Sara Weirich

University of Stuttgart · Institute of Biochemistry

The human protein lysine methyltransferase NSD2 catalyzes dimethylation at H3K36. It has very important roles in development and disease but many mechanistic features and its full spectrum of substrate proteins are unclear. Using peptide SPOT array methylation assays, we investigate the substrate sequence specificity of NSD2 and discover strong rea...

SETDB1 is a major H3K9 methyltransferase involved in heterochromatin formation and silencing of repeat elements. It contains a unique Triple Tudor Domain (3TD) which specifically binds the dual modification of H3K14ac in the presence of H3K9me1/2/3. Here, we explored the role of the 3TD H3-tail interaction for the H3K9 methylation activity of SETDB...

The HEMK2 protein methyltransferase has been described as glutamine methyltransferase catalyzing ERF1‐Q185me1 and lysine methyltransferase catalyzing H4K12me1. Methylation of two distinct target residues is unique for this class of enzymes. To understand the specific catalytic adaptations of HEMK2 allowing it to master this chemically challenging t...

The protein lysine methyltransferase SETD6 has been shown to influence different cellular activities and to be critically involved in the regulation of diverse developmental and pathological processes. However, the upstream signals which regulate the mRNA expression of SETD6 are not known. Bioinformatic analysis revealed that the SETD6 promoter has...

Protein lysine methylation plays important biological roles but its experimental characterization is limited by the lack of suitable mimetics of methylated and unmethylated lysine among the natural amino acids. Here, we summarize the consequent challenges and discuss alternative approaches for biochemical and cellular lysine methylation studies.

The protein lysine methyltransferase SETD6 has been shown to influence different cellular activities and are critically involved in the regulation of diverse developmental and pathological processes. However, the upstream signal which regulates the mRNA expression of SETD6 is not known. Bioinformatics analysis revealed that SETD6 promoter has a bin...

The regulation of cellular activities is a key hallmark in the development of complex life forms. Among other factors, it is facilitated by protein lysine methyltransferases (PKMTs), which modify proteins in a highly specific manner and regulate their biological activities. Here, we describe methods to decipher the PKMT-substrate specificity by bio...

Protein lysine methyltransferases (PKMTs) play essential roles in gene expression regulation and cancer development. Somatic mutations in PKMTs are frequently observed in cancer cells. In biochemical experiments, we show here that the NSD1 mutations Y1971C, R2017Q and R2017L observed mostly in solid cancers are catalytically inactive suggesting tha...

Somatic mutations in protein lysine methyltransferases are frequently observed in cancer cells. We show here that the NSD1 mutations Y1971C, R2017Q and R2017L observed mostly in solid cancers are catalytically inactive suggesting that NSD1 acts as tumor suppressor gene in these tumors. In contrast, the frequent T1150A in NSD2 and its T2029A counter...

The DNMT3A DNA methyltransferase and MECP2 methylation reader are highly expressed in neurons. Both proteins interact via their DNMT3A‐ADD and MECP2‐TRD domains, and the MECP2 interaction regulates the activity and subnuclear localization of DNMT3A. Here, we mapped the interface of both domains using peptide SPOT array binding, protein pull‐down, e...

N-degron pathway plays an important role in the protein quality control and maintenance of cellular protein homeostasis. ZER1 and ZYG11B, the substrate receptors of the Cullin 2-RING E3 ubiquitin ligase (CRL2), recognize N-terminal (Nt) glycine degrons and participate in the Nt-myristoylation quality control through the Gly/N-degron pathway. Here w...

In previous work, we have developed a DNA methylation‐based epigenetic memory system that operates in Escherichia coli to detect environmental signals, trigger a phenotypic switch of the cells and store the information in DNA methylation. The system is based on the CcrM DNA methyltransferase and a synthetic zinc finger (ZnF4), which binds DNA in a...

Protein lysine methyltransferases have important regulatory functions in cells, but mechanisms determining their activity and specificity are incompletely understood. Naturally, SETD2 introduces H3K36me3, but previously an artificial super-substrate (ssK36) was identified, which is methylated >100-fold faster. The ssK36-SETD2 complex structure cann...

Oscillations are an important component in biological systems; grasping their mechanisms and regulation, however, is difficult. Here, we use the theory of dynamical systems to support the design of oscillatory systems based on epigenetic control elements. Specifically, we use results that extend the Poincaré-Bendixson theorem for monotone control s...

The H3.3 G34W mutation has been observed in 90% of the patients affected by giant cell tumor of bone (GCTB). It had been shown to reduce the activity of the SETD2 H3K36 protein lysine methyltransferase (PKMT) and lead to genome wide changes in epigenome modifications including a global reduction in DNA methylation. Here, we investigated the effect...

Posttranslational methylation of amino acid side chains in proteins mainly occurs on lysine, arginine, glutamine, and histidine residues. It is introduced by different protein methyltransferases (PMTs) and regulates many aspects of protein function including stability, activity, localization, and protein/protein interactions. Although the biologica...

Glioblastoma (GBM) is the most malignant and aggressive form of glioma and is associated with a poor survival rate. Latest generation Tumour Necrosis Factor Related Apoptosis-Inducing Ligand (TRAIL)-based therapeutics potently induce apoptosis in cancer cells, including GBM cells, by binding to death receptors. However, the blood–brain barrier (BBB...

Recent evidence has documented the potential roles of histone-modifying enzymes in autism-spectrum disorder (ASD). Aberrant histone H3 lysine 9 (H3K9) dimethylation resulting from genetic variants in histone methyltransferases is known for neurodevelopmental and behavioral anomalies. However, a systematic examination of H3K9 methylation dynamics in...

SUV39H1 and SUV39H2 were the first protein lysine methyltransferases that were identified more than 20 years ago. Both enzymes introduce di- and trimethylation at histone H3 lysine 9 (H3K9) and have important roles in the maintenance of heterochromatin and gene repression. They consist of a catalytically active SET domain and a chromodomain, which...

In recent years, epigenetic memory systems have been developed based on DNA methylation and positive feedback systems. Achieving a robust design for these systems is generally a challenging and multifactorial task. We developed and validated a novel mathematical model to describe methylation‐based epigenetic memory systems that capture switching dy...

The lysine specific demethylase 1 (LSD1) plays a pivotal role in cellular differentiation by regulating the expression of key developmental genes in concert with different coregulatory proteins. This process is impaired in different cancer types and incompletely understood. To comprehensively identify functional coregulators of LSD1, we established...

Post-translational methylation plays a crucial role in regulating and optimizing protein function. Protein histidine methylation, occurring as the two isomers 1- and 3-methylhistidine (1MH and 3MH), was first reported five decades ago, but remains largely unexplored. Here we report that METTL9 is a broad-specificity methyltransferase that mediates...

Clr4 is a histone H3 lysine 9 methyltransferase in Schizosaccharomyces pombe that is essential for heterochromatin formation. Previous biochemical and structural studies have shown that Clr4 is in an autoinhibited state in which an autoregulatory loop (ARL) blocks the active site. Automethylation of lysine residues in the ARL relieves autoinhibitio...

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

SETD2 catalyzes methylation at lysine 36 of histone H3 and it has many disease connections. We investigated the substrate sequence specificity of SETD2 and identified nine additional peptide and one protein (FBN1) substrates. Our data showed that SETD2 strongly prefers amino acids different from those in the H3K36 sequence at several positions of i...

Eukaryotic N-degron pathways are proteolytic systems whose unifying feature is their ability to recognize proteins containing N-terminal (Nt) degradation signals called N-degrons, and to target these proteins for degradation by the 26S proteasome or autophagy. GID4, a subunit of the GID ubiquitin ligase, is the main recognition component of the pro...

Bacterial live cell sensors are potentially powerful tools for the detection of environmental toxins. In this work, we have established and validated a flow cytometry readout for an existing bacterial arabinose sensor system with DNA methylation based memory function (Maier et al., 2017, Nat. Comm., 8:15336). Flow cytometry readout is convenient an...

The SMYD2 protein lysine methyltransferase methylates various histone and non‐histone proteins and is overexpressed in several cancers. Using peptide arrays, we investigated the substrate specificity of the enzyme, revealing a recognition of leucine (or weaker phenylalanine) at the −1 peptide site and disfavor of acidic residues at the +1 to +3 sit...

RomA is a SET-domain containing protein lysine methyltransferase encoded by the Gram-negative bacterium Legionella pneumophila. It is exported into human host cells during infection and has been previously shown to methylate histone H3 at lysine 14 (Rolando et al., 2013, Cell Host Microbe 13:395-405). Here, we investigated the substrate specificity...

SETDB1 is an essential H3K9 methyltransferase involved in silencing of retroviruses and gene regulation. We show here that its triple Tudor domain (3TD) specifically binds to doubly modified histone H3 containing K14 acetylation and K9 methylation. Crystal structures of 3TD in complex with H3K14ac/K9me peptides reveal that peptide binding and K14ac...

Table S1. Compilation of P‐values for the data shown in this manuscript.

Somatic missense mutations in the MLL1 histone H3K4 methyltransferase are often observed in cancers. MLL1 forms a complex with WDR5, RBBP5 and ASH2L (WRA) which stimulates its activity. The MM-102 compound prevents the interaction between MLL1 and WDR5 and functions as an MLL1 inhibitor. We have studied the effects of four cancer mutations in the c...

Somatic mutations in epigenetic enzymes are frequently found in cancer tissues. The MLL3 H3K4-specific protein lysine monomethyltransferase is an important epigenetic enzyme, and it is among the most recurrently mutated enzymes in cancers. MLL3 mainly introduces H3K4me1 at enhancers. We investigated the enzymatic properties of MLL3 variants that ca...

It has been reported that the Numb protein is methylated at lysine 158 and 163 and that this methylation is introduced by the SET8 protein lysine methyltransferase [Dhami et al., (2013) Molecular Cell 50, 565-576]. We studied this methylation in vitro using peptide arrays and recombinant Numb protein as substrates. Numb peptides and protein were in...

Lysine methylation is an emerging post-translation modification and it has been identified on several histone and non-histone proteins, where it plays crucial roles in cell development and many diseases. Approximately 5,000 lysine methylation sites were identified on different proteins, which are set by few dozens of protein lysine methyltransferas...

β-Barrel membrane proteins have regular structures with extensive hydrogen-bond networks between their transmembrane (TM) β-strands, which stabilize their protein fold. Nevertheless, weakly stable TM regions, which are important for the protein function and interaction with other proteins, exist. Here, we report on the apparent stability of human T...