DNA-guided transcription factor interactions extend human gene regulatory code

In the same way that the mRNA-binding specificities of transfer RNAs define the genetic code, the DNA-binding specificities of transcription factors (TFs) form the molecular basis of the gene regulatory code^1,2. The human gene regulatory code is much more complex than the genetic code, in particular because there are more than 1,600 TFs that commonly interact with each other. TF-TF interactions are required for specifying cell fate and executing cell-type-specific transcriptional programs. Despite this, the landscape of interactions between DNA-bound TFs is poorly defined. Here we map the biochemical interactions between DNA-bound TFs using CAP-SELEX, a method that can simultaneously identify individual TF binding preferences, TF-TF interactions and the DNA sequences that are bound by the interacting complexes. A screen of more than 58,000 TF-TF pairs identified 2,198 interacting TF pairs, 1,329 of which preferentially bound to their motifs arranged in a distinct spacing and/or orientation. We also discovered 1,131 TF-TF composite motifs that were markedly different from the motifs of the individual TFs. In total, we estimate that the screen identified between 18% and 47% of all human TF-TF motifs. The novel composite motifs we found were enriched in cell-type-specific elements, active in vivo and more likely to be formed between developmentally co-expressed TFs. Furthermore, TFs that define embryonic axes commonly interacted with different TFs and bound to distinct motifs, explaining how TFs with a similar specificity can define distinct cell types along developmental axes.