Epigenetic control of Transcription Factor activity

• Applicant: Arnaud Krebs, Ph.D.
• Subject Area: General Genetics and Functional Genome Biology
• Term: from 2019 to 2023
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 418196409

Successful development of complex organisms relies on accurate execution of transcriptional programs. Binding of transcription factors (TFs) to cis-regulatory elements (CREs) translates genetic information into appropriate gene expression patterns. In Eukaryotes, DNA is wrapped around histones representing a physical barrier preventing the spurious binding of TFs. The compaction level of chromatin is tightly regulated by multiple processes including post-translational modification of histones. Therefore activity of a CREs is defined by the interpretation of its genetic information by TFs as well as the epigenetic signals present in its chromatin environment. It is commonly assumed that interactions between these two regulatory layers is a key to precisely control complex gene expression patterns required for development. However, the mechanisms by which chromatin modifications influence the binding of TFs are largely unknown. If binding of certain TFs depends on the presence of histone modifications at their target regions, these TFs should preferentially associate with chromatin molecules carrying that modification. Based on this assumption, we propose to use the levels of co-occupancy between chromatin modifications and TFs as mean to identify dependency between them. Current approaches used to probe presence of chromatin marks and binding of TFs require to average signals arising from millions of cells. Consequently, these assays only allow to measure the relative abundance of each feature individually and therefore do not inform on the frequency of co-occurrence of histone modifications and binding of TFs at the molecular level. This proposal aims to move beyond this boundary by developing a novel genomics assay that probes TFs binding and presence of chromatin modifications in the genome simultaneously at the resolution of single DNA molecules. We will apply this protocol to a set of chromatin modifications known to positively or negatively correlate with the activity of CREs. The resulting genome-wide single molecule data will reveal if presence of these chromatin marks associate with higher or lower frequency of TF binding at CREs. We will define putative dependencies based on these interaction data and build a network describing relations between chromatin marks and TFs in pluripotent mouse embryonic stem cells and its rewiring during their commitment to a neuronal lineage. Importantly, we will further experimentally test the functional relevance of the identified dependencies by measuring the effect of global and targeted perturbation of chromatin modification levels through chemical inhibition, genetic deletions and epigenome editing. Together, the proposed work will characterise how epigenetic modifications contribute to transcriptional control by modulating the access of TFs to their target CREs in the context of cell fate choices.

• DFG Programme: Research Grants