A5006847738- Epigenetics and DNA Methylation
- Cancer-related gene regulation
- RNA modifications and cancer
- Genomics and Chromatin Dynamics
- Protein Degradation and Inhibitors
- CRISPR and Genetic Engineering
- Histone Deacetylase Inhibitors Research
- Chromatin Remodeling and Cancer
- Genetics and Neurodevelopmental Disorders
- Pluripotent Stem Cells Research
- Ovarian cancer diagnosis and treatment
- Acute Myeloid Leukemia Research
- RNA Research and Splicing
- Multiple Myeloma Research and Treatments
- RNA Interference and Gene Delivery
- Chemical Reactions and Isotopes
- Peptidase Inhibition and Analysis
- Estrogen and related hormone effects
- Amino Acid Enzymes and Metabolism
- Microtubule and mitosis dynamics
- Aluminum toxicity and tolerance in plants and animals
- Genetic Syndromes and Imprinting
- Genetics, Aging, and Longevity in Model Organisms
- Virus-based gene therapy research
- Reproductive System and Pregnancy
University of Stuttgart
2017-2025
Research Institute of Molecular Pathology
2015-2021
Vienna Biocenter
2015-2019
Constructor University
2007-2014
Max Planck Society
2010
Max Planck Innovation
2010
International University
2006
The Dnmt3a DNA methyltransferase contains in its N-terminal part a PWWP domain that is involved chromatin targeting. Here, we have investigated the interaction of with modified histone tails using peptide arrays and show it specifically recognizes 3 lysine 36 trimethylation mark. H3K36me3 known to be repressive modification correlated methylation mammals heterochromatin <i>Schizosaccharomyces pombe</i>. These results were confirmed by equilibrium binding studies pulldown experiments native...
Using peptide arrays and binding to native histone proteins, we show that the ADD domain of Dnmt3a specifically interacts with H3 1–19 tail. Binding is disrupted by di- trimethylation K4, phosphorylation T3, S10 or T11 acetylation K4. We did not observe H4 The Dnmt3b shows same specificity, suggesting distinct biological functions both enzymes are related their domains. To establish a functional role unmodified tails, analyzed DNA methylation in vitro reconstituted chromatin Dnmt3a2,...
Chromatin structure is greatly influenced by histone tail post-translational modifications (PTM), which also play a central role in epigenetic processes. Antibodies against modified tails are research reagents chromatin biology and molecular epigenetics. We applied Celluspots peptide arrays for the specificity analysis of 36 commercial antibodies from different suppliers directed towards tails. The contained 384 peptides 8 regions N-terminal histones, viz. H3 1-19, 7-26, 16-35 26-45, H4 1-19...
Investigation of the fundamental role epigenetic processes requires methods for locus-specific detection modifications in living cells. Here, we address this urgent demand by developing four modular fluorescence complementation-based biosensors live-cell microscopy applications. These tools combine engineered DNA-binding proteins with domains recognizing defined marks, both fused to non-fluorescent fragments a fluorescent protein. The presence mark at target DNA sequence leads reconstitution...
Somatic DNMT3A mutations at R882 are frequently observed in AML patients including the very abundant R882H, but also R882C, R882P and R882S. Using deep enzymology, we show here that DNMT3A-R882H has more than 70-fold altered flanking sequence preferences when compared with wildtype DNMT3A. The R882H mainly differ on 3' side of CpG site, where they resemble DNMT3B, while 5' DNMT3A, indicating behaves like a DNMT3A/DNMT3B chimera. Investigation activity other revealed cause similar effects....
The nuclear receptor binding SET [su(var) 3-9, enhancer of zeste, trithorax] domain-containing protein 1 (NSD1) lysine methyltransferase (PKMT) was known to methylate histone H3 36 (H3K36). We show here that NSD1 prefers aromatic, hydrophobic, and basic residues at the -2, -1 +2, +1 sites its substrate peptide, respectively. methylation 25 nonhistone peptide substrates by NSD1, two which were (weakly) methylated level, suggesting unstructured regions are preferred substrates. Methylation...
At physiological levels, the trace element selenium plays a key role in redox reactions through incorporation of selenocysteine antioxidant enzymes. Selenium has also been evaluated as potential anti-cancer agent, where nanoparticles have proven effective, and are well tolerated vivo at doses that toxic soluble Se. The use such nanoparticles, coated with either serum albumin or naturally occurring alkaline polysaccharide chitosan, serves to enhance biocompatibility bioavailability. Here we...
Abstract Chromatin properties are regulated by complex networks of epigenome modifications. Currently, it is unclear how these modifications interact and if they control downstream effects such as gene expression. We employed promiscuous chromatin binding a zinc finger fused catalytic domain DNMT3A to introduce DNA methylation in HEK293 cells at many CpG islands (CGIs) systematically investigated the dynamics introduced consequent changes network. observed efficient thousands CGIs, but was...
Despite their central importance in mammalian development, the mechanisms that regulate DNA methylation machinery and thereby generation of genomic patterns are still poorly understood. Here, we identify 5mC-binding protein MeCP2 as a direct strong interactor methyltransferase 3 (DNMT3) proteins. We mapped interaction interface to transcriptional repression domain ADD DNMT3A find binding strongly inhibits activity vitro. This effect was reinforced by cellular studies where global reduction...
Abstract The lysine specific demethylase 1 (LSD1) plays a pivotal role in cellular differentiation by regulating the expression of key developmental genes concert with different coregulatory proteins. This process is impaired cancer types and incompletely understood. To comprehensively identify functional coregulators LSD1, we established novel tractable fluorescent reporter system to monitor LSD1 activity living cells. Combining this state-of-the-art multiplexed RNAi screen, DEAD-box...
Platinum-based combination chemotherapy remains the backbone of first-line treatment for patients with advanced epithelial ovarian cancer (EOC). While most initially respond well to treatment, relapse ultimately develop platinum resistance. This study identified FLYWCH-type zinc finger-containing protein 1 (FLYWCH1) as an important regulator in resistance development process. We showed that loss FLYWCH1 promotes EOC cells, and low expression is correlated poor prognosis patients. In...
AbstractDNA methyltransferase 1 methylates hemi-methylated CpG sites generated during DNA replication. Serine 515 of this enzyme has been shown to be phosphorylated. To explore the importance S515 phosphorylation, we mutants Dnmt1 which removed phosphorylation potential (S515A) or mimic phosphoserine (S515E), purified proteins from insect cells and analyzed their methylation activity in vitro. The S515E mutant was found active, while S515A had severe loss when compared wild type protein....
The nucleotide analogue azacitidine (AZA) is currently the best treatment option for patients with high-risk myelodysplastic syndromes (MDS). However, only half of treated respond and these almost all eventually relapse. New options are urgently needed to improve clinical management patients. Here, we perform a loss-of-function shRNA screen identify histone acetyl transferase transcriptional co-activator, CREB binding protein (CBP), as major regulator AZA sensitivity. Compounds inhibiting...
We describe a continuous peptide methylation assay using the Neurospora crassa Dim-5 histone H3 lysine 9 (H3K9) methyltransferase as model system. The uses streptavidin FlashPlates coated with target peptide. Since no washing and pipeting steps were required after addition of enzyme/S-adenosyl-l-methionine (AdoMet) mixture to microplate, readout reaction progress was possible. show that this is highly reproducible (with errors in order ±3%). well suited for simultaneous analysis up 384...
The authors describe a continuous protein methylation assay using the G9a lysine methyltransferase and its substrate WIZ (widely interspaced zinc finger motifs). is based on coupling of biotinylated to streptavidin-coated FlashPlates transfer radioactive methyl groups from S-adenosyl-L-methionine substrate. reaction progress monitored continuously by proximity scintillation counting. very accurate, convenient, well suited for automation, highly reproducible with standard errors in range 5%....
Purpose Wnt pathway modulator Dickkopf 2 (Dkk2) and signaling of the G protein-coupled estrogen receptor (GPER) seem to have essential functions in numerous cancer types. For epithelial ovarian (EOC), it has not been proven if either Dkk2 or GPER on its own an independent impact overall survival (OS). So far, correlation both factors their clinical significance systematically investigated before. Methods Expression levels were immunohistochemically analyzed 156 patient samples from different...