Reading the message: Connections between the epigenetic landscape bromodomain activity

Abstract

Bromodomains are conserved epigenetic reader domains known to recognize acetylated lysine residues on the N‐terminal histone tails protruding from the nucleosome. The 61 human bromodomains are found in 41 different proteins, and have been classified into eight different subfamilies based on sequence conservation. These subfamilies often share unique sequence variations that drive differences in histone ligand preference for specific patterns of post‐translational modifications on core and variant histone proteins. The BET bromodomains, which are an attractive drug target in multiple disease states, have been the focus of many studies to elucidate the molecular mechanism of histone recognition. However, much less is known about non‐BET bromodomain‐containing proteins. In this study, we examined the ligand specificity the family IV ATAD2B bromodomain and compared it to its closely related paralog in ATAD2. We show that the ATAD2B bromodomain recognizes mono‐ and di‐acetyllysine modifications on histones H4 and H2A. A structure‐function approach using X‐ray crystallography, isothermal titration calorimetry experiments, and site‐directed mutagenesis coupled to ligand binding assays identified key residues in the acetyllysine binding pocket that dictate the molecular recognition process. Furthermore, our results demonstrate how cross‐talk between multiple modifications alters the binding activity of the ATAD2B bromodomain. The structure of the ATAD2B bromodomain in complex with a small molecule ATAD2 bromodomain inhibitor revealed that critical contacts required for coordination are conserved between the ATAD2/B bromodomains, and many of these residues play a dual role in acetyllysine recognition. We further characterized an alternative splice variant of ATAD2B that results in a loss of bromodomain function. Our results outline the structural and functional features of the ATAD2B bromodomain, and identify novel mechanisms regulating the interaction of the ATAD2B protein with chromatin.