Identification of endogenous TERRA-ribonucleoprotein components and characterization of their function in telomere biology

端粒形成真核生物染色体的末端。一组端粒特异性组分作为基因组完整性的监护者和肿瘤发生的抑制剂起着重要作用。最近，一种新的和意想不到的端粒成分已被描述：端粒重复包含RNA（TERRA）。一些观察结果指出，TERRA在维持端粒完整性方面发挥着至关重要的作用。然而，关于这种长的非编码RNA的调控的精确知识仍然很少。为了解决这一问题，本提案的目的是在TERRA结合蛋白的特性方面，其功能的TERRA和端粒代谢。因此，内源性TERRA及其相关因子，也称为TERRA-核糖核蛋白复合物（TERRA-RNP），将从人细胞系中分离。接下来，TERRA结合蛋白将通过质谱法鉴定。通过遗传操作的确定的因素，它们的作用TERRA代谢以及端粒的维护将进行评估。 这将阐明TERRA如何影响染色体稳定性，并将进一步确定与端粒完整性和癌发生抑制有关的新因素。这不仅可以揭示非编码RNA如何控制端粒/基因组完整性的新机制，而且还可以作为其他非编码转录物的作用和调节模式的范例。此外，表征的因素，调节TERRA，并反过来控制端粒的维护可能会发现新的和有前途的癌症治疗的目标。最后，表征TERRA-RNP将扩展我们对与端粒功能障碍相关的人类疾病的分子机制的认识。

Telomeres form the ends of eukaryotic chromosomes. A set of telomere-specific components plays fundamental roles as guardians of genome integrity and suppressors of tumorigenesis. Recently, a novel and unexpected telomere component has been described: the telomeric repeat containing RNA (TERRA). Several observations point towards an essential role of TERRA in the maintenance of telomere integrity. Nevertheless, precise knowledge about the regulation of this long non-coding RNA is still sparse. To address this issue, this proposal aims at the characterization of TERRA-binding proteins with regard to their function in TERRA and telomere metabolism. Hereby, endogenous TERRA and its associated factors, also called TERRA-ribonucleoprotein complexes (TERRA-RNPs), will be isolated from a human cell line. Next, TERRA-binding proteins will be identified by mass spectrometry. By genetic manipulation of the identified factors, their role in TERRA-metabolism as well as telomere maintenance will be assessed. This will elucidate how TERRA impinges on chromosome stability and will further identify new factors implicated in telomere integrity and the suppression of carcinogenesis. Not only may this uncover a novel mechanism of how a non-coding RNA can control telomere/genome integrity, but also serve as a paradigm for the mode of action and regulation of other non-coding transcripts. Furthermore, the characterization of factors that modulate TERRA and in turn control telomere maintenance may identify novel and promising targets for cancer therapy. Finally, characterizing TERRA-RNPs will expand our knowledge on the molecular mechanisms causative for human diseases associated with telomere dysfunction.