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The BRPF1 bromodomain is a molecular reader of di-acetyllysine.

ABSTRACT: Bromodomain-containing proteins are often part of chromatin-modifying complexes, and their activity can lead to altered expression of genes that drive cancer, inflammation and neurological disorders in humans. Bromodomain-PHD finger protein 1 (BRPF1) is part of the MOZ (monocytic leukemic zinc-finger protein) HAT (histone acetyltransferase) complex, which is associated with chromosomal translocations known to contribute to the development of acute myeloid leukemia (AML). BRPF1 contains a unique combination of chromatin reader domains including two plant homeodomain (PHD) fingers separated by a zinc knuckle (PZP domain), a bromodomain, and a proline-tryptophan-tryptophan-proline (PWWP) domain. BRPF1 is known to recruit the MOZ HAT complex to chromatin by recognizing acetylated lysine residues on the N-terminal histone tail region through its bromodomain. However, histone proteins can contain several acetylation modifications on their N-terminus, and it is unknown how additional marks influence bromodomain recruitment to chromatin. Here, we identify the BRPF1 bromodomain as a selective reader of di-acetyllysine modifications on histone H4. We used ITC assays to characterize the binding of di-acetylated histone ligands to the BRPF1 bromodomain and found that the domain binds preferentially to histone peptides H4K5acK8ac and H4K5acK12ac. Analytical ultracentrifugation (AUC) experiments revealed that the monomeric state of the BRPF1 bromodomain coordinates di-acetylated histone ligands. NMR chemical shift perturbation studies, along with binding and mutational analyses, revealed non-canonical regions of the bromodomain-binding pocket that are important for histone tail recognition. Together, our findings provide critical information on how the combinatorial action of post-translational modifications can modulate BRPF1 bromodomain binding and specificity.

SUBMITTER: Obi JO

PROVIDER: S-EPMC7861561 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Bromodomain-PHD finger protein 1 (BRPF1) belongs to the BRPF family of bromodomain-containing proteins. Bromodomains are exclusive reader modules that recognize and bind acetylated histones and non-histone transcription factors to regulate gene expression. The biological functions of acetylated histone recognition by BRPF1 bromodomain are well characterized; however, the function of BRPF1 regulation via non-histone acetylation is still unexplored. Therefore, identifying the non-histone interactome of BRPF1 is pivotal in deciphering its role in diverse cellular processes, including its misregulation in diseases like cancer. Herein, we identified the non-histone interacting partners of BRPF1 utilizing a protein engineering-based approach. We site-specifically introduced the unnatural photo-cross-linkable amino acid 4-azido-L-phenylalanine into the bromodomain of BRPF1 without altering its ability to recognize acetylated histone proteins. Upon photoirradiation, the engineered BRPF1 generates a reactive nitrene species, cross-linking interacting partners with spatio-temporal precision. We demonstrated the robust cross-linking efficiency of the engineered variant with reported histone ligands of BRPF1 and further used the variant reader to cross-link its interactome. We also characterized novel interacting partners by proteomics, suggesting roles for BRPF1 in diverse cellular processes. BRPF1 interaction with interleukin enhancer-binding factor 3, one of these novel interacting partners, was further validated by isothermal titration calorimetry and co-IP. Lastly, we used publicly available ChIP-seq and RNA-seq datasets to understand the colocalization of BRPF1 and interleukin enhancer-binding factor 3 in regulating gene expression in the context of hepatocellular carcinoma. Together, these results will be crucial for full understanding of the roles of BRPF1 in transcriptional regulation and in the design of small-molecule inhibitors for cancer treatment.

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The BRPF1 bromodomain is a molecular reader of di-acetyllysine.

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