Junction-binding–defective Cdc13 promotes adaptive mutation and transcriptomic rewiring under telomere stress

Telomeres protect chromosome ends from degradation and inappropriate DNA repair. In budding yeast, the telomeric protein Cdc13 binds G-rich overhangs to maintain telomere integrity. Here, we show that Cdc13 also weakly interacts with adjacent duplex DNA at the ss/dsDNA junction, a property essential for inhibiting Exo1- and Sgs1–Dna2–mediated resection. Mutation of a conserved residue (K504E) disrupts this junction recognition, impairing resection inhibition, RPA displacement, and cooperation with Ku. cdc13-K504E cells are hypersensitive to Exo1 overexpression and accumulate non-pigmented revertant colonies during chronic growth. These white colonies exhibit faster growth, altered telomere-proximal gene expression, upregulation of metabolic pathways, and increased 4N DNA content. Our findings identify a dual DNA-binding mode of Cdc13 that protects telomeres by blocking resection and regulating Ku positioning, and reveal a stress-induced adaptive state associated with telomere dysfunction. Overall design: 3 biological replicates per condition; RNA-seq of WT-red, K504E-red, K504E-white colonies.

端粒（Telomeres）可保护染色体末端免受降解与异常DNA修复。在出芽酵母中，端粒蛋白Cdc13通过结合富含G的单链悬突（G-rich overhangs）维持端粒完整性。本研究发现，Cdc13还可与单链/双链DNA（ss/dsDNA）连接处的邻近双链DNA发生弱相互作用，这一特性是抑制Exo1及Sgs1–Dna2介导的DNA末端切除的关键。
保守残基K504E的突变会破坏该连接处识别能力，进而削弱末端切除抑制、复制蛋白A（Replication Protein A）置换以及与Ku蛋白的协同作用。cdc13-K504E突变体细胞对Exo1过表达表现出超敏性，且在长期传代培养过程中会积累非色素化回复突变菌落。此类白色菌落生长速率更快，端粒近端基因表达谱发生改变，代谢通路上调，且4N DNA含量升高。
本研究揭示了Cdc13的双重DNA结合模式：通过阻断DNA末端切除与调控Ku蛋白定位来保护端粒，并发现了与端粒功能异常相关的应激诱导适应性状态。
整体实验设计：每组设置3个生物学重复；对野生型（Wild Type）红色菌落、K504E红色菌落及K504E白色菌落开展RNA测序（RNA-seq）。