Chromosome instability resulting from double-strand breaks near telomeres

端粒附近双链断裂导致染色体不稳定

• 批准号: 7073078
• 负责人: John P. Murnane
• 金额: $ 26.45万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-07-06 至 2011-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7073078
• 关键词: chromosomes; human; proteins; siblings; telomere

DESCRIPTION (provided by applicant): Exposure of mammalian cells to ionizing radiation is known to induce chromosome instability (CIN), which can promote the rapid accumulation of genetic changes leading to cancer. We have demonstrated that DSBs occurring near the ends of chromosomes, called telomeres, can promote CIN by causing sister chromatid fusions that initiate breakage/fusion/bridge (B/F/B) cycles. B/F/B cycles occur when fused sister chromatids break during anaphase as their centromeres are pulled in opposite directions. Because breakage does not occur at the site effusion, one daughter cell acquires a chromosome with an inverted repeat at its end, while the other has a chromosome with a terminal deletion. This cycle is then repeated in subsequent cell cycles, resulting in further gene amplification and large terminal deletions. B/F/B cycles continue until the chromosome acquires a new telomere, which we have shown can occur by multiple mechanisms. One of the most common mechanisms is nonreciprocal translocation, which results in the transfer of instability to the chromosome donating the translocation due to the loss of its telomere. As a result, a single DSB near a telomere can result in the instability in multiple chromosomes. This proposal has two specific aims. In the first specific aim, we will address the hypothesis that sister chromatid fusions resulting from DBSs near telomeres result from a deficiency in nonhomologous end joining (NHEJ) near telomeres, as has been demonstrated in yeast. For these studies, we will create an assay system using isogenic cell lines to compare the efficiency of NHEJ at DSBs occurring at various distances from a telomere. We will also use this assay system to address differences in the repair proteins involved in NHEJ and sister chromatid fusion, which has been proposed to involve microhomology-mediated homologous recombinational repair. In the second specific aim, we will address the hypothesis that the p53 and Rb proteins involved in replicative senescence are important in preventing CIN due to DSB-induced telomere loss. In addition, we will establish a genetic screen using green fluorescent protein to monitor the duration of B/F/B cycles to identify additional proteins that can prevent B/F/B cycles. These studies are important in that they will provide new insights into mechanisms of chromosome instability in cancer and lead to new approaches for the development of therapies for preventing CIN in cancer cells.

描述（由申请人提供）：已知哺乳动物细胞暴露于电离辐射会诱发染色体不稳定（CIN），这会促进导致癌症的遗传变化的快速积累。我们已经证明，发生在染色体末端（称为端粒）附近的 DSB 可以通过引发姐妹染色单体融合（启动断裂/融合/桥接 (B/F/B) 循环）来促进 CIN。当融合的姐妹染色单体在后期因着丝粒被向相反方向拉动而断裂时，就会发生 B/F/B 循环。由于渗出位点不会发生断裂，因此一个子细胞获得了末端带有反向重复序列的染色体，而另一个子细胞则获得了末端缺失的染色体。然后在随后的细胞周期中重复该循环，导致进一步的基因扩增和大的末端缺失。 B/F/B 循环持续进行，直到染色体获得新的端粒，我们已经证明这可以通过多种机制发生。最常见的机制之一是非相互易位，由于端粒丢失，导致不稳定转移到提供易位的染色体上。因此，端粒附近的单个 DSB 可能会导致多条染色体不稳定。该提案有两个具体目标。在第一个具体目标中，我们将解决这样的假设：端粒附近的 DBS 产生的姐妹染色单体融合是由于端粒附近的非同源末端连接 (NHEJ) 缺陷造成的，正如在酵母中所证明的那样。对于这些研究，我们将使用同基因细胞系创建一个测定系统，以比较 NHEJ 对距端粒不同距离处发生的 DSB 的效率。我们还将使用该检测系统来解决 NHEJ 和姐妹染色单体融合所涉及的修复蛋白的差异，这已被提议涉及微同源介导的同源重组修复。在第二个具体目标中，我们将提出以下假设：参与复制衰老的 p53 和 Rb 蛋白对于预防 DSB 诱导的端粒丢失所致的 CIN 非常重要。此外，我们将使用绿色荧光蛋白建立遗传筛选来监测B/F/B循环的持续时间，以识别可以阻止B/F/B循环的其他蛋白质。这些研究很重要，因为它们将为癌症染色体不稳定机制提供新见解，并为开发预防癌细胞 CIN 的疗法提供新方法。