EmbryoPAINT · PAINTing the architecture of the totipotency gene network during early mammalian development

The three-dimensional architecture of the genome regulates its fundamental functions such as the transcription or replication of DNA. Thus, chromatin organisation is crucially important for key aspects of cell biology, such as the differentiation of stem cells in the early embryo. While recent studies have shown that the mammalian genome rearranges extensively towards a more ordered state after the first few embryonal divisions, many fundamental questions remain unanswered. For example, it is not known whether totipotent cells have a well-defined genomic architecture or whether this architecture is highly heterogeneous between different cells and embryos. Further, it is unclear if early cell fate decisions are driven by a reproducible coordinated rearrangement of pluripotency-related genes, or if this is stochastic process. These questions could best be tackled by directly assessing the physical genome structure and architecture of pluripotency genes in single stem cells inside the whole embryo. In my project, I will pursue this ambitious aim by exploiting recent breakthroughs in 3D super-resolution microscopy, namely the development of an inverted lattice light-sheet microscope, highly multiplexed oligo-DNA-PAINT, and advanced computational algorithms, to study the physical 3D architecture of the genomic network of totipotency and pluripotency genes. Thus, I will for the first time be able to unravel the structural determinants of the transition from totipotency to the pluripotent and differentiated state during early mammalian development.