Dieldrin induces apoptosis by promoting caspase-3-dependent proteolytic cleavage of protein kinase Cδ in dopaminergic cells:: Relevance to oxidative stress and dopaminergic degeneration
Dieldrin induces apoptosis by promoting caspase-3-dependent proteolytic cleavage of protein kinase Cδ in dopaminergic cells:: Relevance to oxidative stress and dopaminergic degeneration
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DOI：
10.1016/s0306-4522(03)00226-4
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发表时间：
2003-01-01
期刊：
影响因子：
3.3
通讯作者：
Kanthasamy, AG
中科院分区：
文献类型：
--
作者：
Kitazawa, M;Anantharam, V;Kanthasamy, AG
我们以前报道，狄氏剂，帕金森氏病的发展的潜在环境危险因素之一，诱导多巴胺能细胞凋亡，通过产生氧化应激。在这里，我们证明，caspase-3依赖的蛋白激酶C δ（PKC δ）的蛋白水解激活介导，以及调节多巴胺能细胞中狄氏剂诱导的凋亡级联反应。将PC 12细胞暴露于狄氏剂（100-300 μ M）导致细胞色素C的快速释放，随后以时间和剂量依赖性方式激活半胱天冬酶-9和半胱天冬酶-3。超氧化物歧化酶模拟锰（III）四（4-苯甲酸）卟啉氯化物显着衰减狄氏醇诱导的细胞色素C的释放，表明活性氧可能有助于激活促凋亡因子。有趣的是，狄氏剂蛋白水解裂解天然PKC δ成一个41 kDa的催化亚基和一个38 kDa的调节亚基激活激酶。用50-100 μ M浓度的半胱天冬酶抑制剂苄氧基羰基-Val-Ala-Asp-氟甲基酮（Z-VAD-FMK）和苄氧基羰基-Asp-Glu-Val-Asp-氟甲基酮（Z-DEVD-FMK）预处理完全抑制狄氏醇诱导的PKC δ的蛋白水解裂解和激酶活性的诱导，表明PKC δ的蛋白水解活化是半胱天冬酶-3依赖性的。此外，Z-VAD-FMK、Z-DEVD-FMK或PKC δ特异性抑制剂rottlerin几乎完全阻断狄氏醇诱导的DNA片段化。由于狄氏剂显著增加（40-80倍）caspase-3活性，我们研究了蛋白水解激活的PKC δ是否通过正反馈激活放大caspase-3。PKC 8抑制剂rottlerin（3-20 μ M）剂量依赖性地减弱狄氏醇诱导的caspase-3活性，表明PKC δ对caspase-3的正反馈激活。事实上，通过蛋白质递送系统递送催化活性的重组PKC δ显著激活PC 12细胞中的半胱天冬酶-3。最后，在大鼠中脑多巴胺能神经元细胞中激酶失活的PKC δ（K376 R）突变体的过表达减弱了狄氏剂诱导的caspase-3活性和DNA片段化，进一步证实了PKC δ在多巴胺能细胞中的促凋亡功能。总之，我们得出结论，caspase-3依赖的蛋白水解激活PKC δ是一个关键的事件，狄氏剂诱导的多巴胺能细胞的凋亡细胞死亡。(C)2003年IBRO。由爱思唯尔科技有限公司出版。保留所有权利。
We previously reported that dieldrin, one of the potential environmental risk factors for development of Parkinson's disease, induces apoptosis in dopaminergic cells by generating oxidative stress. Here, we demonstrate that the caspase-3-dependent proteolytic activation of protein kinase Cdelta (PKCdelta) mediates as well as regulates the dieldrin-induced apoptotic cascade in dopaminergic cells. Exposure of PC12 cells to dieldrin (100-300 muM) results in the rapid release of cytochrome C, followed by the activation of caspase-9 and caspase-3 in a time- and dose-dependent manner. The superoxide dismutase mimetic Mn(III)tetrakis(4-benzoic acid)porphyrin chloride significantly attenuates dieldrin-induced cytochrome C release, indicating that reactive oxygen species may contribute to the activation of pro-apoptotic factors. Interestingly, dieldrin proteolytically cleaves native PKCdelta into a 41 kDa catalytic subunit and a 38 kDa regulatory subunit to activate the kinase. The dieldrin-induced proteolytic cleavage of PKCdelta and induction of kinase activity are completely inhibited by pretreatment with 50-100 muM concentrations of the caspase inhibitors benzyloxycarbonyl-Val-Ala-Asp-fluoromethylketone (Z-VAD-FMK) and benzyloxycarbonyl-Asp-Glu-Val-Asp-fluoromethylketone (Z-DEVD-FMK), indicating that the proteolytic activation of PKCdelta is caspase-3-dependent. Additionally, Z-VAD-FMK, Z-DEVD-FMK or the PKCdelta specific inhibitor rottlerin almost completely block dieldrin-induced DNA fragmentation. Because dieldrin dramatically increases (40-80-fold) caspase-3 activity, we examined whether proteolytically activated PKCdelta amplifies caspase-3 via positive feedback activation. The PKC8 inhibitor rottlerin (3-20 muM) dose-dependently attenuates dieldrin-induced caspase-3 activity, suggesting positive feedback activation of caspase-3 by PKCdelta. Indeed, delivery of catalytically active recombinant PKCdelta via a protein delivery system significantly activates caspase-3 in PC12 cells. Finally, overexpression of the kinase-inactive PKCdelta(K376R) mutant in rat mesencephalic dopaminergic neuronal cells attenuates dieldrin-induced caspase-3 activity and DNA fragmentation, further confirming the pro-apoptotic function of PKCdelta in dopaminergic cells. Together, we conclude that caspase-3-dependent proteolytic activation of PKCdelta is a critical event in dieldrin-induced apoptotic cell death in dopaminergic cells. (C) 2003 IBRO. Published by Elsevier Science Ltd. All rights reserved.