Department of Cancer Genetics and Epigenetics, City of Hope, Duarte, California 91010, USA.
Irell and Manella Graduate School of Biological Sciences, City of Hope, Duarte, California 91010, USA.
Genes Dev. 2018 Apr 1;32(7-8):524-536. doi: 10.1101/gad.311084.117. Epub 2018 Apr 10.
Chromosomal deletion rearrangements mediated by repetitive elements often involve repeats separated by several kilobases and sequences that are divergent. While such rearrangements are likely induced by DNA double-strand breaks (DSBs), it has been unclear how the proximity of DSBs relative to repeat sequences affects the frequency of such events. We generated a reporter assay in mouse cells for a deletion rearrangement involving repeats separated by 0.4 Mb. We induced this repeat-mediated deletion (RMD) rearrangement with two DSBs: the 5' DSB that is just downstream from the first repeat and the 3' DSB that is varying distances upstream of the second repeat. Strikingly, we found that increasing the 3' DSB/repeat distance from 3.3 kb to 28.4 kb causes only a modest decrease in rearrangement frequency. We also found that RMDs are suppressed by KU70 and RAD51 and promoted by RAD52, CtIP, and BRCA1. In addition, we found that 1%-3% sequence divergence substantially suppresses these rearrangements in a manner dependent on the mismatch repair factor MSH2, which is dominant over the suppressive role of KU70. We suggest that a DSB far from a repeat can stimulate repeat-mediated rearrangements, but multiple pathways suppress these events.
由重复元件介导的染色体缺失重排通常涉及相隔几千个碱基对的重复序列和差异序列。虽然这种重排可能是由 DNA 双链断裂 (DSB) 诱导的，但 DSB 相对于重复序列的接近程度如何影响这种事件的频率尚不清楚。我们在小鼠细胞中生成了一种涉及相隔 0.4Mb 的重复序列的缺失重排报告基因检测。我们通过两个 DSB 诱导这种重复介导的缺失 (RMD) 重排：第一个重复下游的 5' DSB 和第二个重复上游不同距离的 3' DSB。引人注目的是，我们发现将 3' DSB/重复距离从 3.3kb 增加到 28.4kb 仅导致重排频率适度下降。我们还发现，KU70 和 RAD51 抑制 RMD，RAD52、CtIP 和 BRCA1 促进 RMD。此外，我们发现 1%-3%的序列差异以依赖于错配修复因子 MSH2 的方式显著抑制这些重排，而 MSH2 的抑制作用超过了 KU70 的抑制作用。我们认为，远离重复序列的 DSB 可以刺激重复介导的重排，但有多种途径抑制这些事件。
