Compromised repair of clustered DNA damage in the human acute lymphoblastic leukemia MSH2-deficient NALM-6 cells
Compromised repair of clustered DNA damage in the human acute lymphoblastic leukemia MSH2-deficient NALM-6 cells
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DOI：
10.1016/j.mrgentox.2008.09.014
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发表时间：
2009-03-31
影响因子：
1.9
通讯作者：
Georgakilas, Alexandros G.
中科院分区：
文献类型：
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作者：
Holt, Stewart M.;Scemama, Jean-Luc;Georgakilas, Alexandros G.
电离辐射（IR）可引起两类复杂的DNA损伤，即双链断裂（DSB）和非DSB双链氧化成簇DNA损伤（OCDLs）。OCDLs可能由5- 10 bp的单链断裂、氧化嘌呤/嘧啶和脱碱基位点组成。这些显著的生物学病变被假设为挑战修复机制并具有高致突变潜力。MSH 2是一种经典的DNA错配修复蛋白，也参与了与DSB和基底损伤处理相关的其他修复途径。MSH 2突变已在急性淋巴细胞白血病（ALL）患者以及其他类型的癌症中发现。我们的研究模型涉及两种前体B（pre-B）ALL人类细胞系，NALM-6细胞，纯合的MSH 2无效细胞和野生型697细胞。采用改良的中性和碱性单细胞凝胶电泳（SCGE）技术，以大肠杆菌修复酶为损伤探针，研究了NALM-6和697细胞在放射治疗相关剂量为5戈伊的γ射线照射下对单链断裂（SSB）、双链断裂（DSB）和OCDLs的处理能力。利用逆转录酶PCR和蛋白质印迹法，我们证实了在转录和翻译水平上在NALM-6细胞中完全缺乏MSH 2的表达。NALM-6和697细胞暴露于γ射线后，SSB、DSB和OCDLs的诱导水平无差异。然而，697细胞更有效地修复每个损伤，在照射后1-3小时观察到显著差异。最后，我们的结果表明，与NALM-6细胞相比，辐射后6-24小时凋亡697细胞的数量显著更高。我们的研究表明，MSH 2可能参与了电离辐射诱导的具有生物学意义的成簇DNA损伤的处理以及细胞凋亡的执行。(C)2008 Elsevier B. V.保留所有权利。
Ionizing radiation (IR) induces two classes of complex DNA damage, double-strand breaks (DSBs) and non-DSB bi-stranded oxidative clustered DNA lesions (OCDLs). OCDLs may consist of single strand breaks (SSBs), oxidized purines/pyrimidines and abasic sites within 5-10bp. These significant biological lesions are hypothesized to challenge the repair machinery and carry a high mutagenic potential. MSH2, a classical DNA mismatch repair protein, has been also implicated in other repair pathways associated with DSB and base lesion processing. MSH2 mutations have been identified in acute lymphoblastic leukemia (ALL) patients as well as in other types of cancers. Our research model involves two precursors B (pre-B) ALL human cell lines, NALM-6 cells, homozygous null for MSH2, and wild type 697 cells. Using a modified version of neutral and alkaline single cell gel electrophoresis (SCGE) with Escherichia coli repair enzymes as damage probes, the processing capacity of single strand breaks (SSBs), DSBs and OCDLs was assessed in NALM-6 and 697 cells exposed to a radiotherapy relevant gamma-ray dose of 5 Gy. Using reverse transcriptase PCR and Western blotting we verified the complete lack of expression of MSH2 in the NALM-6 cells at the transcriptional and translational level. No differences were measured between NALM-6 and 697 cells in the induction levels of SSBs, DSBs and OCDLs after exposure to gamma-rays. However, 697 cells repaired each lesion more efficiently with significant differences observed after 1-3 h post-irradiation. Lastly, our results indicate a significantly higher population of apoptotic 697 cells compared to NALM-6 cells 6-24 h post-irradiation. Our studies suggest that MSH2 is probably involved in the processing of the biologically significant clustered DNA damages as well as the execution of apoptosis induced by ionizing radiation. (C) 2008 Elsevier B.V. All rights reserved.