The broad, long-term goal of this project is to understand the functional significance of interactions between promoter specific activators and the general transcription factors in eukaryotes. The Epstein-Barr virus lytic activator, Zta, is a model activator protein that clearly interacts with the general transcription factors TFIIA and TFIID to form a stable preinitiation complex, referred to as Z-D-A. TFIIA is one of several RNA polymerase II accessory proteins essential for activator regulated transcription. However, the fundamental role of TFIIA in activator regulated transcription remains poorly understood. The formation of the Z- D-A complex is dependent on the Zta activation domain, the TAFs in the TFIID fraction, and the number of Zta binding sites in the promoter. These requirements strongly support the functional significance of the Z-D-A complex. However, not all transcriptional activators stimulate a TFIIA- TFIID complex. To investigate the significance of this activator function, we have constructed specific amino acid substitution mutations in the Zta activation domain that fail to stimulate D-A formation, yet are only conditionally defective for transcriptional activation. Preliminary evidence suggests that Z-D-A complex formation is required only under conditions where TFIIA and TFIID binding is rate limiting. The biological significance of this specialized activator function remains unclear. Biologically, Zta functions to disrupt the transcriptional silence of the latent Epstein-Barr virus genome. We hypothesize that Zta has evolved a unique capacity to recruit TFIIA to overcome conditions which maintain transcriptional silence of the latent viral genome. In this proposal, we plan to refine our knowledge of the intermolecular contacts that underpin the Z-D-A complex, and to further investigate the role of TFIIA and the TAFs in mediating activator function. Furthermore, we attempt to identify core promoter elements and cell growth conditions which make TFIIA and TFIID binding rate limiting. These investigations promise to provide general information concerning the role of TFIIA in eukaryotic transcriptional activation mechanisms, to provide molecular details of a unique virus-host interaction, and to correlate the biochemical activity of an activator with a defined biological function.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM054687-05
Application #
6181125
Study Section
Molecular Biology Study Section (MBY)
Program Officer
Tompkins, Laurie
Project Start
1996-08-01
Project End
2002-03-31
Budget Start
2000-08-01
Budget End
2002-03-31
Support Year
5
Fiscal Year
2000
Total Cost
$224,018
Indirect Cost
Name
Wistar Institute
Department
Type
DUNS #
042250712
City
Philadelphia
State
PA
Country
United States
Zip Code
19104
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Wang, Pu; Day, Latasha; Dheekollu, Jayaraju et al. (2005) A redox-sensitive cysteine in Zta is required for Epstein-Barr virus lytic cycle DNA replication. J Virol 79:13298-309
Chen, C J; Deng, Z; Kim, A Y et al. (2001) Stimulation of CREB binding protein nucleosomal histone acetyltransferase activity by a class of transcriptional activators. Mol Cell Biol 21:476-87
Deng, Z; Chen, C J; Zerby, D et al. (2001) Identification of acidic and aromatic residues in the Zta activation domain essential for Epstein-Barr virus reactivation. J Virol 75:10334-47
Solow, S; Salunek, M; Ryan, R et al. (2001) Taf(II) 250 phosphorylates human transcription factor IIA on serine residues important for TBP binding and transcription activity. J Biol Chem 276:15886-92
Bell, P; Lieberman, P M; Maul, G G (2000) Lytic but not latent replication of epstein-barr virus is associated with PML and induces sequential release of nuclear domain 10 proteins. J Virol 74:11800-10
Ozer, J; Moore, P A; Lieberman, P M (2000) A testis-specific transcription factor IIA (TFIIAtau) stimulates TATA-binding protein-DNA binding and transcription activation. J Biol Chem 275:122-8
Zerby, D; Chen, C J; Poon, E et al. (1999) The amino-terminal C/H1 domain of CREB binding protein mediates zta transcriptional activation of latent Epstein-Barr virus. Mol Cell Biol 19:1617-26
Moore, P A; Ozer, J; Salunek, M et al. (1999) A human TATA binding protein-related protein with altered DNA binding specificity inhibits transcription from multiple promoters and activators. Mol Cell Biol 19:7610-20
Solow, S P; Lezina, L; Lieberman, P M (1999) Phosphorylation of TFIIA stimulates TATA binding protein-TATA interaction and contributes to maximal transcription and viability in yeast. Mol Cell Biol 19:2846-52

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