S-Nitrosylation of the Death Receptor Fas Promotes Fas Ligand-Mediated Apoptosis in Cancer Cells
S-Nitrosylation of the Death Receptor Fas Promotes Fas Ligand-Mediated Apoptosis in Cancer Cells
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DOI：
10.1053/j.gastro.2011.02.053
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发表时间：
2011-06-01
期刊：
影响因子：
29.4
通讯作者：
Bettaieb, Ali
中科院分区：
文献类型：
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作者：
Leon-Bollotte, Lissbeth;Subramaniam, Selvakumar;Bettaieb, Ali
背景与目的：Fas属于肿瘤坏死因子受体家族，可诱导细胞凋亡。许多癌细胞表达Fas，但不经历Fas介导的凋亡。一氧化氮通过增加质膜上Fas的水平来逆转这种抗性。我们研究了NO影响Fas功能的机制。方法：将结肠癌和乳腺癌细胞系与NO供体三硝酸甘油酯或脂质A孵育;使用生物素开关测定监测Fas的S-亚硝基化。Fas构建体含有突变的半胱氨酸残基，防止S-亚硝基化被用来调查参与的S-亚硝基化Fas介导的细胞死亡。根据形态学标准监测细胞凋亡。结果：NO诱导Fas胞质部分半胱氨酸残基199和304的S-亚硝基化。在过度表达野生型Fas的癌细胞中，S-亚硝基化诱导Fas向脂筏募集并使细胞对Fas配体敏感。在表达半胱氨酸304被缬氨酸残基取代的突变型Fas的细胞中，NO介导的Fas向脂筏的转运受到影响，并且死亡诱导信号复合物和甘油三硝酸酯-Fas配体的协同作用被显著抑制。在表达半胱氨酸199突变的Fas的细胞中未观察到这些效应。结论：我们鉴定了Fas胞质结构域的翻译后修饰（半胱氨酸残基199和304的S-亚硝基化）。半胱氨酸304的S-亚硝基化促进Fas向脂筏的再分布，形成死亡诱导信号复合物，并诱导细胞死亡。
BACKGROUND & AIMS: Fas belongs to the family of tumor necrosis factor receptors which induce apoptosis. Many cancer cells express Fas but do not undergo Fas-mediated apoptosis. Nitric oxide reverses this resistance by increasing levels of Fas at the plasma membrane. We studied the mechanisms by which NO affects Fas function. METHODS: Colon and mammary cancer cell lines were incubated with the NO donor glyceryl trinitrate or lipid A; S-nitrosylation of Fas was monitored using the biotin switch assay. Fas constructs that contained mutations at cysteine residues that prevent S-nitrosylation were used to investigate the involvement of S-nitrosylation in Fas-mediated cell death. Apoptosis was monitored according to morphologic criteria. RESULTS: NO induced S-nitrosylation of cysteine residues 199 and 304 in the cytoplasmic part of Fas. In cancer cells that over-expressed wild-type Fas, S-nitrosylation induced Fas recruitment to lipid rafts and sensitized the cells to Fas ligand. In cells that expressed a mutant form of Fas in which cysteine 304 was replaced by valine residue, NO-mediated translocation of Fas to lipid rafts was affected and the death-inducing signal complex and synergistic effect of glyceryl trinitrate-Fas ligand were inhibited significantly. These effects were not observed in cells that expressed Fas with a mutation at cysteine 199. CONCLUSIONS: We identified post-translational modifications (S-nitrosylation of cysteine residues 199 and 304) in the cytoplasmic domain of Fas. S-nitrosylation at cysteine 304 promotes redistribution of Fas to lipid rafts, formation of the death-inducing signal complex, and induction of cell death.