Retention of epigenetic reader domains in the monocytic leukemic zinc-finger (MOZ) histone acetyltransferase (HAT) leukemic fusion protein is speculated to direct aberrant HAT activity and may be responsible for oncogenic transformations. However, the molecular mechanisms connecting the MOZ HAT complex to its histone substrates are unknown. Progress is hampered because we do not know how chromatin reader domains bridge the MOZ HAT complex to histones. The bromodomain-PHD finger protein 1 (BRPF1) associates with MOZ in leukemic translocations and links the MOZ catalytic subunit with the inhibitor of growth 5 (ING5) and hEaf6 subunits in the MOZ HAT complex promoting activity. BRPF1 contains multiple epigenetic reader domains including a unique double plant homeodomain (PHD) and zinc finger (ZnF) assembly (PZP), a bromodomain (BRD) and a PWWP domain. The overall objective of this proposal is to elucidate the role of the PZP and BRD regions in recruiting BRPF1 to histones. The central hypothesis is that the BRD and PZP regions independently bridge BRPF1 to the histone tail, and the adjacent PHD and zinc finger motifs modulate the histone target of PHD1 diversifying its functionality. This proposal aims to: (1) establish the molecular basis of histone H3 recognition by the unique PHD finger region of BRPF1 and (2) identify the histone ligand(s) of the BRPF1 BRD and structurally characterize the specificity determinants targeting BRPF1 to the histone tail. A unique combination of biochemical, biophysical and structural biology techniques will be used to characterize the structural and functional role of the PZP and BRD regions in BRPF1. The histone tail ligands will be identified and verified using peptide array assays in combination with nuclear magnetic resonance (NMR) chemical shift perturbation techniques. Tryptophan fluorescence, isothermal titration calorimetry (ITC) and/or NMR titration experiments will be used to investigate the effects of post-translational modifications on binding. The atomic resolution structures of the BRPF1 PZP and BRD domains bound to their histone tail ligands will be solved by NMR or X-ray crystallography. The structural data will be used to model the BRPF1 PZP or BRD interaction with histone ligands and to design mutations testing which residues are important for the interaction with the histone tail. The results will establish a uniqe mechanism of chromatin recognition by a novel PHD finger domain and increase our understanding of the molecular mechanism utilized by structurally diverse BRDs to recognize their histone ligands. This information will elucidate the principles targeting the MOZ HAT complex to chromatin substrates and provide insight into how multiple histone reader domains function cooperatively within the BRPF1 subunit. These data will provide critical structural insights into the mechanism used by BRPF1 to target the histone tail, and will impart a greater understanding on how these chromatin binding effectors link epigenetic signals to the regulation of normal and pathological gene expression.

Public Health Relevance

The MOZ histone acetyltransferase (HAT) is involved in chromosomal changes found in a subtype of acute myeloid leukemia (AML) associated with a poor prognosis. In leukemic translocations a subunit of this complex called BRPF1, for bromodomain-PHD finger protein 1 (BRPF1), is associated with MOZ and likely contributes to its aberrant function in leukemia. Our research is designed to investigate the structure and function of the bromodomain and PHD finger region within the BRPF1 subunit to unravel their role in normal and disease processes. This fundamental knowledge will aid in a deeper understanding of how critical epigenetic signaling pathways can be therapeutically manipulated and may help to identify new diagnostic markers and targets to prevent and treat many types of disease, including leukemia.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Academic Research Enhancement Awards (AREA) (R15)
Project #
1R15GM104865-01
Application #
8432160
Study Section
Macromolecular Structure and Function C Study Section (MSFC)
Program Officer
Gerratana, Barbara
Project Start
2013-01-01
Project End
2015-12-31
Budget Start
2013-01-01
Budget End
2015-12-31
Support Year
1
Fiscal Year
2013
Total Cost
$353,400
Indirect Cost
$125,400
Name
Albany College of Pharmacy
Department
Pharmacology
Type
Schools of Pharmacy
DUNS #
797866969
City
Albany
State
NY
Country
United States
Zip Code
12208
Carlson, Samuel; Glass, Karen C (2014) The MOZ histone acetyltransferase in epigenetic signaling and disease. J Cell Physiol 229:1571-4
Lubula, Mulu Y; Poplawaski, Amanda; Glass, Karen C (2014) Crystallization and preliminary X-ray diffraction analysis of the BRPF1 bromodomain in complex with its H2AK5ac and H4K12ac histone-peptide ligands. Acta Crystallogr F Struct Biol Commun 70:1389-93
Lubula, Mulu Y; Eckenroth, Brian E; Carlson, Samuel et al. (2014) Structural insights into recognition of acetylated histone ligands by the BRPF1 bromodomain. FEBS Lett 588:3844-54
Poplawski, Amanda; Hu, Kaifeng; Lee, Woonghee et al. (2014) Molecular insights into the recognition of N-terminal histone modifications by the BRPF1 bromodomain. J Mol Biol 426:1661-76
Lee, Woonghee; Hu, Kaifeng; Tonelli, Marco et al. (2013) Fast automated protein NMR data collection and assignment by ADAPT-NMR on Bruker spectrometers. J Magn Reson 236:83-8