Swiss Institute for Experimental Cancer Research (ISREC), School of Life Sciences, Ecole Polytechnique Fédérale de Lausanne (EPFL), Lausanne, Switzerland.
Department of Biology and National Centre for Biomolecular Research, Masaryk University, Brno, Czech Republic.
Nature. 2020 Nov;587(7833):303-308. doi: 10.1038/s41586-020-2815-6. Epub 2020 Oct 14.
Telomeres-repeated, noncoding nucleotide motifs and associated proteins that are found at the ends of eukaryotic chromosomes-mediate genome stability and determine cellular lifespan. Telomeric-repeat-containing RNA (TERRA) is a class of long noncoding RNAs (lncRNAs) that are transcribed from chromosome ends; these RNAs in turn regulate telomeric chromatin structure and telomere maintenance through the telomere-extending enzyme telomerase and homology-directed DNA repair. The mechanisms by which TERRA is recruited to chromosome ends remain poorly defined. Here we develop a reporter system with which to dissect the underlying mechanisms, and show that the UUAGGG repeats of TERRA are both necessary and sufficient to target TERRA to chromosome ends. TERRA preferentially associates with short telomeres through the formation of telomeric DNA-RNA hybrid (R-loop) structures that can form in trans. Telomere association and R-loop formation trigger telomere fragility and are promoted by the recombinase RAD51 and its interacting partner BRCA2, but counteracted by the RNA-surveillance factors RNaseH1 and TRF1. RAD51 physically interacts with TERRA and catalyses R-loop formation with TERRA in vitro, suggesting a direct involvement of this DNA recombinase in the recruitment of TERRA by strand invasion. Together, our findings reveal a RAD51-dependent pathway that governs TERRA-mediated R-loop formation after transcription, providing a mechanism for the recruitment of lncRNAs to new loci in trans.
端粒是真核染色体末端的重复、非编码核苷酸序列和相关蛋白，它们介导基因组稳定性并决定细胞寿命。端粒重复 RNA（TERRA）是一类从染色体末端转录而来的长链非编码 RNA（lncRNA）；这些 RNA 通过端粒延伸酶端粒酶和同源定向 DNA 修复来调节端粒染色质结构和端粒维持。TERRA 被招募到染色体末端的机制仍未完全定义。在这里，我们开发了一个报告系统来剖析潜在的机制，并表明 TERRA 的 UUAGGG 重复序列既是必需的，也是将 TERRA 靶向染色体末端的充分条件。TERRA 通过形成可以反式形成的端粒 DNA-RNA 杂交（R 环）结构，优先与短端粒结合。端粒结合和 R 环形成引发端粒脆弱性，并受重组酶 RAD51 及其相互作用蛋白 BRCA2 促进，但被 RNA 监测因子 RNaseH1 和 TRF1 抵消。RAD51 与 TERRA 物理相互作用，并在体外与 TERRA 催化 R 环形成，表明这种 DNA 重组酶直接参与了通过链入侵招募 TERRA。总之，我们的发现揭示了一种依赖 RAD51 的途径，该途径控制转录后 TERRA 介导的 R 环形成，为 lncRNA 向新的反式位点募集提供了一种机制。
