Cohesin chromatin loop formation by an extrinsic motor (ChIP-Seq)

The ring-shaped cohesin complex topologically entraps two DNAs to establish sister chromatid cohesion. Cohesin also shapes the interphase chromatin landscape, with wide-ranging implications for gene regulation, which cohesin is thought to achieve by actively extruding DNA loops without topologically entrapping DNA. The 'loop extrusion' model find motivation from in vitro observations - whether this process indeed underlies chromatin loop formation in vivo remains untested. Here, using the budding yeast S. cerevisiae, we generate cohesin variants that have lost their ability to extrude DNA loops but retain their ability to topologically entrap DNA. Analysis of these variants suggests that in vivo chromatin loops form independently of loop extrusion. Instead, we find that transcription promotes loop formation, likely by generating DNA substrates for topological loop capture. Transcription furthermore acts as an extrinsic motor that, by pushing cohesin along transcription units, extends chromatin loops and defines their ultimate positions. Our results necessitate a re-evaluation of the loop extrusion hypothesis and point to an alternative mechanism for cohesin-dependent chromatin organisation. Loop formation by DNA-DNA capture, akin to sister chromatid cohesion establishment at replication forks, unifies cohesin's two roles in chromosome segregation and interphase genome organisation. Overall design: Cohesin calibrated ChIP sequencing and Micro-C. Two independent replicates are provided

环形黏连蛋白（cohesin）复合物可通过拓扑捕获两条DNA分子，建立姐妹染色单体黏连（sister chromatid cohesion）。黏连蛋白同时塑造间期染色质的三维构象，对基因调控具有广泛影响，此前研究认为，黏连蛋白可通过不依赖DNA拓扑捕获的主动DNA环挤出过程，实现上述功能。“环挤出（loop extrusion）”模型的理论依据源于体外实验观察，但该过程是否确实是体内染色质环形成的核心机制，目前仍未得到验证。本研究以酿酒酵母（S. cerevisiae）为模式生物，构建了丧失DNA环挤出能力但仍保留DNA拓扑捕获功能的黏连蛋白变体。对这些变体的分析结果表明，体内染色质环的形成并不依赖环挤出过程。与之相反，我们发现转录过程可促进染色质环的形成，其机制可能是为拓扑环捕获提供DNA底物。此外，转录还可作为一种外在动力，通过推动黏连蛋白沿转录单元移动，延长染色质环并确定其最终位置。我们的研究结果要求对环挤出假说进行重新评估，并提出了依赖黏连蛋白的染色质组织的替代机制。通过DNA-DNA捕获形成染色质环的方式，与复制叉处姐妹染色单体黏连的建立机制类似，统一了黏连蛋白在染色体分离和间期基因组组织中的两项核心功能。整体实验设计：黏连蛋白校准型染色质免疫共沉淀测序（cohesin calibrated ChIP sequencing）与Micro-C测序，本研究提供了两组独立重复样本。