The proposed research in this training plan will focus on investigating the mechanisms by which the toxicity of oncohistones in cells can be abated by manipulating histone post-translational modifications. In eukaryotic cells, DNA is stored as chromatin, which consists of repeating units called nucleosomes made up of DNA wrapped around histone proteins. Extending out from each histone are peptide tails that are post-translationally modified in a variety of ways, including by acetylation, methylation, glycosylation and ubiquitylation. These modifications (PTMs) modulate access to the DNA, which in turn controls transcription, replication, and other cellular processes. Importantly, misregulation of these epigenetic processes has been linked to cancer, and recently it has also been shown by our group and others that mutations in the core histone proteins are found in cancer cells. In particular, mutations in histone 3 (H3) are associated with a deadly form of pediatric brain tumor. In a majority of diffuse pontine glioblastomas (DIPGs), lysine 27 in H3 is mutated to methionine (H3K27M) in a small percentage of nucleosomes. This mutation acts as a potent inhibitor of the polycomb repressor complex 2 (PRC2), which is a methyltransferase. In general, methylation of H3K27 by PRC2 is associated with gene silencing, so reduced levels of H3K27me3 due to H3K27M lead to disruption of genomic programs and thus perturbation of cellular development. Since PRC2 is able to sense chromatin states and modulate its methyltransferase output based on those signals, we hypothesized that PRC2 inhibition could be impacted by existing PTMs on the same H3 tail (i.e. in cis). Indeed, we found that PTMs, particularly polyacetylation of H3, have been found to diminish the inhibitory effect of H3K27M peptides. Therefore, we postulate that the deliberate manipulation of PTM levels relevant to PRC2 could be used to abate the pathogenicity of H3K27M. The strategy explored in this project is to use histone deacetylase (HDAC) inhibitors to achieve elevated levels of acetylation of H3 in H3K27M mutant cells and to study the mechanisms by which this manipulation impacts the PTM cross-talks. The work described herein employs a chemical biology approach to probe specific mechanisms associated with H3K27M detoxification as well as to provide a better understanding of epigenetic misregulation by oncohistones in general.
The specific aims of this project are: (1) To screen HDAC inhibitors for their effect on methyltransferase activity in cells expressing the mutant H3K27M. (2) To characterize mechanisms by which HDACs impact methyltransferase activity in cells expressing H3K27M. (3) To determine the effect of HDAC inhibitors on the pathogenicity of glioblastoma cells that express the H3K27M mutant. This research applies chemical biology tools to elucidate mechanisms by which this cancerous mutation is affected by manipulating the acetylation/methylation cross-talk in cells via HDAC inhibitors. The long term goal of performing these fundamental studies is to illuminate these interconnected PTM pathways and inform the future development of therapeutic procedures.
Histone mutations have been implicated in deadly pediatric brain cancers, including diffused pontine glioblastomas (DIPGs) that often carry a lysine-to-methionine mutation on histone 3 (H3K27M) that leads to decreased methylation of H3K27 and, consequently, gene misregulation. The goal of this project is to investigate how this mutation can be 'detoxified' by manipulating post-translational modifications on H3 to restore methylation levels. Efforts will focus on characterizing the mechanisms by which increasing H3 acetylation with histone deacetylase (HDAC) inhibitors rescues H3K27 methylation in mutant cells.
|Dann, Geoffrey P; Liszczak, Glen P; Bagert, John D et al. (2017) ISWI chromatin remodellers sense nucleosome modifications to determine substrate preference. Nature 548:607-611|