”

“…The HES1 promoter has two well-characterized RBPJ binding sites adjacent to the promoter that are required for NOTCH-mediated transcription (17). An additional RBPJ binding site important for HES1 expression in response to NOTCH activation approximately 5,000 bp upstream of the promoter has also been described (18). These sites in the HES1 enhancer (forward, 5= CCTCCCAGGATAGCTCTTG 3=; reverse, 5= TTTGCCT GAGGACTTGAAGC 3=) and promoter (forward, 5= CAAGACC AAAGCGGAAAGAA 3=; reverse, 5= GGTACCTGTGTGATCCCT AGGC 3=) were assessed using specific primer pairs (Fig.…”

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“…EBNA2 is tethered to cell DNA mostly through RBPJ (11). EBNA2 is similar to its cell counterpart, Notch, which induces RBPJ DNA binding (22). RBPJ ChIP-seq signals also distinguish ESEs from ETEs (26).…”

“…Its C-terminal transactivation domain recruits basal transcription machinery and activates gene transcription (11,(18)(19)(20). EBNALP coactivates with EBNA2 by removing repressors (21,22). EBNA3C binds to the p14 ARF promoter and represses CDKN2A expression (23,24), whereas EBNA3A binds to sites further upstream of CDKN2B (25).…”