Regulation of retrotransposable element activity in Drosophila.

项目概要（项目2） 本项目的重点是利用果蝇模型系统研究反转录转座因子在果蝇中的作用。 元素（RTEs）在衰老过程中发生的细胞功能障碍的进展，并发展间 抑制RTE活动和延长健康寿命的静脉。维持压制性的异性恋- 染色质随着年龄的增长而下降，导致跨物种的RTEs表达和动员增加， 像果蝇和老鼠一样多种多样。我们推测，RTE表达和动员的增加， 导致体细胞基因组损伤的积累，导致细胞和生物体的损失， 体内平衡，从而促进衰老。我们将使用强大的分子和遗传工具和短暂的生命 果蝇的寿命来确定RTE对衰老的影响，特别是，我们将在整个 生物体的寿命，这在人类身上是不可能的，而且会是令人望而却步的时间- 在任何大规模的哺乳动物模型系统中消耗和昂贵。确认一个机械 RTE活动与衰老之间的关系将为制定干预措施提供基础， 减少RTE活动和衰老过程中的基因组损伤，并应延长健康寿命。 本提案的目的是检验以下假设：（i）染色质调节的抑制作用的丧失， 随着年龄的增长，RTE活性与DNA损伤和基因组不稳定性增加有关;（ii）RTE活性降低， 随着年龄的增长，沉默调节蛋白导致RTE活性升高，这可以通过增加沉默调节蛋白活性来逆转;以及（iii） 抑制体细胞中RTE活性的干预可以延长健康寿命。 为了更好地了解RTE活性、基因组损伤和寿命之间的关系， 开发新的干预措施，可以延长健康的寿命，在目标1，我们将开发高通量DNA 测序方法（与项目1，核心B合作），部署可用的和新的RTE报告基因 （与Core B合作）测量衰老果蝇中RTE的动员，并进行了一项前瞻性研究。 基因筛选，以确定参与抑制RTEs的新基因和途径。我们会研究 （i）年龄，（ii）饮食限制（DR）和（iii）稳定异染色质的遗传干预的影响， 增加Su（var）3-9、HP 1a、Dicer-2、Piwi表达或降低阿达尔表达对RTE活性和动员的影响。 在目的2中，为了确定Sir 2和Sirt 6在衰老过程中抑制RTE活性中的作用，我们将联合收割机 在目标1中开发和利用的工具，通过基因操作监测RTE活动， 降低Sir 2和Sirt 6活性的表达（与项目3和核心B合作）。在目标3中，我们 (i)确定目标1和目标2中制定的遗传干预措施对果蝇寿命的影响 （与项目1和3以及核心B合作），以及（ii）测试核苷逆转录酶的作用 抑制剂（nRTIs）和RNAi敲除特定反转录转座子对RTE动员和寿命的影响 （与项目1和3以及核心项目B合作）。

PROJECT SUMMARY (PROJECT 2) The focus of this project is to exploit the Drosophila model system to study the role of retrotransposable elements (RTEs) in the progression of cellular dysfunction that occurs during aging, and to develop inter- ventions that suppress RTE activity and extend healthy life span. The maintenance of repressive hetero- chromatin declines with age, resulting in increased expression and mobilization of RTEs across species as diverse as fruitflies and mice. We hypothesize that the increase in the expression and mobilization of RTEs leads to an accumulation of damage to the genome of somatic cells, causing loss of cellular and organismal homeostasis, and thus promoting aging. We will use the powerful molecular and genetic tools and the short life span of Drosophila to determine the effects of RTEs on aging, and in particular, we will do this over the entire life span of the organism, something that would not be possible in humans, and would be prohibitively time- consuming and expensive in any mammalian model system on a large scale. Confirmation of a mechanistic relationship between RTE activity and aging will provide the groundwork for developing interventions that diminish RTE activity and genomic damage during aging, and should extend healthy life span. The aims of this proposal are to test the hypotheses that: (i) loss of chromatin-regulated suppression of RTE activity with age is associated with increased DNA damage and genome instability; (ii) decreases in sirtuins with age lead to elevated RTE activity that can be reversed by increasing sirtuin activity; and (iii) interventions that suppress RTE activity in somatic cells can extend healthy life span. To better understand the relationship between RTE activity, genomic damage and longevity, as well as to develop new interventions that can extend healthy life span, in Aim 1 we will develop high throughput DNA sequencing methods (collaboration with Project 1, Core B), deploy available and new RTE reporters (collaboration with Core B) to measure mobilization of RTEs in aging Drosophila, and carry out a forward genetic screens to identify new genes and pathways involved in the suppression of RTEs. We will examine the effects of (i) age, (ii) dietary restriction (DR), and (iii) genetic interventions that stabilize heterochromatin, such as increasing Su(var)3-9, HP1a, Dicer-2, Piwi or decreasing Adar expression, on RTE activity and mobilization. In Aim 2, to determine the role of Sir2 and Sirt6 in the repression of RTE activity during aging, we will combine the tools developed and utilized in Aim 1 to monitor RTE activity with genetic manipulations that increase or decrease the expression of Sir2 and of Sirt6 activity (collaboration with Project 3 and Core B). In Aim 3, we will (i) determine the effects of the genetic interventions developed in Aims 1 and 2 on the life span of flies (collaboration with Projects 1 and 3 and Core B), and (ii) test the effects of nucleoside reverse transcriptase inhibitors (nRTIs) and RNAi knockdowns of specific retrotransposons on RTE mobilization and lifespan (collaboration with Projects 1 and 3 and Core B).