Role of APE1 in differentiated neuroblastoma SH-SY5Y cells in response to oxidative stress: use of APE1 small molecule inhibitors to delineate APE1 functions.
Role of APE1 in differentiated neuroblastoma SH-SY5Y cells in response to oxidative stress: use of APE1 small molecule inhibitors to delineate APE1 functions.
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DOI：
10.1016/j.dnarep.2009.08.003
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发表时间：
2009-11-02
期刊：
影响因子：
3.8
通讯作者：
Kelley MR
中科院分区：
文献类型：
--
作者：
Jiang Y;Guo C;Fishel ML;Wang ZY;Vasko MR;Kelley MR
氧化性DNA损伤与许多中枢神经系统的病理有关。碱基切除修复(BER)途径是应答DNA氧化损伤最重要的细胞保护机制之一。人无嘌呤(无嘧啶)核酸内切酶/氧化还原效应因子(APE1/Ref-1或APE1)是BER途径中的一个重要酶，在人类有丝分裂和有丝分裂后的细胞中均有表达。在神经元中，APE1表达的减少增加了化疗引起的细胞毒性，而APE1的过度表达则保护细胞免受细胞毒性的影响。然而，鉴于APE1的多重功能，总APE1的敲除并不能完全反映它是氧化还原或DNA修复活性，还是与其他蛋白质的相互作用。因此，使用选择性的小分子可以独立地阻断每一种功能，对于确定有丝分裂后细胞的APE1功能是非常有益的。在这项研究中，我们选择分化的SH-SY5Y细胞作为有丝分裂后的细胞系模型，以探讨药物诱导的APE1DNA修复或氧化还原活性的降低是否有助于有丝分裂后细胞在DNA氧化损伤条件下的生长和存活。在这里，我们证明了WT-APE1或C65-APE1(具有修复能力)的过表达导致了过氧化氢暴露后细胞存活率的显著增加。然而，177/226-APE1(修复缺陷)没有显示出保护作用。甲氧胺(MX)的使用进一步证实了这一现象，甲氧胺(MX)阻断了APE1的修复活性，导致分化的SH-SY5Y细胞和氧化DNA损伤处理后的神经元培养中细胞杀伤和凋亡增加。用小分子抑制剂阻断APE1氧化还原功能，BQP不会降低氧化DNA损伤处理后SH-SY5Y细胞或神经元培养的存活率。我们的结果表明，APE1的DNA修复功能有助于DNA氧化损伤后未分裂的有丝分裂后细胞的存活。
Oxidative DNA damage has been implicated in a number of central nervous system pathologies. The base excision repair (BER) pathway is one of the most important cellular protection mechanisms that respond to oxidative DNA damage. Human apurinic (apyrimidinic) endonuclease/redox effector factor (APE1/Ref-1 or APE1) is an essential enzyme in the BER pathway and is expressed in both mitotic and post-mitotic cells in humans. In neurons, a reduction of APE1 expression increases chemotherapy-induced cytotoxicity, while overexpression of APE1 protects cells against the cytotoxicity. However, given the multiple functions of APE1, knockdown of total APE1 is not completely informative of whether it is the redox or DNA repair activity, or interactions with other proteins. Therefore, the use of selective small molecules that can block each function independent of the other is of great benefit in ascertaining APE1 function in post-mitotic cells. In this study, we chose differentiated SH-SY5Y cells as our post-mitotic cell line model to investigate whether a drug-induced decrease in APE1 DNA repair or redox activity contributes to the growth and survival of post-mitotic cells under oxidative DNA damaging conditions. Here, we demonstrate that overexpression of WT-APE1 or C65-APE1 (repair competent) results in significant increase in cell viability after exposure to H2O2. However, the 177/226-APE1 (repair deficient) did not show a protective effect. This phenomenon was further confirmed by the use of methoxyamine (MX), which blocks the repair activity of APE1 that results in enhanced cell killing and apoptosis in differentiated SH-SY5Y cells and in neuronal cultures after oxidative DNA damaging treatments. Blocking APE1 redox function by a small molecule inhibitor, BQP did not decrease viability of SH-SY5Y cells or neuronal cultures following oxidative DNA damaging treatments. Our results demonstrate that the DNA repair function of APE1 contributes to the survival of nondividing post-mitotic cells following oxidative DNA damage.
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DOI：
10.1016/j.mrfmmm.2008.04.008
发表时间：
2008-08-25
影响因子：
2.3
作者：
Georgiadis, M. M.;Luo, M.;Kelley, M. R.
通讯作者：
Kelley, M. R.
影响因子：
14.9
作者：
Kulkarni, Avanti;McNeill, Daniel R.;Wilson, David M., III
通讯作者：
Wilson, David M., III
影响因子：
11.2
作者：
Jiang, Yanlin;Guo, Chunlu;Vasko, Michael R.;Kelley, Mark R.
通讯作者：
Kelley, Mark R.
影响因子：
--
作者：
CHANG, MY;MING, S;FORMAN, HJ
通讯作者：
FORMAN, HJ
影响因子：
11.5
作者：
Fishel, Melissa L.;He, Ying;Kelley, Mark R.
通讯作者：
Kelley, Mark R.