Thiol agents and Bcl-2 identify an alphavirus-induced apoptotic pathway that requires activation of the transcription factor NF-kappa B.
Thiol agents and Bcl-2 identify an alphavirus-induced apoptotic pathway that requires activation of the transcription factor NF-kappa B.
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DOI：
10.1083/jcb.131.5.1149
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发表时间：
1995-12
期刊：
影响因子：
--
通讯作者：
Ratan RR
中科院分区：
文献类型：
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作者：
Lin KI;Lee SH;Narayanan R;Baraban JM;Hardwick JM;Ratan RR
氧化应激已被认为是凋亡性死亡的常见介质。为了进一步研究氧化剂在这一过程中的作用，我们研究了抗氧化剂对辛德毕斯病毒（SV）诱导的两种细胞系AT-3（前列腺癌细胞系）和N18（神经母细胞瘤细胞系）凋亡的影响。巯基抗氧化剂，N-乙酰半胱氨酸（NAC），在30 mM以上的浓度，完全废除SV诱导的AT-3和N18细胞凋亡。NAC的作用不能归因于抑制病毒进入或病毒复制、细胞外渗透压的变化或细胞谷胱甘肽水平的增加，也不能被痕量金属螯合剂、脂质过氧化抑制剂或过氧化物清除剂所模拟。相比之下，包括吡咯烷二硫代氨基甲酸酯（PDTC，75 μ M）的其他硫醇试剂是保护性的。由于NAC和PDTC是转录因子NF-κ B最有效的抑制剂，我们研究了SV在细胞凋亡的形态学或生化证据出现之前激活NF-κ B的能力。在感染后数小时内，SV诱导AT-3和N18细胞核NF-κ B B活性显著增加; NAC和PDTC可抑制这种激活。AT-3细胞中bcl-2的过表达已被证明可抑制SV诱导的细胞凋亡，也可抑制SV诱导的NF-κ B活化。为了确定NF-κ B活化是否是SV诱导的这些细胞凋亡所必需的，我们使用具有共有NF-κ B序列的双链寡核苷酸作为转录因子诱饵（TFD）来抑制NF-κ B与天然DNA位点的结合。野生型，但不是突变体，TFD抑制SV诱导的AT-3细胞凋亡。相反，TFD抑制N18细胞中NF-κ B核活性并不能阻止SV诱导的细胞凋亡。总之，这些观察结果定义了一个细胞类型特异性，转录因子信号通路所需的SV诱导的细胞凋亡。了解Bcl-2和巯基药物抑制AT-3细胞中SV诱导的核NF-κ B活性的确切机制，可能有助于了解这些药物的多能抗凋亡作用。
Oxidative stress has been proposed as a common mediator of apoptotic death. To investigate further the role of oxidants in this process we have studied the effects of antioxidants on Sindbis virus (SV)-induced apoptosis in two cell lines, AT-3 (a prostate carcinoma line) and N18 (a neuroblastoma line). The thiol antioxidant, N-acetylcysteine (NAC), at concentrations above 30 mM, completely abrogates SV-induced apoptosis in AT-3 and N18 cells. The effects of NAC cannot be attributed to inhibition of viral entry or viral replication, changes in extracellular osmolarity or to increases in cellular glutathione levels, nor can they be mimicked by chelators of trace metals, inhibitors of lipid peroxidation or peroxide scavengers. In contrast, other thiol agents including pyrrolidine dithiocarbamate (PDTC, 75 microM) are protective. Because NAC and PDTC are among the most effective inhibitors of the transcription factor NF-kappa B, we examined SV's ability to activate NF-kappa B before the onset of morphologic or biochemical evidence of apoptosis. Within hours of infection, SV induced a robust increase in nuclear NF-kappa B activity in AT-3 and N18 cells; this activation was suppressible by NAC and PDTC. Over-expression of bcl-2 in AT-3 cells, which has been shown to inhibit SV-induced apoptosis, also inhibits SV-induced NF-kappa B activation. To determine if NF-kappa B activation is necessary for SV- induced apoptosis in these cells, we used double stranded oligonucleotides with consensus NF-kappa B sequences as transcription factor decoys (TFDs) to inhibit NF-kappa B binding to native DNA sites. Wild-type, but not mutant, TFDs inhibit SV-induced apoptosis in AT-3 cells. In contrast, TFD inhibition of NF-kappa B nuclear activity in N18 cells did not prevent SV-induced apoptosis. Taken together, these observations define a cell type-specific, transcription factor signaling pathway necessary for SV-induced apoptosis. Understanding the precise mechanism by which Bcl-2 and thiol agents inhibit SV-induced nuclear NF-kappa B activity in AT-3 cells may provide insights into the pluripotent antiapoptotic actions of these agents.