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. The cohesin protein complex plays a key role in the formation and dynamics of DNA loops in the cell nucleus. How these loops guide genes to distal enhancers and influence their expression remains unclear. We depleted two opposing regulators of cohesin activity, NIPBL and WAPL, to compare their independent and shared effects on nascent gene expression in human cells. We show patterns of dysregulation that are both unique and highly similar between the depletions, with some large genomic regions showing amplified sensitivity to both. Finally, we show that co-depletion of NIPBL and WAPL restores the majority of gene misexpression as compared to either knockdown alone. This indicates that the balance between the factors, rather than their absolute levels, is especially important for regulating cohesin dynamics and DNA looping that can influence gene expression.

黏连蛋白介导的染色质环化与基因表达之间的关系仍不明确。NIPBL和WAPL是黏连蛋白活性的两种相反的调节因子；在多种细胞类型中，二者中任何一种的缺失都与染色质折叠和转录的变化相关。然而，缺乏在同一细胞类型中对它们各自以及联合作用于基因表达的直接比较。我们发现，在人HCT116细胞中，NIPBL或WAPL的缺失各自改变了约2000个基因的表达，且在两种条件下只有约30%的基因是相同的。我们发现，在同一拓扑相关结构域（TAD）内的差异表达基因簇呈现出协同的错误表达，这表明一些基因组结构域对黏连蛋白的增多或减少尤其敏感。最后，与单独敲低其中任何一个相比，同时敲低NIPBL和WAPL可恢复大多数基因的错误表达。在同时敲低CTCF后，一组类似的对NIPBL敏感的基因得到恢复。总之，这表明由于黏连蛋白活性降低而导致的转录改变，可以通过去除其负调节因子（WAPL）或限制环挤压事件的物理屏障（CTCF）在功能上得到补偿。 黏连蛋白复合物在细胞核中DNA环的形成和动态变化中起关键作用。这些环如何引导基因与远端增强子结合并影响其表达仍不明确。我们去除了黏连蛋白活性的两种相反的调节因子NIPBL和WAPL，以比较它们对人细胞中新生基因表达的独立作用和共同作用。我们展示了在去除这两种因子时独特且高度相似的失调模式，一些大的基因组区域对二者都表现出增强的敏感性。最后，我们表明与单独敲低其中任何一个相比，同时敲低NIPBL和WAPL可恢复大多数基因的错误表达。这表明这些因子之间的平衡，而非它们的绝对水平，对于调节黏连蛋白动态变化和可影响基因表达的DNA环化尤为重要。