Budding yeast Cdc13–Stn1–Ten1 (CST) complex plays an essential role in telomere protection and maintenance. Despite extensive studies, only structural information of individual domains of CST is available; the architecture of CST still remains unclear. Here, we report crystal structures of Kluyveromyces lactis Cdc13–telomeric-DNA, Cdc13–Stn1 and Stn1–Ten1 complexes and propose an integrated model depicting how CST assembles and plays its roles at telomeres. Surprisingly, two oligonucleotide/oligosaccharide-binding (OB) folds of Cdc13 (OB2 and OB4), previously believed to mediate Cdc13 homodimerization, actually form a stable intramolecular interaction. This OB2–OB4 module of Cdc13 is required for the Cdc13–Stn1 interaction that assembles CST into an architecture with a central ring-like core and multiple peripheral modules in a 2:2:2 stoichiometry. Functional analyses indicate that this unique CST architecture is essential for both telomere capping and homeostasis regulation. Overall, our results provide fundamentally valuable structural information regarding the CST complex and its roles in telomere biology.

酵母Cdc13–Stn1–Ten1（CST）复合芽在端粒保护和维持中起着至关重要的作用。尽管进行了广泛的研究，但仅提供了CST各个领域的结构信息；CST的架构仍不清楚。在这里，我们报告乳酸克鲁维酵母的晶体结构Cdc13–端粒DNA，Cdc13–Stn1和Stn1–Ten1复合物，并提出了一个集成模型，描述了CST如何组装并在端粒中发挥作用。出人意料的是，以前认为可以介导Cdc13均二聚的Cdc13的两个寡核苷酸/寡糖结合（OB）折叠（OB2和OB4）实际上形成了稳定的分子内相互作用。Cdc13–Stn1交互将CST组装到具有中心环状核心和多个化学计量比为2的多个外围模块的体系结构中，此Cdc13的OB2-OB4模块是必需的。功能分析表明，这种独特的CST体系结构对于端粒封端和体内平衡调节至关重要。总体而言，我们的结果提供了有关CST复合物及其在端粒生物学中的作用的根本有价值的结构信息。