Determining the role of TCAB1 in shaping telomerase function

确定 TCAB1 在塑造端粒酶功能中的作用

• 批准号: 9366667
• 负责人: STEVEN E ARTANDI
• 金额: $ 44.83万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-09-15 至 2022-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9366667
• 关键词: Adopted; Affect; Aging; Binding; Binding Proteins; Bioinformatics; Biological Assay; Catalysis; Catalytic Domain; Cell Aging; Cell Nucleus; Cells; Complement; Complex; Core Protein; Development; Disease; Elements; Embryo; Engineering; Enzymes; Evolution; Fibroblasts; Functional disorder; Genetic Polymorphism; Germ Cells; Germ-Line Mutation; Holoenzymes; Human; Impairment; Incidence; Investigation; Knockout Mice; Messenger RNA; Molecular Conformation; Morbidity - disease rate; Mus; Mutation; Pathway interactions; Patients; Pattern; Phenotype; Population; Process; Proteins; Pseudouridine; RNA; RNA Folding; RNA Splicing; RNA-Directed DNA Polymerase; Recruitment Activity; Reproducibility; Role; Series; Shapes; Structure; Susceptibility Gene; Telomerase; Telomerase RNA Component; Telomere Maintenance; Telomere Shortening; Untranslated RNA; crosslinking and immunoprecipitation sequencing; embryonic stem cell; genome-wide; human tissue; in vivo; mutant; protein complex; scaffold; single molecule; telomere; trafficking; transcriptome sequencing

Abstract Impaired maintenance of telomeres by telomerase causes cellular senescence and underlies many aging-related diseases in humans. Furthermore, telomere shortening is one of the most reproducible hallmarks of aging in human tissues, suggesting that telomere dysfunction contributes broadly to aging phenotypes in people. The human telomerase enzyme is comprised of a catalytic core – the telomerase reverse transcriptase (TERT) and the telomerase RNA component (TERC) – but also depends upon other holoenzyme proteins that bind the H/ACA element within TERC. During evolution, vertebrate telomerase hijacked this H/ACA sequence from other ancient non-coding RNAs, bringing the dyskerin core protein complex, which recognizes the H/ACA element, and TCAB1, which binds the CAB box element, into the telomerase holoenzyme. Telomerase function is critically dependent upon a series of steps, including assembly of the enzyme, trafficking within the nucleus, recruitment to telomeres and finally catalytic extension of telomeres. Each of these steps is poorly understood in humans and each step can be disrupted by germline mutations in patients with telomere diseases. The dyskerin complex is required for assembly and stability of telomerase. We identified TCAB1 as a protein that interacts with dyskerin and showed that TCAB1 binds the CAB box element common to TERC and to all small Cajal body RNAs (scaRNAs). We previously showed that TCAB1 is required for localization of telomerase in Cajal bodies, required for telomere maintenance and is important in recruitment to telomeres. We recently made the surprising finding that TCAB1 is also critical for telomerase catalytic activity and that TCAB1 is required for proper TERC conformation. Telomerase and other RNPs depend on an RNA molecule that needs to fold in a precise conformation, and this correct conformation represents one of countless potential structures. In this proposal, we pursue the hypothesis that TCAB1 is essential in shaping the telomerase RNA, and that this activity of TCAB1 is conserved in the splicing RNA pathway. We propose that the principal advantages conferred upon telomerase by adopting an H/ACA RNA fate is facilitating assembly and proper folding of TERC, and that numerous downstream steps depend upon this proper folding. We will pursue the following aims: (1) To understand how telomerase catalytic function depends upon TCAB1 (2) To determine how TCAB1 influences telomerase trafficking and recruitment (3) To determine how TCAB1 loss affects scaRNA function and RNA splicing.

摘要 端粒酶对端粒的维护受损，导致细胞衰老， 许多与衰老有关的疾病。此外，端粒缩短是最重要的因素之一。 人体组织中可重复的衰老标志，表明端粒功能障碍 在很大程度上导致了人类的衰老表型。人类端粒酶是 由催化核心-端粒酶逆转录酶（TERT）和端粒酶 RNA组分（TERC）-但也取决于其他全酶蛋白，结合 TERC中的H/ACA元件。在进化过程中，脊椎动物的端粒酶劫持了这个H/ACA 从其他古老的非编码RNA序列，使dyskerin核心蛋白复合物， 识别H/ACA元件的TCAB 1和结合CAB盒元件的TCAB 1， 端粒酶全酶端粒酶的功能主要依赖于一系列的步骤， 包括酶的组装，细胞核内的运输，端粒的募集， 最后是端粒的催化延伸。这些步骤中的每一个在人类中都知之甚少， 每个步骤都可能被端粒疾病患者的生殖细胞突变所破坏。的 dyskerin复合物是端粒酶组装和稳定所必需的。我们将TCAB 1鉴定为 与dyskerin相互作用并显示TCAB 1结合CAB盒元件的蛋白质 TERC和所有小Cajal体RNA（scaRNA）共有。我们之前已经证明， TCAB 1是Cajal小体中端粒酶定位所必需的，是端粒形成所必需的。 维持并在端粒的募集中起重要作用。我们最近做出了令人惊讶的 发现TCAB 1对于端粒酶催化活性也是至关重要的，并且TCAB 1是 正确的TERC构象。端粒酶和其他RNP依赖于一种RNA分子， 需要以精确的构象折叠，而这种正确的构象代表了 无数潜在的结构。在这个提议中，我们继续假设TCAB 1是 TCAB 1的这种活性在端粒酶RNA的形成中是必需的，并且TCAB 1的这种活性在端粒酶RNA的形成中是保守的。 剪接RNA途径。我们认为，端粒酶的主要优势是由 采用H/ACA RNA命运有利于TERC的组装和正确折叠， 许多下游步骤取决于这种适当的折叠。我们将继续努力 目的：（1）了解TCAB 1对端粒酶催化活性的影响;（2）研究TCAB 1对端粒酶活性的影响。 确定TCAB 1如何影响端粒酶的运输和募集（3）确定TCAB 1如何影响端粒酶的运输和募集 TCAB 1缺失影响scaRNA功能和RNA剪接。