Development of chemical tools to interrogate the roles of lysine and arginine methylation in epigenetic regulation

Epigenetic regulation of gene expression is critical to development, cellular homeostasis, and disease pathology. One important factor of the epigenetic signaling system is chromatin methylation. In particular, histone protein methylation of lysine and arginine residues is widespread and influences myriad cellular processes. To aid in this research, a facile method was developed to install methylarginine analogs on recombinant protein for use in biochemical, biophysical and structural studies. Through chemical conjugation of novel alpha,beta-unsaturated amidine precursors with proteins, methylarginine mimics can be displayed with control of methylation site, extent and regiospecificity. Moreover, a discussion of the enzymes responsible for installing chromatin methylation further highlights the importance of protein and nucleic acid methylation in cellular physiology and disease. Complementary to the activity of methylating enzymes, the removal of methylation marks by demethylase proteins is essential for dynamic gene expression. Of particular interest is the demethylation of methyllysine residues in histone proteins, which has been shown to strongly influence transcription of associated genes. In certain cancer cells overexpressing such lysine demethylase enzymes, epigenetic signaling is dysregulated, resulting in aberrant gene expression conducive to cancer phenotypes. Therefore, chemical probes were developed against a cancer-associated demethylase, JMJD2C, to study the cellular role of said enzyme during normal transcription as well as in cancer pathology. This work has produced several novel and potent inhibitors, which will be used in cellular assays. With the acquisition of structural data from crystallography efforts, structure-based design is feasible for compound optimization in regard to potency and paralog selectivity. Ultimately, these chemical probes are intended to explore the cellular functions of JMJD2C and its role in cancer. Taken together, this body of work represents a broad investigation of chromatin methylation and its influence on epigenetics from the perspective of chemical biology