In vertebrates, epithelial permeability is regulated by the tight junction (TJ) formed by specialized adhesive membrane proteins, adaptor proteins, and the actin cytoskeleton. Despite the TJ’s critical physiological role, a molecular-level understanding of how TJ assembly sets the permeability of epithelial tissue is lacking. Here, we identify a 28-amino-acid sequence in the TJ adaptor protein ZO-1, which is responsible for actin binding, and show that this interaction is essential for TJ permeability. In contrast to the strong interactions at the adherens junction, we find that the affinity between ZO-1 and actin is surprisingly weak, and we propose a model based on kinetic trapping to explain how affinity could affect TJ assembly. Finally, by tuning the affinity of ZO-1 to actin, we demonstrate that epithelial monolayers can be engineered with a spectrum of permeabilities, which points to a promising target for treating transport disorders and improving drug delivery.

在脊椎动物中，上皮细胞的通透性受紧密连接 (TJ) 的调节，紧密连接 (TJ) 由专门的粘附膜蛋白、衔接蛋白和肌动蛋白细胞骨架形成。尽管 TJ 具有重要的生理作用，但缺乏对 TJ 组装如何设置上皮组织渗透性的分子水平理解。在这里，我们确定了负责肌动蛋白结合的 TJ 衔接蛋白 ZO-1 中的 28 个氨基酸序列，并表明这种相互作用对于 TJ 通透性至关重要。与粘附连接处的强相互作用相反，我们发现 ZO-1 和肌动蛋白之间的亲和力出奇地弱，我们提出了一个基于动力学捕获的模型来解释亲和力如何影响 TJ 组装。最后，通过调整 ZO-1 与肌动蛋白的亲和力，