In order for an individual cell to properly execute its gene expression program, the appropriate combinations of factors must be present at specific regulatory regions of the genome at the correct time. In large part, the diversity of the promoters of individual genes directs these requirements; therefore, eukaryotic promoter structure and function have been an area of intense study. RNA polymerase II-dependent promoters can be broken down into two basic components. The core promoter recruits the general transcriptional apparatus and supports basal transcription, while the proximal promoter recruits transcriptional activators, which are necessary for appropriately activated transcription (26). In vitro transcription reconstitution assays have provided an important framework for the specific mechanistic roles the general transcription factors (GTFs) play at core promoters (32, 36). For example, an important step in the assembly of a preinitiation complex at the promoter is the binding of the GTF TFIID (7). TFIID is comprised of the TATA binding protein (TBP) and 10 to 12 TBP-associated factors (TAFs) (53). For core promoters that contain TATA elements, the TBP component of TFIID is important for TFIID binding and, hence, supports basal transcription (26). Another common functional element, termed the initiator (Inr), can also be found in eukaryotic core promoters (39). For this class of core promoter, the largest TFIID component, TAF1 (formerly referred to as TAFII250), has been implicated in recruiting TFIID by binding the Inr in conjunction with TAF2 (formerly referred to as TAFII150) (6). Interestingly, TAF1 offers additional functions at promoters such as acetyltransferase, E1 ubiquitin activation, E2 ubiquitin conjugation, and two Ser/Thr kinase activities, in addition to two bromodomains, which play a role in binding to acetylated histones (52). The precise targets of these activities are not known, but the importance of TAF1 is highlighted by findings that the transcription of 18 and 30% of cellular genes in hamster and Saccharomyces cerevisiae, respectively, is absolutely dependent on its function (18, 31). Furthermore, TAF1 inactivation in Drosophila melanogaster is lethal (51). A major obstacle that must be overcome for transcription to occur is a repressive chromatin structure. The regulation of chromatin dynamics is achieved in large part through the acetylation and deacetylation of the N-terminal tails on nucleosomal histone proteins (23). Histone acetylation is catalyzed by histone acetyltransferase enzymes and strongly correlates with transcriptional activity (15, 33). Indeed, transcriptional activators, in addition to influencing the thermodynamics and kinetics of preinitiation complex assembly, also recruit coactivator complexes which contain histone acetyltransferase enzymes, such as p300/CBP and PCAF (42). Corepressor complexes, conversely, contain histone deacetylase (HDAC) activities and are recruited to promoters by transcriptional repressors (33). Although histones are the best-defined substrates for these activities, other proteins involved in transcription, such as p53, EKLF, GATA-1, TFIIE, and TFIIF, are also acetylated and deacetylated by these enzymes (42). Uncoupling of the fine balance between acetylation and deacetylation is thought to occur upon treatment of cells with HDAC inhibitors (HDIs) such as trichostatin A (TSA) and butyrate. Indeed, treatment with HDIs leads to the accumulation of hyperacetylated nuclear histones (50). Current excitement surrounding these agents stems from their ability to elicit G1/S arrest, differentiation, and/or apoptosis of transformed cells in culture and animal models (28). It would be expected that treatment with these agents would cause a general induction of many cellular genes. However, the present view is that HDIs reprogram gene expression and only affect a very specific subset of genes (27). For example, the most well characterized response to HDI treatment is the p53-independent transcriptional induction of the WAF1 gene, which encodes the cell cycle inhibitor p21WAF1 (19, 30). Induction of WAF1 has been demonstrated as essential for the growth inhibitory effects of these agents (2). However, HDIs can also directly repress genes such as cyclin D1 (25) and c-Myc (16, 41). The repression of these growth-promoting genes offers further explanation for the anticancer effectiveness of HDIs. At the present time, it is largely unknown how HDIs repress gene expression. Recent studies, however, have suggested that the mechanism of HDI-mediated gene expression modulation is direct and that changes in the promoters' chromatin structure, resulting from disrupted acetylation or deacetylation dynamics, are secondary effects (22, 29). We have recently demonstrated that c-Src mRNA and protein expression are inhibited by treatment of a diverse array of cancer cell lines with HDIs (24). Activation and/or overexpression of the c-Src tyrosine kinase has been a consistent finding in colon and other cancers (3), and it has been shown that this activation is at the level of SRC transcription in a subset of human colon cancer cell lines (9). c-Src is the human homologue of the transforming v-Src oncogene encoded in the Rous sarcoma virus genome. SRC transcription is controlled by two disparate promoters separated by approximately 1 kb (5). Despite their apparent dissimilarity, both promoters are directly inhibited following treatment with TSA and butyrate. These observations, coupled with the lack of reports describing the mechanism of gene repression by HDIs, prompted an investigation into the mechanism of SRC inhibition by these agents. A fundamental regulatory similarity was observed for these two promoters in that they both contain Inr elements in their core regions and are TAF1 dependent. Interestingly, the effects of TSA and butyrate on the SRC1A promoter were blocked in cells harboring a temperature-sensitive TAF1 mutant, suggesting that TAF1 could play a role in the repression of transcription mediated by HDIs. The generation of chimeric promoters demonstrated that proximal and core promoter elements from both SRC promoters could independently confer HDI-mediated repression on the WAF1 promoter. Further analysis of these chimeric promoters showed that they were TAF1 dependent even though WAF1 is normally TAF1 independent. In summary, these findings represent the first, potentially functional link between promoter architecture, TAF1 dependence, and HDI-mediated transcriptional repression.