3D chromatin organisation in direct neuronal reprogramming – reshaping the nucleus to mold new neurons

How transcription factors can rewire 3D genome architecture and how such regulation affects cell fate decisions is one of the fundamental questions in chromatin biology. Reprogramming of glial cells into neurons by a single neurogenic factor provides an excellent system to study this phenomenon and to determine the global changes in chromatin architecture which occur in direct neuronal reprogramming. To tackle this fascinating project Boyan Bonev, an expert in chromatin architecture and high resolution Hi-C and Magdalena Götz, who pioneered the glia-to-neuron reprogramming, propose to combine their unique expertise and dissect the relationship between 3D nuclear architecture, chromatin accessibility and gene expression during neuronal reprogramming. We first aim to determine the 3D genome organization genome-wide in astrocytes and induced neurons using high-resolution Hi-C. We will also exploit an in vitro model of astrocytes that are more resistant to reprogramming to determine if global changes in chromatin compaction and long-range interactions underlie this lack of plasticity. We will then explore how transcriptional changes (RNA-seq data have been generated already) correspond to changes in chromatin looping and examine by Hi-ChIP to which extent direct binding of the reprogramming factor (Neurogenin2) is involved in re-shaping chromatin architecture. Furthermore, using genome engineering to tether Ngn2 to specific loci and delete Ngn2-bound enhancers, we will determine if Ngn2 binding is necessary and/or sufficient to cause chromatin looping. In the last part of the proposal, we will examine if YY1 is important for formation of cell type specific regulatory interactions during neuronal reprogramming and use ChIP-MS and locus-specific proteomics approach to identify other factors relevant for rewiring the chromatin during glia-to-neuron reprogramming. Taken together, this project will not only yield unprecedented insights into how a single TF can reorganize 3D chromatin architecture, but also help to improve this process to perfect the induced neurons.