MECHANISMS OF TRANSCIPTIONAL REGULATION BY ATF

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Long term objective: To better understand the regulation of eukaryotic gene expression. From atherosclerosis to cancer, many diseases are the result of under-expression or over-expression of a gene or a group of genes. Thus, a better understanding of gene expression will lead to a better understanding of, and possibly even cures to, different diseases. Specific aim: To study the molecular mechanisms by which a group of activating transcription factor (ATF) dimers regulate transcription. One recent discovery in transcription research is that many different transcription factors can bind to a given DNA regulatory sequence in vitro. Since a given regulatory sequence may occur in different promoters, this discovery raises several questions. For example, do these different transcription factors regulate different promoters? If so, how is the specificity achieved? Do they differ from one another in ways other than regulating different promoters? This proposal addresses these questions by describing experiments to study a superfamily of transcription factors: the ATF/CREB and Fos/Jun "leucine zipper" proteins. These transcription factors form selective dimers with each other via the leucine zipper regions, and bind to similar DNA sequences. In addition, they are induced by many extracellular stimuli, such as viral infection, growth factors and peptide hormones that increase cellular cAMP level. The goal of this proposal is to use ATF as a model to elucidate fundamental principles of transcriptional regulation. The strategy is to focus on a group of stable ATF homodimers and heterodimers: ATF-1, ATF-3, ATF-4, ATF-3/c-Jun and ATF-4/Fra-1. Specific methods: (1) A "random mutagenesis" analysis to study the importance of DNA sequences on ATF dimer binding, (2) In vitro transcription and in vivo transfection experiments to find out whether different ATF dimers regulate different genes; (3) Pulse-chase labeling, chemical cross-linking, immunoprecipitation and two-dimensional analysis to study how different ATF proteins are induced to become transcriptionally active by extracellular stimuli.