Acute cadmium exposure inactivates thioltransferase (glutaredoxin), inhibits intracellular reduction of protein-glutathionyl-mixed disulfides, and initiates apoptosis
Acute cadmium exposure inactivates thioltransferase (glutaredoxin), inhibits intracellular reduction of protein-glutathionyl-mixed disulfides, and initiates apoptosis
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DOI：
10.1074/jbc.m004097200
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发表时间：
2000-08-25
影响因子：
4.8
通讯作者：
Mieyal, JJ
中科院分区：
文献类型：
--
作者：
Chrestensen, CA;Starke, DW;Mieyal, JJ
氧化应激广泛影响细胞，启动调节途径以及细胞凋亡和坏死。一个关键的分子事件是蛋白质 S-谷胱甘肽化，而硫醇转移酶（谷氧还蛋白）是蛋白质-SSG 还原的特异性且有效的催化剂。在这项研究中，H9 和 Jurkat 细胞暴露于镉 30 分钟会抑制细胞内蛋白质-SSG 的减少，这与硫醇转移酶系统的抑制相关，这与硫醇转移酶是去谷胱甘肽的主要细胞内催化剂一致。硫氧还蛋白系统对总去谷胱甘肽酶活性的贡献很小。原位硫醇转移酶和 GSSG 还原酶显示出相似的剂量反应曲线（细胞外缓冲液中 10 μM 镉附近有 50% 抑制）。急性镉暴露也会引发细胞凋亡，H9 细胞比 Jurkat 更敏感。此外，用反义硫醇转移酶cDNA转染与细胞存活不相容。总的来说，这些数据表明硫醇转移酶在巯基稳态和细胞存活中具有至关重要的作用。在单独的实验中，镉抑制了硫醇转移酶和硫氧还蛋白系统的分离组分酶，与其活性位点的邻位二硫醇性质一致：硫醇转移酶（IC50大约为1μM），GSSG还原酶（IC50大约为μM），硫氧还蛋白（IC50大约为8μM），硫氧还蛋白还原酶（IC50大约为0.2μM）硫醇转移酶上邻位二硫醇的破坏（通过氧化为 C22-SS-C25；或 C25S 突变）可防止镉，与二硫醇螯合失活机制一致。
Oxidative stress broadly impacts cells, initiating regulatory pathways as well as apoptosis and necrosis. A key molecular event is protein S-glutathionylation, and thioltransferase (glutaredoxin) is a specific and efficient catalyst of protein-SSG reduction. In this study 30-min exposure of H9 and Jurkat cells to cadmium inhibited intracellular protein-SSG reduction, and this correlated with inhibition of the thioltransferase system, consistent with thioltransferase being the primary intracellular catalyst of deglutathionylation. The thioredoxin system contributed very little to total deglutathionylase activity. Thioltransferase and GSSG reductase in situ displayed similar dose-response curves (50% inhibition near 10 mu M cadmium in extracellular buffer). Acute cadmium exposure also initiated apoptosis, with H9 cells being more sensitive than Jurkat. Moreover, transfection with antisense thioltransferase cDNA was incompatible with cell survival. Collectively, these data suggest that thioltransferase has a vital role in sulfhydryl homeostasis and cell survival. In separate experiments, cadmium inhibited the isolated component enzymes of the thioltransferase and thioredoxin systems, consistent with the vicinal dithiol nature of their active sites: thioltransferase (IC50 approximate to 1 mu M), GSSG reductase (IC50 approximate to mu M), thioredoxin (IC50 approximate to 8 mu M), thioredoxin reductase (IC50 approximate to 0.2 mu M) Disruption of the vicinal dithiol on thioltransferase (via oxidation to C22-SS-C25; or C25S mutation) protected against cadmium, consistent with a dithiol chelation mechanism of inactivation.