The DNA-binding protein CCCTC-binding factor (CTCF) and the cohesin complex function together to shape chromatin architecture in mammalian cells, but the molecular details of this process remain unclear. Here, we demonstrate that a 79-aa region within the CTCF N terminus is essential for cohesin positioning at CTCF binding sites and chromatin loop formation. However, the N terminus of CTCF fused to artificial zinc fingers was not sufficient to redirect cohesin to non-CTCF binding sites, indicating a lack of an autonomously functioning domain in CTCF responsible for cohesin positioning. BORIS (CTCFL), a germline-specific paralog of CTCF, was unable to anchor cohesin to CTCF DNA binding sites. Furthermore, CTCF-BORIS chimeric constructs provided evidence that, besides the N terminus of CTCF, the first two CTCF zinc fingers, and likely the 3D geometry of CTCF-DNA complexes, are also involved in cohesin retention. Based on this knowledge, we were able to convert BORIS into CTCF with respect to cohesin positioning, thus providing additional molecular details of the ability of CTCF to retain cohesin. Taken together, our data provide insight into the process by which DNA-bound CTCF constrains cohesin movement to shape spatiotemporal genome organization.

中文翻译：

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CTCF 通过 N 端结构域依赖性粘连蛋白保留介导染色质环化。


DNA 结合蛋白 CCCTC 结合因子 (CTCF) 和粘连蛋白复合物共同发挥作用，塑造哺乳动物细胞中的染色质结构，但这一过程的分子细节仍不清楚。在这里，我们证明 CTCF N 末端内的 79 个氨基酸区域对于粘连蛋白在 CTCF 结合位点的定位和染色质环的形成至关重要。然而，与人工锌指融合的 CTCF 的 N 末端不足以将粘连蛋白重定向到非 CTCF 结合位点，表明 CTCF 中缺乏负责粘连蛋白定位的自主功能结构域。 BORIS (CTCFL) 是 CTCF 的种系特异性旁系同源物，无法将粘连蛋白锚定到 CTCF DNA 结合位点。此外，CTCF-BORIS 嵌合构建体提供的证据表明，除了 CTCF 的 N 末端之外，前两个 CTCF 锌指以及可能的 CTCF-DNA 复合物的 3D 几何形状也参与粘连蛋白保留。基于这些知识，我们能够将 BORIS 转化为关于粘连蛋白定位的 CTCF，从而提供 CTCF 保留粘连蛋白能力的更多分子细节。总而言之，我们的数据提供了对 DNA 结合 CTCF 限制粘连蛋白运动以塑造时空基因组组织的过程的深入了解。