Area B: Modifications on Proteins

Project B01: A. Hoffmann-Röder
O-GlcNAc profiling and epigenetic regulation

The dynamic O-GlcNAcylation at specific protein sites is controlled by the two enzymes Ogt and Oga, with the former closely interacting with Tet proteins during epigenetic regulation. Since very little is known about this O-GlcNAc-based Ogt-Tet cross-talk, we plan to map O-GlcNAcylated proteins at specific cellular differentiation stages using chemical labeling and modern MS-based GlcNAcomic techniques. Thus, by developing novel metabolic labels and analyzing specific O-GlcNAcylation profiles of Tet proteins and their binding partners, we hope to decipher the molecular basics of these regulative Ogt-Tet interactions.

Project B02: B. Küster
Dynamics of epigenetic protein modifications and RNA interactions

In this project, we ask two main questions: first, do post-synthetic modifications influence the interaction of proteins with mRNAs or miRNAs to modulate their function? Second, what are the post-translational modifications that characterize differentiating stem cells and how can we exploit this chemically to kill cancer stem cells. Both questions will be approached by a combination of mass spectrometry based quantitative proteomics and bioinformatics.

Project B03: A. Imhof
Dynamics and function of posttranslational protein methylation and acetylation

The methylation and acetylation of lysine residues within proteins that regulate gene expression has been shown to play a key role in various diseases and the enzymes that catalyze these modifications have therefore gained considerable interest as potential drug targets. Surprisingly little is known about the proteins that are modified, the regulation of the enzymes, the dynamics of their turnover or their functional implications. Within this project we plan to use a variety of biochemical strategies to determine the complete lysine methylome of a human cell line. We will furthermore investigate the substrate specificity of a set of putative methyltransferases by applying a combination of different in vitro and in vivo assays. In addition, we will establish MALDI imaging as a new technology to investigate the distribution of inhibitors targeting such modifying enzymes and simultaneously the corresponding histone proteoform within tissues. Finally, we aim to use MALDI imaging as a new and antibody independent way to detect, quantify and characterize modified nucleic acids in situ.

Project B04: A. Ladurner
PARP1-mediated recruitment of nucleic acid regulators and their function in DNA demethylation and the circadian clock

DNA damage recruits nuclear factors to chromatin, often in a PARP1-dependent manner. This includes Cryptochrome 1 (CRY1), a circadian clock component, METTL3/14, the m6A methyltransferase METTL3, and ATP-dependent chromatin remodelers. Moreover, PARP1 and the PAR-dependent remodeler ALC1/CHD1L promote cell reprogramming by Yamanaka factors and DNA demethylation. We will establish the mechanism(-s) through which PARylation promotes recruitment and test the role of PARylation in DNA demethylation and circadian gene expression.

Project B05: S. Michalakis
Role of TET3-mediated 5mC oxidation for neuronal differentiation and plasticity

The physiological significance of TET3 proteins and their enzymatic products in the CNS has not been characterized and will be addressed with specific genetic mouse and cellular models in Aim A. TET enzymes act in concert with chromatin remodeling proteins and transcription factors. In Aim B we will assess the potential of interacting proteins to engage with TET3 and modulate its enzymatic activity. The genomic 5hmC and 5fC content in neurons varies depending on differentiation and/or activity state. In Aim C we will explore the underlying mechanism of this phenomenon.

Project B06: R. Schneider
Lysine acetylation and succinylation in the core of the nucleosome

Histones form the building blocks of chromatin. Covalent modifications of these histone proteins regulate all DNA dependent processes. Currently very little is known about modifications within the core of the nucleosome. We want to study the role of lysine acetylations in the nucleosomal core and in particular combinations of these acetylations in chromatin dynamics and transcription. Furthermore, we want to unravel the function of a novel acylation, lysine succinylation. Together this will give us new insights in how PTMs can regulate chromatin function.

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