谷胱甘肽S-转移酶π通过抑制ASK1-MKK7-JNK通道保护血清撤离诱导的293细胞死亡

摘要

Glutathione S-transferase π(GSTπ) protects cells from death by altering intracellular oxidative stress. In order to understand the mechanism of GSTπ protection, a cell death model induced by serum depletion as the stress was established. Cotransfection of apoptosis signal-regulating kinase 1 (ASK1) and GSTπ cDNA was performed to elucidate the impact of GSTπ on ASK1 activity, as well as on its downstream signals, MKK7 and JNK, and to elucidate the potential protection of GSTπ on 293 cell death induced by serum depletion. The dominant negative mutant of JNK was used to explore if the blocking of the JNK pathway led to cell death inhibition. It was found that GSTπ had a dose-dependent inhibitory effect on activation induced by serum depletion,and also on inhibition both on MKK7 and JNK. Intracellular expression of GSTπ significantly inhibited serum depletion-induced cell death. Blocking the JNK pathway by transfection of a dominant negative form of JNK (JNKAPF) brought about significant inhibition of cell death induced by serum depletion with an inhibiting rate as high as 15 %. All the results suggest that the mechanism of GSTπ protection on serum depletion-induced cell death works through an ASK1-MKK7-JNK pathway.%谷胱甘肽S-转移酶π(glutathione S-transferaseπ，GSTπ)通过消除细胞内的氧化应激(oxidative stress)来保护细胞免于死亡。为了阐明细胞内GSTπ保护细胞的机制，建立了血清撤离(serum depletion)诱导细胞凋亡的模型。用凋亡信号调节激酶(apoptosis signal-regulating kinase 1，ASK1)和GSTπ cDNA转染293细胞，来研究GSTπ对ASK1激酶活性及其对下游激酶MKK7和JNK活性的影响及对细胞死亡的保护作用；使用负显性(dominantnegative)JNK突变体转染293细胞，来探讨阻断JNK通路对细胞死亡的抑制作用。结果发现GSTπ对血清撤离带来的ASK1激活呈剂量依赖性抑制效应，对MKK7和JNK也具有明显的抑制作用；GSTπ的细胞内表达可明显抑制血清撤离带来的细胞死亡；用JNK无活性突变体转染阻断JNK通路，对血清撤离引起的细胞死亡有明显抑制作用，抑制率达15％。表明GSTπ对血清撤离引起的细胞损伤的保护机制是通过抑制ASK1-MKK7-JNK通路而发挥作用的。