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A Potential Role for Phase Separation in NUT Carcinoma

NUT carcinoma (NC) is a highly aggressive and poorly differentiated type of cancer, mostly occurring in human midline organs. The disease is characterized by chromosomal translocations that involve NUT on chromosome 15 and a partner gene. In about 75% of cases that partner gene has been reported to be BRD4 on chromosome 19. The mechanism by which the BRD4-NUT fusion product causes cancer remains poorly understood. It has been reported that BRD4 protein by itself can form liquid-like intranuclear condensates, which have been shown to be associated with super-enhancers, to control gene expression. Previous studies have also shown that the BRD4-NUT fusion protein forms large, round condensates in cells. I hypothesize that these condensates represent aberrant phase separation of BRD4, which alters transcription, leading to cell transformation.

In my studies, I observed that upon expression in cells, BRD4-NUT fusion protein forms large condensates, that are highly dynamic, suggesting that they might exhibit liquid-like properties. I also determined that there is only a small part of NUT that is required to be in fusion with BRD4 for the formation of these nuclear condensates and that same part is responsible for the interaction with a histone acetyltransferase, p300. Therefore, I subsequently tested the expression of an engineered BRD4-p300 fusion in cells and learned that this fusion is also sufficient to form condensates, which exhibit similar properties to the ones formed by BRD4-NUT. I then expressed different mutants of BRD4-p300 fusion in cells and found that different fragments of p300 collectively contribute to condensates formation.

I am currently in the process of testing how the expression of the different fusion constructs influences gene expression. By combining the RNAseq and ChIP-seq experiments with microscopy, I aim to find out if there is a link between the aberrant phase separation of BRD4-NUT and cell transformation from healthy to diseased cells in Nut Carcinoma.

Publication:

Pak C.W., Kosno M., Holehouse A.S., Padrick S.B., Mittal A., Ali R., Yunus A.A., Liu D.R., Pappu R.V., Rosen M.K. “Sequence Determinants of Intracellular Phase Separation by Complex Coacervation of a Disordered Protein.” Molecular Cell. 2016. Jul 7;63(1):72-85.

https://www.ncbi.nlm.nih.gov/pubmed/27392146

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