Metabolically Derived Lysine Acylations and Neighboring Modifications Tune the Binding of the BET Bromodomains to Histone H4

Recent
proteomic studies discovered histone lysines are modified
by acylations beyond acetylation. These acylations derive from acyl-CoA
metabolites, potentially linking metabolism to transcription. Bromodomains
bind lysine acylation on histones and other nuclear proteins to influence
transcription. However, the extent bromodomains bind non-acetyl acylations
is largely unknown. Also unclear are the effects of neighboring post-translational
modifications, especially within heavily modified histone tails. Using
peptide arrays, binding assays, sucrose gradients, and computational
methods, we quantified 10 distinct acylations for binding to the bromodomain
and extraterminal domain (BET) family. Four of these acylations (hydroxyisobutyrylation,
malonylation, glutarylation, and homocitrullination) had never been
tested for bromodomain binding. We found N-terminal BET bromodomains
bound acetylated and propionylated peptides, consistent with previous
studies. Interestingly, all other acylations inhibited binding of
the BET bromodomains to peptides and nucleosomes. To understand how
context tunes bromodomain binding, effects of neighboring methylation,
phosphorylation, and acylation within histone H4 tails were determined.
Serine 1 phosphorylation inhibited binding of the BRD4 N-terminal
bromodomain to polyacetylated H4 tails by >5-fold, whereas methylation
had no effect. Furthermore, binding of BRDT and BRD4 N-terminal bromodomains
to H4K5acetyl was enhanced 1.4–9.5-fold by any neighboring
acylation of lysine 8, indicating a secondary H4K8acyl binding site
that is more permissive of non-acetyl acylations than previously appreciated.
In contrast, C-terminal BET bromodomains exhibited 9.9–13.5-fold
weaker binding for polyacylated than for monoacylated H4 tails, indicating
the C-terminal bromodomains do not cooperatively bind multiple acylations.
These results suggest acyl-CoA levels tune or block recruitment of
the BET bromodomains to histones, linking metabolism to bromodomain-mediated
transcription.