Mitzi Kuroda headshot
Mitzi I. Kuroda, Ph.D.
Professor of Genetics

Analysis of chromatin organization and epigenetic gene regulation in health and disease

We study epigenetic regulators using genetics, genomics and proteomics. The factors we study include the MSL dosage compensation complex in fruit flies, the Polycomb Group in both flies and humans, and a translocation oncoprotein, BRD4-NUT, that drives an aggressive form of squamous cell cancer in humans. The common thread is that each is strongly implicated in the creation of active or silent chromatin domains that are integral to the fidelity of gene regulation. One serious obstacle to understanding the interactions of such factors with additional proteins and RNAs on chromatin has been the trade-off between removal from the DNA, to allow purification, and the resultant loss of interactions with key partners in function. Therefore, we have adapted a crosslinking approach that allows us to affinity-purify fragmented chromatin with protein and RNAs attached, to avoid disruption of interactions that may only occur on DNA. After reversal of crosslinks, the DNA, protein, histone peptides, and RNA fractions can be separately analyzed using comprehensive sequencing and mass spectrometry. Our current results are providing us with a rich and comprehensive view of key epigenetic complexes bound to their chromatin templates.

An example is our recent work with BRD4-NUT, a translocation-encoded fusion protein that plays a defining role in NUT midline carcinoma (NMC). In collaboration with Christopher French’s lab at BWH, we discovered that nuclear foci containing BRD4-NUT protein correspond to extremely broad, cell type-specific, hyperacetylated chromatin domains in patient tissue and cell lines. These are much larger than typical activated regions or ‘super-enhancers’, ranging from 100 kb to 2 Mb. These ‘megadomains’ appear to reflect a pathologic, feed-forward regulatory loop in which hyperacetylation drives further bromodomain-dependent binding and aberrant transcriptional activity. The novelty of megadomains is that they spread from select pre-existing enhancers, surprisingly not enriched for recently described ‘super-enhancers’, to fill individual topologically associating domains (TADs). Although the selected TADs generally differ by cell type, the c-MYC and TP63 regions are targeted in all NMC patient cells examined to date. The ability to spread to fill whole regulatory compartments surrounding genes encoding proteins like MYC and p63 is likely to explain the extremely aggressive nature of NUT midline carcinoma.

Combination Targeting of the Bromodomain and Acetyltransferase Active Site of p300/CBP.
Authors: Authors: Zucconi BE, Makofske JL, Meyers DJ, Hwang Y, Wu M, Kuroda MI, Cole PA.
Biochemistry
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Sex-specific phenotypes of histone H4 point mutants establish dosage compensation as the critical function of H4K16 acetylation in Drosophila.
Authors: Authors: Copur Ö, Gorchakov A, Finkl K, Kuroda MI, Müller J.
Proc Natl Acad Sci U S A
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"Z4" Complex Member Fusions in NUT Carcinoma: Implications for a Novel Oncogenic Mechanism.
Authors: Authors: Shiota H, Elya JE, Alekseyenko AA, Chou PM, Gorman SA, Barbash O, Becht K, Danga K, Kuroda MI, Nardi V, French CA.
Mol Cancer Res
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Bivalent complexes of PRC1 with orthologs of BRD4 and MOZ/MORF target developmental genes in Drosophila.
Authors: Authors: Kang H, Jung YL, McElroy KA, Zee BM, Wallace HA, Woolnough JL, Park PJ, Kuroda MI.
Genes Dev
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Ectopic protein interactions within BRD4-chromatin complexes drive oncogenic megadomain formation in NUT midline carcinoma.
Authors: Authors: Alekseyenko AA, Walsh EM, Zee BM, Pakozdi T, Hsi P, Lemieux ME, Dal Cin P, Ince TA, Kharchenko PV, Kuroda MI, French CA.
Proc Natl Acad Sci U S A
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upSET, the Drosophila homologue of SET3, Is Required for Viability and the Proper Balance of Active and Repressive Chromatin Marks.
Authors: Authors: McElroy KA, Jung YL, Zee BM, Wang CI, Park PJ, Kuroda MI.
G3 (Bethesda)
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Dosage Compensation in Drosophila-a Model for the Coordinate Regulation of Transcription.
Authors: Authors: Kuroda MI, Hilfiker A, Lucchesi JC.
Genetics
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Streamlined discovery of cross-linked chromatin complexes and associated histone modifications by mass spectrometry.
Authors: Authors: Zee BM, Alekseyenko AA, McElroy KA, Kuroda MI.
Proc Natl Acad Sci U S A
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The Oncoprotein BRD4-NUT Generates Aberrant Histone Modification Patterns.
Authors: Authors: Zee BM, Dibona AB, Alekseyenko AA, French CA, Kuroda MI.
PLoS One
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The oncogenic BRD4-NUT chromatin regulator drives aberrant transcription within large topological domains.
Authors: Authors: Alekseyenko AA, Walsh EM, Wang X, Grayson AR, Hsi PT, Kharchenko PV, Kuroda MI, French CA.
Genes Dev
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