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. 3 biological replicates per condition; RNA-seq of WT-red, K504E-red, K504E-white colonies.