Ectopic protein interactions within BRD4–chromatin complexes drive oncogenic megadomain formation in NUT midline carcinoma

To investigate the mechanism that drives dramatic mistargeting of active chromatin in NUT midline carcinoma (NMC), we have identified protein interactions unique to the BRD4–NUT fusion oncoprotein compared with wild-type BRD4. Using cross-linking, affinity purification, and mass spectrometry, we ide...

Full description

Saved in:
Bibliographic Details
Published in:Proceedings of the National Academy of Sciences - PNAS 2017-05, Vol.114 (21), p.E4184-E4192
Main Authors: Alekseyenko, Artyom A., Walsh, Erica M., Zee, Barry M., Pakozdi, Tibor, Hsi, Peter, Lemieux, Madeleine E., Dal Cin, Paola, Ince, Tan A., Kharchenko, Peter V., Kuroda, Mitzi I., French, Christopher A.
Format: Article
Language:English
Subjects:
Acetyltransferase
Affinity
Biochemistry
Biological Sciences
Carcinoma, Squamous Cell - genetics
Carcinoma, Squamous Cell - pathology
Cell Cycle Proteins
Cell Line, Tumor
Cell Proliferation - genetics
Cells
Chromatin
Chromatin - metabolism
Crosslinking
E1A-Associated p300 Protein - metabolism
Epithelial Cells - pathology
Female
Functional anatomy
HEK293 Cells
Humans
In Situ Hybridization, Fluorescence
Lung Neoplasms - genetics
Lung Neoplasms - pathology
Mass spectrometry
Mass spectroscopy
Medical diagnosis
Middle Aged
Multiprotein Complexes - genetics
Nuclear Proteins - genetics
Nuclear Proteins - metabolism
Oncogene Proteins - genetics
Oncogene Proteins - metabolism
Oncogene Proteins, Fusion - genetics
Players
PNAS Plus
Propagation
Protein Domains - genetics
Protein interaction
Proteins
Purification
Repressor Proteins - genetics
Repressor Proteins - metabolism
RNA Interference
RNA, Small Interfering - genetics
Transcription Factors - genetics
Transcription Factors - metabolism
Translocation
Zinc Fingers - genetics
Citations: Items that this one cites
Items that cite this one
Online Access:Get full text
Tags: Add Tag
No Tags, Be the first to tag this record!
Description
Summary:To investigate the mechanism that drives dramatic mistargeting of active chromatin in NUT midline carcinoma (NMC), we have identified protein interactions unique to the BRD4–NUT fusion oncoprotein compared with wild-type BRD4. Using cross-linking, affinity purification, and mass spectrometry, we identified the EP300 acetyltransferase as uniquely associated with BRD4 through the NUT fusion in both NMC and non-NMC cell types. We also discovered ZNF532 associated with BRD4–NUT in NMC patient cells but not detectable in 293T cells. EP300 and ZNF532 are both implicated in feed-forward regulatory loops leading to propagation of the oncogenic chromatin complex in BRD4–NUT patient cells. Adding key functional significance to our biochemical findings, we independently discovered a ZNF532–NUT translocation fusion in a newly diagnosed NMC patient. ChIP sequencing of the major players NUT, ZNF532, BRD4, EP300, and H3K27ac revealed the formation of ZNF532–NUT–associated hyperacetylated megadomains, distinctly localized but otherwise analogous to those found in BRD4–NUT patient cells. Our results support a model in which NMC is dependent on ectopic NUT-mediated interactions between EP300 and components of BRD4 regulatory complexes, leading to a cascade of misregulation.
ISSN:0027-8424
1091-6490
DOI:10.1073/pnas.1702086114