Uncoupling Histone Modification Crosstalk by Engineering Lysine Demethylase LSD1

Biochemical crosstalk between two or more histone modifications is often observed in epigenetic enzyme regulation but its functional significance in cells has been difficult to discern. Prior enzymatic studies have revealed that Lys14 acetylation of histone H3 can inhibit Lys4 demethylation by lysine specific demethylase 1 (LSD1). Here we have engineered a mutant form of LSD1, Y391K, which renders the nucleosome demethylase activity of LSD1 insensitive to Lys14 acetylation. Y391K LSD1 knockin cells show increased repression of a set of genes associated with cellular adhesion. Chromatin profiling revealed that the cis-regulatory regions of these silenced genes display a higher level of H3 Lys14 acetylation than the baseline in unedited, parental cells. Y391K LSD1 knockin cells show diminished H3 mono-methyl Lys4 in the vicinity of these silenced genes, consistent with a role for enhanced LSD1 demethylase activity in these regions. These findings illuminate the functional consequences of disconnecting histone modification crosstalk for a key epigenetic enzyme in gene and chromatin regulation. Cleavage Under Targets & Release Using Nuclease (CUT&RUN) experiments for GFP (LSD1-GFP), H3K4me1, H3K4me2, H3K14ac, and IgG control in K562 cells with WT LSD1-GFP and Y391K LSD1-GFP knockin