Characterization of Lysine Methylation During Neuronal Differentiation of LUHMES cells.

Over one-third of human lysine methyltransferases (KMTs) and lysine demethylases (KDMs)-the enzymes responsible for adding or removing methylation on lysine residues within proteins-are linked to neurodevelopmental disorders (NDDs). Consequently, several studies have explored the roles of specific KMTs or KDMs in neuronal differentiation, and alterations in histone methylation patterns have been identified. It is now widely recognized that KMTs and KDMs also target non-histone proteins, yet knowledge of how non-histone lysine methylation changes during neuronal differentiation remains limited. Here, we address this gap using quantitative mass spectrometry-based proteomics to identify and measure changes in non-histone lysine methylation at three different stages in the Lund human mesencephalic (LUHMES) neuronal differentiation model. We identify 74 lysine methylation sites with significant differences in abundance across differentiation. Our analysis reveals lysine methylation on many non-histone proteins involved in neuronal differentiation and neurodevelopment, including signaling molecules, cytoskeletal proteins, RNA splicing factors, and transcription factors. Overall, this work broadens the understanding of non-histone lysine methylation in a neuronal differentiation model and offers a valuable resource of lysine methylation sites on proteins of biological and clinical significance for future research.