A role for nuclear actin filaments at damaged telomeres

描述(申请人提供)：肌动蛋白是真核生物中最具特性的蛋白质之一，在许多胞浆过程中都是必需的，包括细胞运动、细胞内运输和细胞结构的完整性。最近的报道表明，肌动蛋白还参与了多种细胞类型的一系列核过程，尽管肌动蛋白核池的作用在很大程度上是未知的。以前关于核肌动蛋白的工作一直受到限制，因为在完整的体细胞核中显示肌动蛋白的难度很大。我最近开发了关于活细胞中核肌动蛋白单体池和丝状池的新颖记者。与布莱克本实验室合作使用这些工具，我们已经取得了令人兴奋的发现，肌动蛋白细丝与未封顶的端粒共存。为了研究端粒肌动蛋白的作用，我建议确定核肌动蛋白在有保护和去保护的端粒上的活细胞动力学。我还将调查这种关联是否依赖于DNA损伤反应，以及负责招募和调节端粒肌动蛋白的因素。 公共卫生相关性：几十年来对端粒生物学的实验研究已经提供了端粒功能和衰老之间的明确联系。在体内和体外，端粒酶活性的破坏或丢失都被证明会导致端粒缩短并触发细胞衰老(Yu等人。1990年，哈雷等人。1990年)。当活性端粒酶被引入低端粒酶活性的细胞培养时，端粒延长，细胞可以永生(Bodnar等人)。1998年，Counter等人。1998年)。从这些观察中，出现了一种传统的观点，即端粒“临界长度”的丧失会促进衰老。然而，最近的证据表明，限制细胞增殖的不是端粒本身的长度，而是端粒缩短导致的端粒保护的丧失(“去封顶”)(Blackburn 2000)。这项拟议的研究确定了核肌动蛋白是一种存在于无帽端粒的新因子，并研究了端粒肌动蛋白在细胞周期退出、细胞凋亡和基因组不稳定所涉及的机制中的潜在作用。这项研究将提供对无上限端粒的后果的洞察，端粒负责促进细胞衰老。

DESCRIPTION (provided by applicant): Actin is among the most well characterized proteins in eukaryotic organisms and is required for many cytosolic processes, including cell motility, intracellular transport and the structural integrity of the cell. Recent reports have suggested that actin also participates in a range of nuclear processes in numerous cell types, though the role of this nuclear pool of actin is largely unknown. Previous work on nuclear actin has been limited due to the difficulty associated with visualizing actin in intact somatic nuclei. I have recently developed novel reporters of both monomeric and filamentous pools of nuclear actin in live cells. Using these tools in collaboration with the Blackburn lab, we have made the exciting discovery that actin filaments colocalize with uncapped telomeres. To investigate the role of telomeric actin, I propose to determine the live cell dynamics of nuclear actin with respect to both capped and deprotected telomeres. I will also investigate whether the association is dependent on the DNA damage response, as well as factors responsible for recruitment and regulation of telomeric actin. PUBLIC HEALTH RELEVANCE: Decades of experimental research on telomere biology have provided a clear link between telomere function and aging. Disruption or loss of active telomerase has been shown to cause telomeres to shorten and trigger cellular senescence, both in vivo and in vitro (Yu et al. 1990, Harley et al. 1990). When active telomerase is introduced into cell cultures with low telomerase activity, telomeres lengthen and the cells can become immortalized (Bodnar et al. 1998, Counter et al. 1998). From these observations, the traditional view that emerged was that the loss of telomere "critical length" promotes senescence. However, more recent evidence suggests that rather than telomere length per se, it is the loss of protection ("uncapping") of the telomere that results from telomere shortening that limits cell proliferation (Blackburn 2000). The proposed research identifies nuclear actin as a novel factor present at uncapped telomeres, and investigates potential roles for telomeric actin in signaling to the machineries involved in cell cycle exit, apoptosis and genomic instability. This research will provide insight into the consequences of uncapped telomeres that are responsible for promoting cellular senescence.