Biology subjects

Hanquier, J. N.

Publications and source records attributed to Hanquier, J. N..

3 recordsLinked to original sources

bioRxiv2026

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.

neuroscience↗

bioRxiv2024

Quantitative analysis of non-histone lysine methylation sites and lysine demethylases in breast cancer cell lines

Growing evidence shows that lysine methylation is a widespread protein post-translational modification that regulates protein function on histone and non-histone proteins. Numerous studies have demonstrated that dysregulation of lysine methylation mediators contributes to cancer growth and chemotherapeutic resistance. While changes in histone methylation are well documented with extensive analytical techniques available, there is a lack of high-throughput methods to reproducibly quantify changes in the abundances of the mediators of lysine methylation and non-histone lysine methylation (Kme) simultaneously across multiple samples. Recent studies by our group and others have demonstrated that antibody enrichment is not required to detect lysine methylation, prompting us to investigate the use of Tandem Mass Tag (TMT) labeling for global Kme quantification sans antibody enrichment in four different breast cancer cell lines (MCF-7, MDA-MB-231, HCC1806, and MCF10A). To improve the quantification of KDMs, we incorporated a lysine demethylase (KDM) isobaric trigger channel, which enabled 96% of all KDMs to be quantified while simultaneously quantifying 326 Kme sites. Overall, 142 differentially abundant Kme sites and eight differentially abundant KDMs were identified between the four cell lines, revealing cell line-specific patterning.

cancer biology↗

bioRxiv2022

Substrate selectivity of the PRDM9 lysine methyltransferase domain.

Lysine methylation is a dynamic, post-translational mark that regulates the function of histone and non-histone proteins. Many of the enzymes that mediate lysine methylation, known as lysine methyltransferases (KMTs), were originally identified to modify histone proteins but have also been discovered to methylate non-histone proteins. In this work, we investigate the substrate selectivity of the lysine methyltransferase PRDM9 to identify both potential histone and non-histone substrates. Though normally expressed in germ cells, PRDM9 is significantly upregulated across many cancer types. The methyltransferase activity of PRDM9 is essential for double-strand break formation during meiotic recombination. PRDM9 has been reported to methylate histone H3 at lysine residues 4 and 36; however, PRDM9 KMT activity had not previously been evaluated on non-histone proteins. Using lysine-oriented peptide (K-OPL) libraries to screen potential substrates of PRDM9, we determined that PRDM9 preferentially methylates peptide sequences not found in any histone protein. We confirmed PRDM9 selectivity through in vitro KMT reactions using peptides with substitutions at critical positions. A multisite {lambda}-dynamics computational analysis provided a structural rationale for the observed PRDM9 selectivity. The substrate selectivity profile was then used to identify putative non-histone substrates, which were tested by peptide spot array. Finally, PRDM9 methylation non-histone substrates were validated at the protein level by in vitro KMT assays on recombinant proteins. The selectivity profile of PRDM9 will be useful in identifying putative PRDM9 substrates in different cellular contexts, and future studies are required to determine whether PRDM9 methylates non-histone proteins in the context of meiotic recombination or cancer.

biochemistry↗