Activation of retrotransposable elements in cellular senescence and aging

 描述(申请人提供)：逆转座子元件(RTE)约占人类基因组的45%。RTE是可移动的DNA元件，可以使用复制和粘贴机制插入到新的基因组位置。这个过程被称为逆转座，可以在多个水平上造成有害的，导致突变和基因组结构不稳定，触发表观遗传变化，并扰乱正常的基因调控模式。人类中的许多单基因突变已被证明是由生殖系逆转录转座引起的。生物体已经进化出多种转录和转录后沉默机制来保护他们的基因组免受RTE的影响。直到最近，RTE还被认为在胞体中是沉默的，然而，新的证据表明，大脑和癌细胞中都有活动。事实上，初步迹象表明，体细胞逆转位的频率比之前预期的要高得多。我们已经报道，逆转座子在衰老和细胞衰老过程中被激活，并假设它可能代表一种迄今未被认识的分子衰老过程。我们研究的长期目标是确定逆转座对衰老过程中基因组完整性的影响。作为第一步，我在这个提案中的目标是，使用高通量DNA测序方法，研究细胞衰老过程中RTE的动员。在目标1中，我将进行全基因组分析，以确定新的插入发生在哪里，并评估这些事件是否有可能通过扰乱基因组中对细胞功能重要的区域而产生有害影响。在目标2中，我将研究个别细胞中独特的逆转位的发生情况。我们有证据表明，在单个有丝分裂后细胞停止分裂后，可能会出现许多新的插入。为了全面描述这些事件的光谱和频率，我将使用单细胞全基因组DNA测序。我的研究将采用创新的最先进的高通量测序策略，我将开发新的生物信息学工具来整合和验证来自多种算法的输出。所获得的信息将解决这个问题：反转座会在多大程度上损害我们体内的体细胞基因组，这是一种可信的衰老机制吗？所获得的经验和开发的工具将为未来旨在检查这些直接在老化组织中处理的研究提供高度信息。我的努力也将告诉我们，是否应该调查使用抑制逆转位的干预措施的原则验证研究，作为年龄相关疾病的潜在疗法。

 DESCRIPTION (provided by applicant): Retrotransposable elements (RTEs) comprise approximately 45% of the human genome. RTEs are mobile DNA elements that can insert into new genomic positions using a copy and paste mechanism. This process, termed retrotransposition, can be deleterious at multiple levels by causing mutagenesis and genome structural instability, triggering epigenetic changes, and disrupting normal patterns of gene regulation. Numerous single- gene mutations in humans have been documented to result from germline retrotransposition. Organisms have evolved multiple transcriptional and post-transcriptional silencing mechanisms to protect their genomes against RTEs. Until recently RTEs were thought to be silent in the soma, however, new evidence points to activity in the brain and in cancer cells. Indeed, initial indications are that somatic retrotransposition is much more frequent than previously anticipated. We have reported that retrotransposition is activated during aging and cellular senescence, and hypothesized that it may represent a hitherto unappreciated molecular aging process. The long-term goal of our research is to determine the impact of retrotransposition on genome integrity during aging. As a first step, my objective in thi proposal is to study, using high throughput DNA sequencing methods, the mobilization of RTEs during cellular senescence. In Aim 1 I will perform genome-wide profiling to determine where new insertions occur and evaluate whether these events have the potential to exert deleterious effects by disrupting regions of the genome important for cell function. In Aim 2 I will examine the occurrence of unique retrotranspositions in individual cells. We have evidence that many new insertions likely occur in individual post-mitotic cells after they have ceased dividing. To comprehensively profile the spectrum and frequency of these events I will use single-cell whole genome DNA sequencing. My research will employ innovative state-of-the-art high-throughput sequencing strategies, and I will develop new bioinformatic tools to integrate and validate the output from multiple algorithms. The information obtained will address the question: To what extent is retrotransposition damaging to the genomes of somatic cells in our bodies, and is this a plausible mechanism of aging? The experience gained and tools developed will be highly informative for future studies aimed at examining these processed directly in aging tissues. My efforts will also inform whether proof- of-principle studies using interventions that inhibit retrotransposition should be investigated as potential therapeutics for age-associated diseases.