Potent, Selective, and Cell-active Chemical Probe of Protein Lysine Methyltransferases G9a and GLP: Cell-Based Assays
Résumé
Histone lysine methylation is important epigenetic mark that regulates chromatin structure and gene expression. G9a and GLP lysine methyltransferases dimethylate lysine 9 on histone H3 and repress gene transcription playing roles in development, cancer, pluripotency and other biological processes. We report the development of UNC0638 as a chemical probe of G9a/GLP, active in a cellular system at nM concentrations, with excellent toxicity/potency ratio and low nM in vitro potency and selectivity for G9a/GLP over a wide range of targets. Cell immunofluorescence assay/ Incell Western as fast, reproducible and high throughput cellular assay measuring the levels of dimethylated histone H3 lysine 9 was developed and validated using G9a and GLP knockdowns. High G9a/GLP selectivity for lysine 9 dimethyl modification on histone H3 was also established by the mass spectrometry. In mouse ES cells, UNC0638 reactivated silenced GFP reporter