Deciphering Transcriptional Regulation of Human Core Promoters

Although traditionally viewed as universal stretches of DNA, core promoters are diverse regulatory sequenceswith a high impact on transcriptional activity in promoters and enhancers. However, our understanding of their function, architecture, and cis-regulatory elementsis still lacking. Here, we devised a high-throughput assay to quantify the core promoter activity of ~15,000 fully designed sequences that we integrated and expressed from a fixed location within the human genome. By measuring ~500 Pre-Initiation Complex (PIC) binding sequences, we find functional differences between promoters and enhancers and that both exhibit unidirectional activity. Our systematic investigation of all possible combinations of core promoter elements reveals a positive effect on expression of TATA and Initiator, and a negative effect of BREu and BREd. Moreover, we observe a 10bp periodicity in the optimal distance between the TATA and the Inr. By comprehensively screening TF binding-sites, we show that site orientation has little effect, that the effect of binding site number on expression is factor-specific, and that there is a striking agreement between the effect of binding site multiplicity and TF appearance in endogenous homotypic clusters. Thus, we provide a systematic view of the core- and proximal- promoter regions and shed light on organization principlesof regulatory regions in the human genome.