Understanding how changes in spatial chromatin organization cause defective gene expression in acute myeloid leukemia with cohesin mutations.
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Multiple large-scale sequencing projects have identified a large number of mutations recurrently acquired in acute myeloid leukemia (AML). Mutations often cause defective gene regulation and consequently AML subtypes can be recognized based on unique gene expression signatures. Recently it has been shown that mutations may not only occur in coding regions of genes, but also in elements that regulate their expression. Enhancer dislocation as well as mutations that either generate or destroy enhancers have been reported in cancer. In this application I propose to investigate other mechanisms of altered gene regulation by changes in enhancer-to-promoter association, i.e. in AML with mutations in cohesin genes. Members of the cohesin complex have been reported to be mutated in ~15% of AML patients. Since cohesin plays a critical role in the alignment and stabilization of replicated chromosomes, one would expect that mutations in genes encoding for cohesin members are particularly found in leukemias with severe chromosomal defects. However, the opposite appears to be true, as most AML patients with mutations in members of the cohesin complex have a normal karyotype. Therefore these mutant protein members are likely to drive tumorigenesis through other functions of cohesin, i.e. via regulation of three-dimensional chromatin organization, looping and insulation of topologically associated domains. Cohesin consequently shapes genomewide gene expression patterns, regulating cell-type specific gene sets that are essential for healthy hematopoietic development. Cohesin complexes carry one of the two versions of the SA subunit, namely STAG1 (SA1) or STAG2 (SA2), respectively. SA1-cohesin complex has been reported to be particularly involved in stabilization of TADs, via the interaction with CTCF. SA2- cohesin complexes preferentially support interactions between celltype specific enhancers and promoters. These SA2 interactions, may either involve CTCF proteins or they act via the Mediator complex, of which MED12 is key in human hematopoietic progenitor cells (HSPCs). Mutations have been reported in several cohesin members in AML, in particular in STAG1, STAG2 or RAD21. We hypothesize that these mutations affect the cohesin complex formation, the chromatin structure and consequently enhancer-to-promoter interactions, which will cause changes in gene expression. In this proposal we will study the potential differences and functions between the two variant cohesin complexes in HSPCs as well as the consequences of their specific downregulation in spatial chromatin architecture and gene expression. Furthermore we will study the consequences of mutations in cohesin members on chromatin structure and gene expression in cells from patients with AML. The results obtained from this project will reveal fundamental insights into the effect of mutated SA1- and SA-2 cohesin complexes on 3D chromatin folding and gene regulation in primary AML cells.