Hierarchy of grammar rules for the language of transcriptional activation domains

Transcriptional activation domains (ADs) of eukaryotic gene activators have remained enigmatic for decades as short, consensus-less, extremely variable amino acid sequences that lack a specific structure and interact fuzzily with an uncertain number of targets. Understanding AD sequence grammar is critical for solving the enigma. Using rational design of AD sequences and high-throughput in vivo experimentation combined with bioinformatic analysis and machine learning, we refined grammar rules for AD sequences, calculated the relative importance of each rule to define a rule hierarchy, and linked each rule to the biochemical features essential for biological function. The key dominating and common features-redundant representation in the sequence of aromatic residues and the obligatory net negative charge of the sequence are consistent with the novel idea that ADs function as acidic-hydrophobic surfactants, which is crucial for understanding eukaryotic gene regulation and the function of intrinsically disordered protein regions.