Selected Publications of Prof. Jeltsch

Some of the Publications of the Research by Prof. Dr. Albert Jeltsch are shown on this page.

Recent Highlights

2013 - today

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Lungu, Pinter, Broche, Rathert, Jeltsch (2017) Modular fluorescence complementation sensors for live cell detection of epigenetic signals at endogenous genomic sites. Nature Communications 8:649

Investigation of the fundamental role of epigenetic processes requires methods for the locus-specific detection of epigenetic modifications in living cells. Here, we address this urgent demand by developing four modular fluorescence complementation-based epigenetic biosensors for live-cell microscopy applications. These tools combine engineered DNA-binding proteins with domains recognizing defined epigenetic marks, both fused to non-fluorescent fragments of a fluorescent protein. The presence of the epigenetic mark at the target DNA sequence leads to the reconstitution of a functional fluorophore. With this approach, we could for the first time directly detect DNA methylation and histone 3 lysine 9 trimethylation at endogenous genomic sites in live cells and follow dynamic changes in these marks upon drug treatment, induction of epigenetic enzymes and during the cell cycle. We anticipate that this versatile technology will improve our understanding of how specific epigenetic signatures are set, erased and maintained during embryonic development or disease onset.Tools for imaging epigenetic modifications can shed light on the regulation of epigenetic processes. Here, the authors present a fluorescence complementation approach for detection of DNA and histone methylation at endogenous genomic sites allowing following of dynamic changes of these marks by live-cell microscopy.

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Maier , Möhrle, & Jeltsch (2017) Design of synthetic epigenetic circuits featuring memory effects and reversible switching based on DNA methylation. Nature Communcations 8, 15336
 
Epigenetic systems store information in DNA methylation patterns in a durable but reversible manner, but have not been regularly used in synthetic biology. Here, we designed synthetic epigenetic memory systems using DNA methylation sensitive engineered zinc finger proteins to repress a memory operon comprising the CcrM methyltransferase and a reporter. Triggering by heat, nutrients, ultraviolet irradiation or DNA damaging compounds induces CcrM expression and DNA methylation. In the induced on-state, methylation in the operator of the memory operon prevents zinc finger protein binding leading to positive feedback and permanent activation. Using an mf-Lon protease degradable CcrM variant enables reversible switching. Epigenetic memory systems have numerous potential applications in synthetic biology, including life biosensors, death switches or induction systems for industrial protein production. The large variety of bacterial DNA methyltransferases potentially allows for massive multiplexing of signal storage and logical operations depending on more than one input signal.
 

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Kudithipudi & Jeltsch (2016) Approaches and Guidelines for the Identification of Novel Substrates of Protein Lysine Methyltransferases. Cell Chem. Biol. 23, 1049-55

Protein lysine methylation is emerging as a general post-translational modification (PTM) with essential functions regulating protein stability, activity, and protein-protein interactions. One of the outstanding challenges in this field is linking protein lysine methyltransferases (PKMTs) with specific substrates and lysine methylation events in a systematic manner. Inability to validate reported PKMT substrates delayed progress in the field and cast unnecessary doubt about protein lysine methylation as a truly general PTM. Here, we aim to provide a concise guide to help avoid some of the most common pitfalls in studies searching for new PKMT substrates and propose a set of seven basic biochemical rules: (1) include positive controls; (2) use target lysine mutations of substrate proteins as negative controls; (3) use inactive enzyme variants as negative controls; (4) report quantitative methylation data; (5) consider PKMT specificity; (6) validate methyl lysine antibodies; and (7) connect cellular and in vitro results. We explain the logic behind them and discuss how they should be implemented in the experimental work.

 

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Kungulovski & Jeltsch (2016) Epigenome Editing: State of the Art, Concepts, and Perspectives. Trends in Genetics 32, 101-113

Epigenome editing refers to the directed alteration of chromatin marks at specific genomic loci by using targeted EpiEffectors which comprise designed DNA recognition domains (zinc finger, TAL effector, or modified CRISPR/Cas9 complex) and catalytic domains from a chromatin-modifying enzyme. Epigenome editing is a promising approach for durable gene regulation, with many applications in basic research including the investigation of the regulatory functions and logic of chromatin modifications and cellular reprogramming. From a clinical point of view, targeted regulation of disease-related genes offers novel therapeutic avenues for many diseases. We review here the progress made in this field and discuss open questions in epigenetic regulation and its stability, methods to increase the specificity of epigenome editing, and improved delivery methods for targeted EpiEffectors. Future work will reveal if the approach of epigenome editing fulfills its great promise in basic research and clinical applications.

 

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Kungulovski et al. (2014) Application of histone modification-specific interaction domains as an alternative to antibodies. Genome Research 24(11): 1842-53

Post-translational modifications (PTMs) of histones constitute a major chromatin indexing mechanism, and their proper characterization is of highest biological importance. So far, PTM-specific antibodies have been the standard reagent for studying histone PTMs despite caveats such as lot-to-lot variability of specificity and binding affinity. Herein, we successfully employed naturally occurring and engineered histone modification interacting domains for detection and identification of histone PTMs and ChIP-like enrichment of different types of chromatin. Our results demonstrate that histone interacting domains are robust and highly specific reagents that can replace or complement histone modification antibodies. These domains can be produced recombinantly in Escherichia coli at low cost and constant quality. Protein design of reading domains allows for generation of novel specificities, addition of affinity tags, and preparation of PTM binding pocket variants as matching negative controls, which is not possible with antibodies.

 

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Jeltsch & Jurkowska (2014) New concepts in DNA Methylation. Trends in Biochemical Sciences, 39(7):310-18

The widely-cited model of maintenance of DNA methylation at CpG sites implies that DNA methylation is introduced by the Dnmt3 de novo DNA methyltransferases during early development, and methylation at hemimethylated CpG sites is specifically maintained by the Dnmt1 maintenance methyltransferase. However, substantial experimental evidence from the past decade indicates that this simple model needs to be revised. DNA methylation can be described by a dynamic stochastic model, in which DNA methylation at each site is determined by the local activity of DNA methyltransferases (Dnmts), DNA demethylases, and the DNA replication rate. Through the targeting and regulation of these enzymes, DNA methylation is controlled by the network of chromatin marks.

 

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Jurkowska & Jeltsch (2013) Genomic Imprinting—The Struggle of the Genders at the Molecular Level. Angewandte Chemie 52, 13524-36

Genomic imprinting, the parent of origin-dependent expression of genes, has been discovered as a fascinating example of the control of gene expression by epigenetic processes in the human body. It affects about 100 genes, which are often involved in growth and development. In this Review, we discuss the mechanisms leading to the generation of gender-specific imprints in form of DNA methylation marks, their preservation during growth and development of the organism, and the processes that translate parental methylation marks into monoallelic gene expression. We discuss the gender-specific dimorphic nature of imprints from an evolutionary point of view and present the prevalent model that molecular imprinting mediates a conflict of interest between the parents that occurs in viviparous animals. Finally, we summarize the relevance of parental imprinting for human health.

 

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Jeltsch (2013) Oxygen, epigenetic signaling and the evolution of early life. Trends in Biochemical Sciences, 38(4):172-6

After approximately 3 billion years of unicellular life on Earth, multicellular animals appeared some 600 million years ago, followed by the rapid emergence of most animal phyla during the Cambrian radiation. This evolutionary jump was paralleled by an increase in atmospheric oxygen, which I propose allowed the generation of epigenetic signaling systems that are essential for cellular differentiation in animals. Epigenetic signaling is based on the reversible deposition of chemically stable marks in DNA and histone proteins, with methylation of cytosine and lysine residues, respectively, playing a central role. Recent evidence indicates that the removal of such methyl groups critically depends on oxygenases. Hence, reversible epigenetic systems could only appear after accumulation of oxygen in the atmosphere.

 

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Till 2012

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Dhayalan et al. (2011) Specificity analysis based identification of new methylation targets of the SET7/9 protein lysine methyltransferase. Chemistry & Biology 18, 111-120

We applied peptide array methylation to determine an optimized target sequence for the SET7/9 (KMT7) protein lysine methyltransferase. Based on this, we identified 91 new peptide substrates from human proteins, many of them better than known substrates. We confirmed methylation of corresponding protein domains in vitro and in vivo with a high success rate for strongly methylated peptides and showed methylation of nine nonhistone proteins (AKA6, CENPC1, MeCP2, MINT, PPARBP, ZDH8, Cullin1, IRF1, and [weakly] TTK) and of H2A and H2B, which more than doubles the number of known SET7/9 targets. SET7/9 is inhibited by phosphorylation of histone and nonhistone substrate proteins. One lysine in the MINT protein is dimethylated in vitro and in vivo demonstrating that the product pattern created by SET7/9 depends on the amino acid sequence context of the target site.

 

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Bock et al. (2011) Detailed specificity analysis of antibodies binding to modified histone tails with peptide arrays. Epigenetics 6, 265-263

Chromatin structure is greatly influenced by histone tail post-translational modifications (PTM), which also play a central role in epigenetic processes. Antibodies against modified histone tails are central research reagents in chromatin biology and molecular epigenetics. We applied Celluspots peptide arrays for the specificity analysis of 36 commercial antibodies from different suppliers which are directed towards modified histone tails. The arrays contained 384 peptides from 8 different regions of the N-terminal tails of histones, viz. H3 1-19, 7-26, 16-35 and 26-45, H4 1-19 and 11-30, H2A 1-19 and H2B 1-19, featuring 59 post-translational modifications in many different combinations. Using various controls we document the reliability of the method. Our analysis revealed previously undocumented details in the specificity profile. Most of the antibodies bound well to the PTM they have been raised for, but some failed. In addition some antibodies showed high cross-reactivity and most antibodies were inhibited by specific additional PTMs close to the primary one. Furthermore, specificity profiles for antibodies directed towards the same modification sometimes were very different. The specificity of antibodies used in epigenetic research is an important issue. We provide a catalog of antibody specificity profiles for 36 widely used commercial histone tail PTM antibodies. Better knowledge about the specificity profiles of antibodies will enable researchers to implement necessary control experiments in biological studies and allow more reliable interpretation of biological experiments using these antibodies.

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Jeltsch (2010) Phylogeny of Methylomes, Science 328, 837-8

The DNA of most species is methylated, containing the modified base 5-methylcytosine. This modification has a role in silencing gene expression, among other important functions. Advances in sequencing methods have allowed measurement of the first complete genome-wide DNA methylation map (“methylome”) of the model plant Arabidopsis thaliana and human cells. Studies by Feng et al. and by Zemach et al. on page 916 of this issue now expand this list by providing genome-wide methylomes for 20 additional species, revealing important conserved features and phylogenetic relationships of the methylation machinery.

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Rathert et al. (2008) Protein lysine methyltransferase G9a acts on non-histone targets. Nat. Chem. Biol. 4, 344-6

By methylation of peptide arrays, we determined the specificity profile of the protein methyltransferase G9a. We show that it mostly recognizes an Arg-Lys sequence and that its activity is inhibited by methylation of the arginine residue. Using the specificity profile, we identified new non-histone protein targets of G9a, including CDYL1, WIZ, ACINUS and G9a (automethylation), as well as peptides derived from CSB. We demonstrate potential downstream signaling pathways for methylation of non-histone proteins.

 

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Rathert et al. (2008) Analysis of the Substrate Specificity of the Dim-5 Histone Lysine Methyltransferases Using Peptide Arrays. Chemistry & Biology 15, 5-11

Histone methylation is an epigenetic mark essential for gene regulation and development. We introduce peptide SPOT synthesis to study sequence specificity of the Dim-5 histone-3 lysine-9 methyltransferase. Dim-5 recognizes R8-G12 of the H3 tail with T11 and G12 being the most important specificity determinants. Exchange of H3 tail residue S10 and T11 by E strongly reduced methylation by Dim-5, suggesting that phosphorylation of S10 or T11 may regulate the activity of Dim-5. In the Dim-5/peptide structure, E227 interacts with H3R8 and D209 with H3-S10. Mutations of E227 or D209 caused predictable changes in the substrate preference, illustrating that peptide recognition of histone methyltransferases can be altered by protein design. Comparative analyses of peptide arrays with wild-type and mutant enzymes, therefore, are well suited to investigate the target specificity of protein methyltransferases and study epigenetic crosstalk.

 

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Jia et al. (2007) Structure of the Dnmt3L-Dnmt3a Complex Suggests a Model for de novo DNA Methylation. Nature 449, 248-51

Genetic imprinting, found in flowering plants and placental mammals, uses DNA methylation to yield gene expression that is dependent on the parent of origin. DNA methyltransferase 3a (Dnmt3a) and its regulatory factor, DNA methyltransferase 3-like protein (Dnmt3L), are both required for the de novo DNA methylation of imprinted genes in mammalian germ cells. Dnmt3L interacts specifically with unmethylated lysine 4 of histone H3 through its amino-terminal PHD (plant homeodomain)-like domain. Here we show, with the use of crystallography, that the carboxy-terminal domain of human Dnmt3L interacts with the catalytic domain of Dnmt3a, demonstrating that Dnmt3L has dual functions of binding the unmethylated histone tail and activating DNA methyltransferase. The complexed C-terminal domains of Dnmt3a and Dnmt3L showed further dimerization through Dnmt3a-Dnmt3a interaction, forming a tetrameric complex with two active sites. Substitution of key non-catalytic residues at the Dnmt3a-Dnmt3L interface or the Dnmt3a-Dnmt3a interface eliminated enzymatic activity. Molecular modelling of a DNA-Dnmt3a dimer indicated that the two active sites are separated by about one DNA helical turn. The C-terminal domain of Dnmt3a oligomerizes on DNA to form a nucleoprotein filament. A periodicity in the activity of Dnmt3a on long DNA revealed a correlation of methylated CpG sites at distances of eight to ten base pairs, indicating that oligomerization leads Dnmt3a to methylate DNA in a periodic pattern. A similar periodicity is observed for the frequency of CpG sites in the differentially methylated regions of 12 maternally imprinted mouse genes. These results suggest a basis for the recognition and methylation of differentially methylated regions in imprinted genes, involving the detection of both nucleosome modification and CpG spacing.

 

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Horton et al. (2005) Transition from non-specific to specific DNA interaction along the DNA recognition pathway of DAM methyltransferase. Cell 121, 349-61

DNA methyltransferases methylate target bases within specific nucleotide sequences. Three structures are described for bacteriophage T4 DNA-adenine methyltransferase (T4Dam) in ternary complexes with partially and fully specific DNA and a methyl-donor analog. We also report the effects of substitutions in the related Escherichia coli DNA methyltransferase (EcoDam), altering residues corresponding to those involved in specific interaction with the canonical GATC target sequence in T4Dam. We have identified two types of protein-DNA interactions: discriminatory contacts, which stabilize the transition state and accelerate methylation of the cognate site, and antidiscriminatory contacts, which do not significantly affect methylation of the cognate site but disfavor activity at noncognate sites. These structures illustrate the transition in enzyme-DNA interaction from nonspecific to specific interaction, suggesting that there is a temporal order for formation of specific contacts.

 

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