c-Myc (Myc) is a DNA-binding transcription factor that heterodimerizes with an obligatory partner, Max, to regulate transcription of genes important for cell proliferation, differentiation, and apoptosis. Myc is highly regulated and is required for normal embryonic development. Deregulated Myc expression is associated with disorders characterized by excess cell proliferation, including cancer and cardiovascular disease. All known functions of Myc require its N-terminal transcription regulatory domain (TAD). However, the cofactors and molecular mechanisms that mediate Myc TAD functions have remained elusive. Thus, elucidating the mechanisms by which Myc regulates transcription is of general medical relevance. Recent observations, including our own, suggest possible roles of histone acetyltransferases (HATs) in regulating Myc functions. We have purified a multisubunit transcription coactivator complex, STAGA, which contains the human HAT GCN5, TRRAP, and other transcription cofactors. STAGA associates with Myc in vivo and in vitro via direct physical interactions with the TAD domain. We postulate that Myc recruits HAT complexes, including STAGA, to its regulatory DNA sequences on target genes and that this allows transcription activation through mechanisms that we propose to investigate.
In Specific Aim 1 we will analyze the role of chromatin acetylation, STAGA, and specific STAGA/GCN5-interacting coactivators in transcription regulation by Myc. We will use both cell-free (in vitro) and tissue culture (in vivo) transcription assays, including modified chromatin immunoprecipitation (CHIP) protocols to analyze the function of the Myc-coactivator/HAT associations.
In Specific Aim 2 we will address the possible regulation of Myc:Max functions by p300 and the role(s) of Myc:Max acetylation.
In Specific Aim 3 we will characterize the Myc-STAGA/p300 protein interface. We will map the region(s) of Myc TAD that directly interact with STAGA and p300 by using GST pull-down and co-immunoprecipitation assays. We will identify the STAGA subunit(s) that contacts Myc TAD in crosslinking experiments. This proposal constitutes the first phase of our long-term goal to understand the role of coactivators and acetylation in regulation of transcription and cell proliferation by Myc.

National Institute of Health (NIH)
National Cancer Institute (NCI)
Research Project (R01)
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Biochemistry Study Section (BIO)
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Mietz, Judy
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University of California Riverside
Schools of Earth Sciences/Natur
United States
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Zhang, Na; Ichikawa, Wataru; Faiola, Francesco et al. (2014) MYC interacts with the human STAGA coactivator complex via multivalent contacts with the GCN5 and TRRAP subunits. Biochim Biophys Acta 1839:395-405
Wang, Yuan-Liang; Faiola, Francesco; Martinez, Ernest (2012) Purification of multiprotein histone acetyltransferase complexes. Methods Mol Biol 809:427-43
Wang, Yuan-Liang; Faiola, Francesco; Xu, Muyu et al. (2008) Human ATAC Is a GCN5/PCAF-containing acetylase complex with a novel NC2-like histone fold module that interacts with the TATA-binding protein. J Biol Chem 283:33808-15
Liu, Xiaohui; Vorontchikhina, Marina; Wang, Yuan-Liang et al. (2008) STAGA recruits Mediator to the MYC oncoprotein to stimulate transcription and cell proliferation. Mol Cell Biol 28:108-21
Faiola, Francesco; Wu, Yi-Ting; Pan, Songqin et al. (2007) Max is acetylated by p300 at several nuclear localization residues. Biochem J 403:397-407
Zhang, Kangling; Faiola, Francesco; Martinez, Ernest (2005) Six lysine residues on c-Myc are direct substrates for acetylation by p300. Biochem Biophys Res Commun 336:274-80
Faiola, Francesco; Liu, Xiaohui; Lo, Szuying et al. (2005) Dual regulation of c-Myc by p300 via acetylation-dependent control of Myc protein turnover and coactivation of Myc-induced transcription. Mol Cell Biol 25:10220-34