Claudin-based tight junctions (TJs) are formed at the most apical part of cell–cell contacts in epithelial cells. Previous studies suggest that scaffolding proteins ZO-1 and ZO-2 (ZO proteins) determine the location of TJs by interacting with claudins, but this idea is not conclusive. To address the role of the ZO proteins binding to claudins at TJs, a COOH-terminal PDZ domain binding motif-deleted claudin-3 mutant, which lacks the ZO protein binding, was stably expressed in claudin-deficient MDCK cells. The COOH-terminus-deleted claudin-3 was localized at the apicolateral region similar to full-length claudin-3. Consistently, freeze-fracture electron microscopy revealed that the COOH-terminus-deleted claudin-3-expressing cells reconstituted belts of TJs at the most apical region of the lateral membrane and restored functional epithelial barriers. These results suggest that the interaction of claudins with ZO proteins is not a prerequisite for TJ formation at the most apical part of cell–cell contacts.

中文翻译：

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由缺乏 COOH 末端 PDZ 结构域结合基序的 claudin 突变体形成紧密连接

基于密连接蛋白的紧密连接 (TJ) 形成于上皮细胞中细胞-细胞接触的最顶端部分。之前的研究表明，支架蛋白 ZO-1 和 ZO-2（ZO 蛋白）通过与 claudins 的相互作用来决定 TJ 的位置，但这种想法并不是决定性的。为了解决 ZO 蛋白在 TJ 处与密蛋白结合的作用，COOH 末端 PDZ 结构域结合基序缺失的 claudin-3 突变体在密蛋白缺陷型 MDCK 细胞中稳定表达，该突变体缺乏 ZO 蛋白结合。COOH 末端缺失的 claudin-3 位于类似于全长 claudin-3 的顶外侧区域。一致地，冷冻断裂电子显微镜显示，COOH 末端缺失的 claudin-3 表达细胞在侧膜最顶端区域重建了 TJ 带，并恢复了功能性上皮屏障。