The long-term goal of this project is to uncover the role of the core promoter in the regulation of transcription by RNA polymerase II. The core promoter has been traditionally defined to be the sequences that direct the accurate initiation of transcription. However, this simple definition belies a complex and diverse component of the transcription process. Although the core promoter has historically been considered to be a somewhat generic and homogeneous moiety, it is now apparent that many different sequence motifs, such as the BREu, TATA, Inr, and DPE, can each confer specific transcriptional properties to the core promoter. There are no universal core promoter motifs. Moreover, in humans, the best known core promoter element, the TATA box, is present in only about 15% of promoters. It is thus likely that many more core promoter motifs remain to be discovered and characterized. The objective of the proposed research is to undertake a three-tiered approach to the study of the core promoter at the levels of DNA sequences, protein factors, and biological networks. Specific Aim 1 focuses upon the discovery of new core promoter elements. At the present time, there is an incomplete understanding of the DNA sequences that direct the initiation of transcription by RNA polymerase II. There are new motifs as well as transcriptional strategies that remain to be uncovered. Specific Aim 2 involves the identification and characterization of transcription factors that function in a core-promoter-specific manner. For example, Caudal is an enhancer-binding protein and master developmental regulator that functions as a DPE- specific activator. There are likely to be many other factors that regulate transcription via the core promoter. Specific Aim 3 will explore the function of specific core promoter motifs in biological networks. For instance, the DPE is a central component of the Hox gene system. It is likely that other gene networks employ specific core promoter motifs in their regulation. In the future, the core promoter will be an integral part of all analyses of transcriptional regulation. A key aim of this grant is to provide new and important insights into the core promoter and thus contribute critical knowledge for understanding how genes are regulated.