Department of Genetics, The Silberman Institute of Life Sciences, The Hebrew University of Jerusalem, Safra Campus, Givat Ram, 91904, Jerusalem, Israel.
Curr Genet. 2021 Feb;67(1):153-163. doi: 10.1007/s00294-020-01125-4. Epub 2020 Nov 6.
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.
端粒是一种特殊的核蛋白复合物，可保护真核生物染色体的末端，并将其与断裂的 DNA 末端区分开来。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，我们使用芽殖酵母模型生物研究了是什么使端粒保护持久且能抵抗干扰。Rap1 蛋白结合端粒重复序列，负调控端粒长度，并通过抑制同源重组和非同源末端连接（NHEJ）来保护端粒。在 Kluyveromyces lactis 端粒酶 RNA（TER1）模板中，ter1-16T 的一个单核苷酸突变被整合到端粒重复序列中，破坏了 Rap1 的结合，导致端粒显著伸长。然而，细胞活力没有受到显著影响，这表明存在额外的端粒保护机制。为了检验这一假设，我们在 ter1-16T 的背景下研究了重组因子 Rad52 和端粒酶对端粒保护的贡献。为了破坏端粒酶的功能，我们利用 TER1 （Reg2）茎环结构域中的小突变，产生短但稳定的端粒。我们生成了 K. lactis 菌株，其组合了三种不同的突变：ter1-16T、RAD52 缺失和 Reg2 中的两个核苷酸取代。我们的结果表明，在 Rap1 从端粒中耗尽后，端粒酶和重组机制补偿了 Rap1 保护的缺失，并在防止 NHEJ 和维持端粒完整性和细胞活力方面发挥了冗余但关键的作用。这些结果表明冗余途径如何使端粒保护这一至关重要的作用——保护我们的基因组完整性和预防癌症——更加稳健和耐受攻击和干扰。
