Multiple Pathways Suppress Telomere Addition to DNA Breaks in the Drosophila Germline
Multiple Pathways Suppress Telomere Addition to DNA Breaks in the Drosophila Germline
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DOI：
10.1534/genetics.112.138818
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发表时间：
2012-06-01
期刊：
影响因子：
3.3
通讯作者：
Rong, Yikang S.
中科院分区：
文献类型：
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作者：
Beaucher, Michelle;Zheng, Xiao-Feng;Rong, Yikang S.
端粒保护染色体末端不被修复为双链断裂（DSB）。正如DSB修复在端粒处被抑制一样，从头添加端粒在DSB位点处被抑制。为了确定导致这种抑制的因素，我们开发了一种检测方法来监测果蝇中从头端粒的形成，果蝇是一种端粒可以在染色体末端以基本上任何序列建立的生物体。I-SceI核酸内切酶的胚系表达导致在其切割位点以高效率精确地形成端粒。使用该测定，我们定量了在不同遗传背景中具有已知或可能的DNA损伤修复缺陷的端粒形成的频率。我们发现，DSB修复因子（Rad 51或DNA连接酶IV）或DSB传感因子（ATRIP或MDC 1）的破坏导致更有效的端粒形成。有趣的是，通常调节端粒保护的因子（ATM或NBS）的部分破坏也导致端粒形成的频率更高，这表明这些蛋白质在端粒维持与建立中具有相反的作用。在ku 70突变体背景下，端粒的建立之前，DSB末端的过度降解，这是稳定的端粒形成。最引人注目的是，ATRIP的去除导致端粒反转录转座子附着到断裂末端的急剧增加。我们的研究确定了几个途径，抑制端粒添加在DSB，为未来的机制研究铺平了道路。
Telomeres protect chromosome ends from being repaired as double-strand breaks (DSBs). Just as DSB repair is suppressed at telomeres, de novo telomere addition is suppressed at the site of DSBs. To identify factors responsible for this suppression, we developed an assay to monitor de novo telomere formation in Drosophila, an organism in which telomeres can be established on chromosome ends with essentially any sequence. Germline expression of the I-SceI endonuclease resulted in precise telomere formation at its cut site with high efficiency. Using this assay, we quantified the frequency of telomere formation in different genetic backgrounds with known or possible defects in DNA damage repair. We showed that disruption of DSB repair factors (Rad51 or DNA ligase IV) or DSB sensing factors (ATRIP or MDC1) resulted in more efficient telomere formation. Interestingly, partial disruption of factors that normally regulate telomere protection (ATM or NBS) also led to higher frequencies of telomere formation, suggesting that these proteins have opposing roles in telomere maintenance vs. establishment. In the ku70 mutant background, telomere establishment was preceded by excessive degradation of DSB ends, which were stabilized upon telomere formation. Most strikingly, the removal of ATRIP caused a dramatic increase in telomeric retrotransposon attachment to broken ends. Our study identifies several pathways that suppress telomere addition at DSBs, paving the way for future mechanistic studies.