title: Mechanistic analysis of the role of bromodomain-containing protein 4 (BRD4) in BRD4-NUT oncoprotein-induced transcriptional activation.
abstract: NUT midline carcinoma (NMC) is a rare but highly aggressive cancer typically caused by the translocation t(15;19), which results in the formation of the BRD4-NUT fusion oncoprotein. Previous studies have demonstrated that fusion of the NUT protein with the double bromodomains of BRD4 may significantly alter the cellular gene expression profile to contribute to NMC tumorigenesis. However, the mechanistic details of this BRD4-NUT function remain poorly understood. In this study, we examined the NUT function in transcriptional regulation by targeting it to a LacO transgene array integrated in U2OS 2-6-3 cells, which allow us to visualize how NUT alters the in situ gene transcription dynamic. Using this system, we demonstrated that the NUT protein tethered to the LacO locus recruits p300/CREB-binding protein (CBP), induces histone hyperacetylation, and enriches BRD4 to the transgene array chromatin foci. We also discovered that, in BRD4-NUT expressed in NMC cells, the NUT moiety of the fusion protein anchored to chromatin by the double bromodomains also stimulates histone hyperacetylation, which causes BRD4 to bind tighter to chromatin. Consequently, multiple BRD4-interacting factors are recruited to the NUT-associated chromatin locus to activate in situ transgene expression. This gene transcription function was repressed by either expression of a dominant negative inhibitor of the p300-NUT interaction or treatment with (+)-JQ1, which dissociates BRD4 from the LacO chromatin locus. Our data support a model in which BRD4-NUT-stimulated histone hyperacetylation recruits additional BRD4 and interacting partners to support transcriptional activation, which underlies the BRD4-NUT oncogenic mechanism in NMC.
authors.affiliation: From the Department of Microbiology, University of Pennsylvania Perelman School of Medicine, Philadelphia, Pennsylvania 19104-6076.
authors.forename: Ranran
authors.initials: R
authors.lastname: Wang
authors.affiliation: From the Department of Microbiology, University of Pennsylvania Perelman School of Medicine, Philadelphia, Pennsylvania 19104-6076 jianyou@mail.med.upenn.edu.
authors.forename: Jianxin
authors.initials: J
authors.lastname: You
mesh_terms: D018797:Cell Cycle Proteins; D045744:Cell Line, Tumor; D047369:Chromatin Immunoprecipitation; D050881:E1A-Associated p300 Protein / Q000235:genetics / Q000378:metabolism; D006801:Humans; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism*; D015514:Oncogene Proteins, Fusion / Q000235:genetics / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D015533:Transcriptional Activation / Q000235:genetics / Q000502:physiology; D000092468:Bromodomain Containing Proteins
chemical_list: D018797:Cell Cycle Proteins; D050881:E1A-Associated p300 Protein; D009687:Nuclear Proteins; D015514:Oncogene Proteins, Fusion; D014157:Transcription Factors; C441289:BRD4 protein, human; D000092468:Bromodomain Containing Proteins; C471990:BRD4-NUT fusion oncogene protein, human; C494360:EP300 protein, human
keywords: Acetyltransferase; BRD4-NUT; Bromodomain-containing Protein 4 (BRD4); E1A Binding Protein p300 (P300); NUT Midline Carcinoma; Transcription Elongation Factor; Transcription Factor; Transcriptional Activation