Multivalent gene regulatory elements localize condensation of transcription machinery

Enhancers are regulatory elements that cooperatively assemble the transcriptional machinery upon binding by sequence-specific transcription factors. Recently, we and others have hypothesized and demonstrated that the transcriptional apparatus forms liquid-like condensates, particularly at cell-type specific enhancer elements called super-enhancers. Many transcription factors (TFs) and coactivators phase separate in vitro, but only at supra-physiological concentrations. A unified mechanism to describe how and why these condensates form around specific genomi loci at physiological conditions is unknown. Here, using a combination of simulation, and complementary in vitro experiments, we propose that specific and multivalent enhancers nucleate localized condensation of TFs and coactivators at physiologic-like conditions. Using thermodynamic guiding principles, we predict key interactions that regulate condensate formation, as well as motif features encoded in DNA that drive phase separation and higher-order organization of the 3-D genome. Experiments validate our predictions, and many of these features are encoded and leveraged by mammalian genomes to assemble high amounts of transcription machinery, implicating phase separation in shaping the gene regulatory landscape.