High-throughput Oligopaint screen identifies druggable 3D genome regulators.

The human genome functions as a three-dimensional chromatin polymer, driven by a complex collection of chromosome interactions1-3. Although the molecular rules governing these interactions are being quickly elucidated, relatively few proteins regulating this process have been identified. Here, to address this gap, we developed high-throughput DNA or RNA labelling with optimized Oligopaints (HiDRO)-an automated imaging pipeline that enables the quantitative measurement of chromatin interactions in single cells across thousands of samples. By screening the human druggable genome, we identified more than 300 factors that influence genome folding during interphase. Among these, 43 genes were validated as either increasing or decreasing interactions between topologically associating domains. Our findings show that genetic or chemical inhibition of the ubiquitous kinase GSK3A leads to increased long-range chromatin looping interactions in a genome-wide and cohesin-dependent manner. These results demonstrate the importance of GSK3A signalling in nuclear architecture and the use of HiDRO for identifying mechanisms of spatial genome organization.

Co-depletion of NIPBL and WAPL balance cohesin activity to correct gene misexpression.

The relationship between cohesin-mediated chromatin looping and gene expression remains unclear. NIPBL and WAPL are two opposing regulators of cohesin activity; depletion of either is associated with changes in both chromatin folding and transcription across a wide range of cell types. However, a direct comparison of their individual and combined effects on gene expression in the same cell type is lacking. We find that NIPBL or WAPL depletion in human HCT116 cells each alter the expression of ~2,000 genes, with only ~30% of the genes shared between the conditions. We find that clusters of differentially expressed genes within the same topologically associated domain (TAD) show coordinated misexpression, suggesting some genomic domains are especially sensitive to both more or less cohesin. Finally, co-depletion of NIPBL and WAPL restores the majority of gene misexpression as compared to either knockdown alone. A similar set of NIPBL-sensitive genes are rescued following CTCF co-depletion. Together, this indicates that altered transcription due to reduced cohesin activity can be functionally offset by removal of either its negative regulator (WAPL) or the physical barriers (CTCF) that restrict loop-extrusion events.

Out of destruction comes new growth: Pore-forming antimicrobials make pancreas grow.

Gut-residing bacteria are known to regulate the physiologies of distal organs. However, the mechanism behind the long-distance communication between gut microbes and distal organs remains unknown. In this issue of Cell Metabolism, two studies show that ß cell expansion in the pancreas depends on bacterially induced antimicrobials produced in the gut.

Cohesin promotes stochastic domain intermingling to ensure proper regulation of boundary-proximal genes.

The human genome can be segmented into topologically associating domains (TADs), which have been proposed to spatially sequester genes and regulatory elements through chromatin looping. Interactions between TADs have also been suggested, presumably because of variable boundary positions across individual cells. However, the nature, extent and consequence of these dynamic boundaries remain unclear. Here, we combine high-resolution imaging with Oligopaint technology to quantify the interaction frequencies across both weak and strong boundaries. We find that chromatin intermingling across population-defined boundaries is widespread but that the extent of permissibility is locus-specific. Cohesin depletion, which abolishes domain formation at the population level, does not induce ectopic interactions but instead reduces interactions across all boundaries tested. In contrast, WAPL or CTCF depletion increases inter-domain contacts in a cohesin-dependent manner. Reduced chromatin intermingling due to cohesin loss affects the topology and transcriptional bursting frequencies of genes near boundaries. We propose that cohesin occasionally bypasses boundaries to promote incorporation of boundary-proximal genes into neighboring domains.