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Cohesin is a ring-shaped protein complex that controls dynamic chromosome structure. Cohesin activity is important for a variety of biological processes, including formation of DNA loops that regulate gene expression. The precise mechanisms by which cohesin shapes local chromosome structure and gene expression are not fully understood. Recurrent mutations in cohesin complex members have been reported in various cancers, though it is not clear whether many cohesin sequence variants have phenotypes and contribute to disease. Here, we utilized CRISPR/Cas9 genome editing to introduce a variety of cohesin sequence variants into murine embryonic stem cells and investigate their molecular and cellular consequences. Some of the cohesin variants tested caused changes to transcription, including altered expression of gene encoding lineage-specifying developmental regulators. Altered gene expression was also observed at insulated neighborhoods, where cohesin-mediated DNA loops constrain potential interactions between genes and enhancers. Furthermore, some cohesin variants altered the proliferation rate and differentiation potential of murine embryonic stem cells. This study provides a functional comparison of cohesin variants found in cancer within an isogenic system, revealing the relative roles of various cohesin perturbations on gene expression and maintenance of cellular identity. [ABSTRACT FROM AUTHOR]

The cohesin complex spatially organizes interphase chromatin by bringing distal genomic loci into close physical proximity, looping out the intervening DNA. Mutation of cohesin complex subunits is observed in cancer and developmental disorders, but the mechanisms through which these mutations may contribute to disease remain poorly understood. Here, we investigate a recurrent missense mutation to the hinge domain of the cohesin subunit SMC1A, observed in acute myeloid leukemia. Engineering this mutation into murine embryonic stem cells caused widespread changes in gene expression, including dysregulation of the pluripotency gene expression program. This mutation reduced cohesin levels at promoters and enhancers, decreased DNA loops and interactions across short genomic distances, and weakened insulation at CTCF-mediated DNA loops. These findings provide insight into how altered cohesin function contributes to disease and identify a requirement for the cohesin hinge domain in three-dimensional chromatin structure. Author summary: Mammalian genomes consist of multiple meters of DNA which must be highly folded in order to fit inside of the nucleus. This folding is regulated at multiple scales by different biological mechanisms. The spatial organization of the genome is closely linked to its function, including the spatial and temporal expression of genes. Especially important for gene control is the partitioning of chromosomes into DNA loops, which are formed when two distal loci are brought into close contact. The folding of the genome into DNA loops is performed by cohesin and CTCF. The molecular basis for how DNA loops dynamically form and function in gene control is poorly understood. Here, we investigate a recurrent cancer mutation in cohesin and show that it causes altered folding of the genome into DNA loops and misexpression of many genes. This finding is important because cohesin mutations are common in many cancers and yet there is little understanding of how cohesin defects may contribute to disease. [ABSTRACT FROM AUTHOR]

Chromosome structure is a key regulator of gene expression. CTCF and cohesin play critical roles in structuring chromosomes by mediating physical interactions between distant genomic sites. The resulting DNA loops often contain genes and their cis -regulatory elements. Despite the importance of DNA loops in maintaining proper transcriptional regulation and cell identity, there is limited understanding of the molecular mechanisms that regulate their dynamics and function. We report a previously unrecognized role for WIZ (widely interspaced zinc finger-containing protein) in DNA loop architecture and regulation of gene expression. WIZ forms a complex with cohesin and CTCF that occupies enhancers, promoters, insulators, and anchors of DNA loops. Aberrant WIZ function alters cohesin occupancy and increases the number of DNA loop structures in the genome. WIZ is required for proper gene expression and transcriptional insulation. Our results uncover an unexpected role for WIZ in DNA loop architecture, transcriptional control, and maintenance of cell identity. • WIZ generally colocalizes with CTCF and cohesin across the genome • Loss of WIZ increases cohesin occupancy and DNA loops • WIZ maintains proper gene expression and stem cell identity Justice et al. show that WIZ functions with CTCF and cohesin in the structural regulation of DNA loops. Aberrant WIZ function causes many changes in gene expression, including at DNA loops important for regulating stem cell identity genes. [ABSTRACT FROM AUTHOR]