A role for nuclear actin filaments at damaged telomeres

描述（由申请人提供）：肌动蛋白是真核生物中最充分表征的蛋白质之一，是许多胞质过程所必需的，包括细胞运动、细胞内转运和细胞结构完整性。最近的报道表明，肌动蛋白也参与了许多细胞类型的核过程，尽管这种肌动蛋白核库的作用在很大程度上是未知的。由于在完整的体细胞核中观察肌动蛋白的困难，以前对核肌动蛋白的研究受到限制。我最近开发了新的记者在活细胞核肌动蛋白的单体和丝状池。使用这些工具与布莱克本实验室合作，我们已经取得了令人兴奋的发现，肌动蛋白丝与无帽端粒共定位。为了研究端粒肌动蛋白的作用，我建议确定核肌动蛋白的活细胞动力学方面的上限和脱保护端粒。我也将调查是否协会是依赖于DNA损伤反应，以及负责招聘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，活性端粒酶的破坏或丧失已显示导致端粒缩短并触发细胞衰老（Yu et al.1990，哈雷et al.1990）。当将活性端粒酶引入具有低端粒酶活性的细胞培养物中时，端粒延长并且细胞可以变得永生化（Bodnar等1998，Counter等1998）。从这些观察中，传统的观点认为端粒“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长度本身，而是端粒缩短导致的端粒保护的丧失（“脱帽”）（Blackburn 2000）。拟议的研究确定核肌动蛋白作为一种新的因素存在于未加帽的端粒，并调查端粒肌动蛋白在信号转导的机制参与细胞周期退出，凋亡和基因组不稳定性的潜在作用。这项研究将提供深入了解的后果是负责促进细胞衰老的未加帽的端粒。

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.