Significance: The correct and regulated readout of epigenetic marks on chromatin is essential to modulate gene expression in living cells. The regulation of chromatin accessibility is ensured by such epigenetic tags, which form a platform for the binding of specific enzymatic modules. A clear example of this mechanism is represented by the histone demethylase LSD1-CoREST, which removes methylation marks from lysine 4 of histone protein H3. We developed a crosslinking technology to capture this histone demethylase in contact with the nucleosome and used this methodology to explore the structural and biophysical properties of this complex. This is one of the very few successful attempts to visualize the molecular mechanism underlying the recognition of the nucleosomal substrate by a histone-modifying enzyme complex.

The binding mode of newly discovered histone demethylase inhibitors could have applications in the design and repurposing of drugs.

LSD1 and LSD2 are homologous histone demethylases with opposite biological outcomes related to chromatin silencing and transcription elongation, respectively. Unlike LSD1, LSD2 nucleosome-demethylase activity relies on a specific linker peptide from the multidomain protein NPAC. We used single-particle cryoelectron microscopy (cryo-EM), in combination with kinetic and mutational analysis, to analyze the mechanisms underlying the function of the human LSD2/NPAC-linker/nucleosome complex. Weak interactions between LSD2 and DNA enable multiple binding modes for the association of the demethylase to the nucleosome. The demethylase thereby captures mono- and dimethyl Lys4 of the H3 tail to afford histone demethylation. Our studies also establish that the dehydrogenase domain of NPAC serves as a catalytically inert oligomerization module. While LSD1/CoREST forms a nucleosome docking platform at silenced gene promoters, LSD2/NPAC is a multifunctional enzyme complex with flexible linkers, tailored for rapid chromatin modification, in conjunction with the advance of the RNA polymerase on actively transcribed genes.