Determining the role of TCAB1 in shaping telomerase function

摘要 端粒酶的端粒维护受损会导致细胞衰老和基础 人类中的许多与衰老相关的疾病。此外，端粒缩短是最常见的 人类组织中可复制的衰老特征，表明端粒功能障碍 对人的衰老表型有广泛的影响。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)-但也依赖于其他结合 TERC中的H/ACA元素。在进化过程中，脊椎动物端粒酶劫持了这种H/ACA 来自其他古代非编码RNA的序列，带来了dyskerin核心蛋白复合体， 它识别H/ACA元素，而TCAB1将CAB盒元素绑定到 端粒酶全酶。端粒酶的功能关键依赖于一系列步骤， 包括酶的组装，核内的运输，端粒的募集和 最后是端粒的催化延伸。人类对这些步骤中的每一个都知之甚少，而且 每一步都可能被端粒疾病患者的生殖系突变所破坏。这个 Dyskerin复合体是端粒酶组装和稳定所必需的。我们确定TCAB1是一种 与dyskerin相互作用的蛋白质，并表明TCAB1与CAB盒元件结合 TERC和所有小Cajal小体RNA(ScaRNAs)共同的。我们之前已经证明了 端粒酶在Cajal小体中的定位需要TCAB1，端粒也需要TCAB1 维持并在端粒招募中起重要作用。我们最近做了一个令人惊讶的事情 发现TCAB1对端粒酶催化活性也是至关重要的，并且TCAB1是端粒酶活性所必需的 正确的TERC构象。端粒酶和其他RNPs依赖于一种RNA分子 需要折叠成精确的构象，而这个正确的构象代表了 无数潜在的建筑。在这个提议中，我们继续假设TCAB1是 TCAB1的这种活性在形成端粒酶RNA中是必不可少的，并且TCAB1的这种活性在 剪接RNA途径。我们认为端粒酶的主要优势是 采用H/ACA RNA命运有利于TERC的组装和适当折叠， 许多下游步骤依赖于这种适当的折叠。我们将致力于以下工作 目的：(1)了解端粒酶催化功能如何依赖于TCAB1(2) 确定TCAB1如何影响端粒酶的贩运和招募(3)以确定如何 TCAB1缺失影响Scarna功能和RNA剪接。

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.