Modulation of p300/CBP Acetylation of Nucleosomes by Bromodomain Ligand I-CBP112

Abstract

The histone acetyltransferase (HAT) enzymes p300 and CBP are closely related paralogs that serve as transcriptional coactivators and have been found to be dysregulated in cancer and other diseases. p300/CBP is a multidomain protein and possesses a highly conserved bromodomain that has been shown to bind acetylated Lys residues in both proteins and various small molecules, including I-CBP112 and CBP30. Here we show that the ligand I-CBP112 can stimulate nucleosome acetylation up to 3-fold while CBP30 does not. Activation of p300/CBP by I-CBP112 is not observed with the isolated histone H3 substrate but requires a nucleosome substrate. I-CBP112 does not impact nucleosome acetylation by the isolated p300 HAT domain, and the effects of I-CBP112 on p300/CBP can be neutralized by CBP30, suggesting that I-CBP112 likely allosterically activates p300/CBP through bromodomain interactions. Using mass spectrometry and Western blots, we have found that I-CBP112 particularly stimulates acetylation of Lys18 of histone H3 (H3K18) in nucleosomes, an established in vivo site of p300/CBP. In addition, we show that I-CBP112 enhances H3K18 acetylation in acute leukemia and prostate cancer cells in a concentration range commensurate with its antiproliferative effects. Our findings extend the known pharmacology of bromodomain ligands in the regulation of p300/CBP and suggest a novel approach to modulating histone acetylation in cancer.

Role of the CBP catalytic core in intramolecular SUMOylation and control of histone H3 acetylation

“…Binding of Ac-K1596 to the BRD is likely to be enhanced within the intact CBP catalytic core due to its increased effective concentration. In contrast to our observations with ischemin, the bromodomain inhibitor I-CBP112 activates CBP/p300 toward acetylation of a subset of H3 and H4 residues (H3K18, H3K23, H4K5) within nucleosomal substrates, whereas a different inhibitor, CBP30, has no effect on histone acetylation (46). It is…”

Allosteric regulation of epigenetic modifying enzymes

“…This activation of acetylation is seen both in vitro and in cell culture. The requirement of p300/CBP domains beyond the HAT domain and the intact nucleosome substrate points toward a multifaceted interaction shift propagated by I-CBP112, potentially unique to this bromodomain ligand among other small molecules [53]. The importance of the bromodomain to activate p300 acetylation was consistent with an earlier study in which deletion of the bromodomain abolished p300 acetylation of an entire library of nucleosome substrates [54].…”

CRISPR/Cas9 knockouts reveal genetic interaction between strain-transcendent erythrocyte determinants of Plasmodium falciparum invasion

“…Six compounds were identified that displayed substantial induction of JK-1 cell differentiation. Significantly, four of the six top compounds included inhibitors of bromodomain-containing proteins: two compounds, (+)-JQ1 (45) and PFI-1 (46), target mammalian bromodomain and extra terminal domain (BET) proteins; bromosporine is a general bromodomain inhibitor (47); and I-CBP112 targets the bromodomain of cAMP-responsive element-binding protein binding protein (CREBBP)/E1A-associated protein p300 (EP300) (48,49). The only other bromodomain-specific inhibitor in the library, PFI-3, targets a different category of bromodomaincontaining proteins (50) and was not found to be an inducer of JK-1 differentiation.…”