Our research centers around the question: how are genes regulated within the context of the three-dimensional (3D) genome? We use a combination of genetic and acute perturbation experiments in combination with genomics tools to understand how distal regulatory elements (e.g. enhancers) contribute to the regulation of genes. In addition to implementing and developing genomics methods we also develop software for the analysis of chromosome conformation capture data.
It is becoming increasingly appreciated that chromatin inside the nucleus is highly dynamic. Both the proteins binding to DNA and the DNA itself are in a constant flux. These dynamic processes contribute to the regulation of genes. However, because of the highly dynamic nature of nuclear organization it is difficult to determine cause and effect. The reason for this is that whereas the folding of chromosomes influences expression, expression in turn also influences the folding of chromosomes. In order to study the consequences of changes in genome folding we now make use of methods that can acutely (<1hr) and uniformly deplete a protein from cells. This is achieved by fusing a degron tag to a protein of interest which is sensitive to a small molecule. Following the addition of the small molecule the protein is then rapidly degraded.
We have performed acute depletion in mouse embryonic stem cells of a number of proteins that are important for nuclear organization. Our experiments show a rapid change in nuclear organization, reshaping the nucleus within a few hours. Changes in the 3D genome are followed by changes in gene expression. Our experiments are consistent with a model in which the 3D genome, driven by cohesin, is crucial for maintaining proper gene expression and the maintenance of the cellular state.
A key event in the differentiation of multi-cellular organisms is gastrulation. During gastrulation the blastula which is single-layered and symmetrical reorganizes itself into a multilayered and asymmetric gastrula. The process of symmetry breaking leads to the establishment of different cell types in the early embryo. The gene regulatory factors that control this process in mammalian cell systems are still not fully delineated. This is in part due to the difficulty to obtain sufficient material from in utero or in vitro developing embryos. However, this can be overcome by using an in vitro culture system that uses embryonic stem cells that aggregate into so-called gastruloids. These undergo the process of symmetry breaking in vitro.
Gastruloids will allow us to study the role of the 3D genome during the process of gastrulation. By making gastruloids of our acute depletion mouse embryonic stem cell lines, we can influence key regulators of expression during early development and delineate the key transcriptional response. We have set up single cell ATACseq that allows us to measure the open chromatin landscape of single cells, enabling us to deconvolve the different cell types and their regulators in gastruloids.