Telomeres are specialized nucleoprotein complexes that protect the ends of eukaryotic chromosomes and distinguish them from broken DNA ends. Disruption of telomere protection may cause aging-associated pathologies and cancer. Here, we examined what makes telomere protection durable and resistant to perturbations using a budding yeast model organism. The protein Rap1 binds the telomeric repeats, negatively regulates telomere length, and protects telomeres by repressing homologous recombination and non-homologous end joining (NHEJ). A single-nucleotide mutation in the Kluyveromyces lactis telomerase RNA (TER1) template, ter1-16T, is incorporated into the telomeric repeats, disrupting the binding of Rap1 and causing dramatic telomere elongation. However, cell viability is not significantly affected, suggesting the existence of additional mechanism(s) for telomere protection. To examine this hypothesis, we explored the contribution of the recombination factor Rad52 and telomerase to telomere protection in the background of ter1-16T. To disrupt the function of telomerase, we exploited small mutations in a stem-loop domain of TER1 (Reg2), which result in short but stable telomeres. We generated K. lactis strains with combinations of three different mutations: ter1-16T, RAD52 deletion, and a two-nucleotide substitution in Reg2. Our results show that upon Rap1 depletion from telomeres, telomerase and the recombination machinery compensate for the loss of Rap1 protection and play redundant but critical roles in preventing NHEJ and maintaining telomere integrity and cell viability. These results demonstrate how redundant pathways make the essential role of telomeres—protecting our genome integrity and preventing cancer—more robust and resistant to assaults and perturbations.

端粒是专门的核蛋白复合物，可保护真核染色体的末端并将其与断裂的DNA末端区分开。端粒保护的破坏可能导致与衰老相关的病理和癌症。在这里，我们研究了使用发芽酵母模型生物使端粒保护持久且抗干扰的原因。Rap1蛋白结合端粒重复序列，负调节端粒长度，并通过抑制同源重组和非同源末端连接（NHEJ）保护端粒。乳酸克鲁维酵母中的单核苷酸突变端粒酶RNA（TER1）模板ter1-16T被整合到端粒重复序列中，破坏了Rap1的结合并导致端粒显着延长。但是，细胞活力并未受到显着影响，表明存在端粒保护的其他机制。为了验证这一假设，我们在ter1-16T背景下探索了重组因子Rad52和端粒酶对端粒保护的作用。为了破坏端粒酶的功能，我们利用了TER1（Reg2）茎环结构域中的小突变，从而导致了短而稳定的端粒。我们生成了具有三种不同突变组合的乳酸克鲁维酵母菌株：ter1-16T，RAD52缺失，以及Reg2中的两个核苷酸取代。我们的研究结果表明，从端粒中耗尽Rap1后，端粒酶和重组机制可弥补Rap1保护作用的丧失，并在预防NHEJ和维持端粒完整性和细胞活力方面起着多余但至关重要的作用。这些结果表明，多余的途径如何使端粒的重要作用（保护我们的基因组完整性和预防癌症）变得更健壮，并能抵抗攻击和摄动。