Deregulation of DNA Double-Strand Break Repair in Multiple Myeloma: Implications for Genome Stability
Deregulation of DNA Double-Strand Break Repair in Multiple Myeloma: Implications for Genome Stability
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DOI：
10.1371/journal.pone.0121581
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发表时间：
2015-03-19
期刊：
影响因子：
3.7
通讯作者：
Gutierrez, Norma C.
中科院分区：
文献类型：
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作者：
Herrero, Ana B.;San Miguel, Jess;Gutierrez, Norma C.
多发性骨髓瘤(MM)是一种以频繁的染色体异常为特征的血液系统恶性肿瘤。然而，这种基因组不稳定性的分子基础仍然未知。由于受损和过度活跃的双链断裂(DSB)修复通路都可以导致DNA重排，我们研究了MM细胞中DSB修复的功能。彗星实验显示，电离辐射(IR)诱导的DSB在MM和正常对照淋巴母细胞系中的修复动力学相似。然而，在分析的7个MM细胞系中，有4个表现出持续DSB的亚群，以伽马-H_2AX和RAD51为标志，引起G2/M DNA损伤检查点激活和对IR的超敏反应，特别是在检查点抑制剂存在的情况下。对MM细胞中参与DSB修复的蛋白质的分析表明，参与非同源末端连接(NHEJ)的DNA-PKcs、Artemis和XRCC4以及参与同源重组(HR)的RAD51上调。相应地，体内功能分析表明，MM细胞中NHEJ和HR的活性均高于对照组。有趣的是，参与高度突变、易位促进、替代NHEJ亚途径(Alt-NHEJ)的蛋白质水平在所有MM细胞系中也增加，Alt-NHEJ蛋白DNA连接酶IIIα在从MM患者分离的几个浆细胞样本中也过表达。在多发性骨髓瘤细胞中，Alt-NHEJ途径的过度激活是由于修复连接处更大的缺失和更高的序列微同源性，而该途径的化学抑制则减少了该通路。综上所述，我们的结果揭示了多发性骨髓瘤中一种解除调节的DSB修复，这可能是该病特有的基因组不稳定的基础，并可用于治疗。
Multiple myeloma (MM) is a hematological malignancy characterized by frequent chromosome abnormalities. However, the molecular basis for this genome instability remains unknown. Since both impaired and hyperactive double strand break (DSB) repair pathways can result in DNA rearrangements, we investigated the functionality of DSB repair in MM cells. Repair kinetics of ionizing-radiation (IR)-induced DSBs was similar in MM and normal control lymphoblastoid cell lines, as revealed by the comet assay. However, four out of seven MM cell lines analyzed exhibited a subset of persistent DSBs, marked by gamma-H2AX and Rad51 foci that elicited a prolonged G2/M DNA damage checkpoint activation and hypersensitivity to IR, especially in the presence of checkpoint inhibitors. An analysis of the proteins involved in DSB repair in MM cells revealed upregulation of DNA-PKcs, Artemis and XRCC4, that participate in non-homologous end joining (NHEJ), and Rad51, involved in homologous recombination (HR). Accordingly, activity of both NHEJ and HR were elevated in MM cells compared to controls, as determined by in vivo functional assays. Interestingly, levels of proteins involved in a highly mutagenic, translocation-promoting, alternative NHEJ subpathway (Alt-NHEJ) were also increased in all MM cell lines, with the Alt-NHEJ protein DNA ligase III alpha, also overexpressed in several plasma cell samples isolated from MM patients. Overactivation of the Alt-NHEJ pathway was revealed in MM cells by larger deletions and higher sequence microhomology at repair junctions, which were reduced by chemical inhibition of the pathway. Taken together, our results uncover a deregulated DSB repair in MM that might underlie the characteristic genome instability of the disease, and could be therapeutically exploited.