Area A: Modifications on Nucleuc Acids

Project A01: S. Kellner
Discovery of novel RNA modifications and targets of RNA writer enzymes

We will continue our efforts for the discovery of novel modified nucleosides as we believe that the function of RNA modification can only be understood once the complete network of modifications in an organism is known. With isotope labeling in cell culture we will establish a systematic approach to identify RNA writers and erasers and generate internal standards for modification profile analysis. We plan to study the effect of unnatural nucleosides and nucleic acid metabolism inhibitors on the adaptation of modified nucleosides.

Project A02: M. Sattler
Effects of posttranscriptional chemical modifications on RNA structure, dynamics and protein interactions

The molecular, structural and functional consequences of chemical RNA modifications involving adenosine-to-inosine (A-to-I) editing and m6A RNA methylation are studied. Using biochemical methods, solution-state NMR and complementary techniques the effects of A-to-I-editing and base m6A methylation on the conformation, internal dynamics and molecular interactions will be studied. The results will provide unprecedented insight and high-resolution structural information for the effects of chemical modifications of eukaryotic RNAs with important functions in posttranscriptional gene regulation.

Project A03: R. Linser, P. Ro
The effect of DNA modifications on the stability and mobility of nucleosomes

In addition to the canonical DNA bases chemically modified bases have been found recently. In particular for 5-fC a role in epigenetics is certain. How this modification affects gene expression is so-far elusive. On one hand, we will characterize potential changes in structure and dynamics of DNA double strands upon inclusion of 5-fC. On the other, changes in DNA-histone interactions in nucleosomes due to 5-fC will be determined. For these goals, we use NMR spectroscopy in solution and in the solid state.

Project A04: T. Carell
Molecular mechanism of active demethylation

Since 2009 three new DNA bases (5-hydroxymethyl-, 5-formyl-, and 5-carboxy-cytidine, hmdC, fdC and cadC) were discovered that are present in the DNA isolated from stem cells and neurons. We believe that fdC and cadC undergo deformylation and decarboxylation reactions to establish a new, long search for, pathway to active demethylation. In this project, we plan to gain direct in vivo evidence for the existence of the putative decarboxylation reaction and we plan to make the first steps towards identifying the decarboxylating entity in stem cells. In the second part of the project, we plan to develop new, robust sequencing methods that will allow us to determine the positions of hmdC and fdC in the genome.

Project A05: J. Walter
Sequence specific and functional analysis of oxdative modifications in nucleic acids – implication for epigenetic memory in the mouse

Tet mediated oxidative modifications of 5-methylcytosine (5mC) regulate and modulate short term and long term effect of the 5mC mediated epigenetic memory formation. In this project we will analyse the formation and regulation of 5fC and 5hmC in the mouse zygote, ES cells and primary neurons of newborn and adult mice. We will develop novel methods for base specific detection of 5fC and 5hmC and will use these methods to determine the functional consequences of a loss of distinct Tet3 isoforms using NGS data based modeling.

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