Reinehr Roland, Görg Boris, Höngen Andrea, Häussinger Dieter
Clinic for Gastroenterology, Hepatology and Infectiology, Heinrich Heine University Düsseldorf, Germany.
J Biol Chem. 2004 Mar 12;279(11):10364-73. doi: 10.1074/jbc.M311997200. Epub 2003 Dec 16.
Epidermal growth factor receptor-dependent CD95-tyrosine phosphorylation was recently identified as an early step in apoptosis induction via the CD95 system (Reinehr, R., Schliess, F., and Häussinger, D. (2003) FASEB J. 17, 731-733). The effect of peroxynitrite (ONOO(-)) on modulation of the hyperosmotic and CD95 ligand (CD95L)-induced CD95 activation process was studied. Pretreatment of hepatocytes with ONOO(-) inhibited CD95L- and hyperosmolarity-induced CD95 membrane trafficking and formation of the death-inducing signaling complex, but not epidermal growth factor receptor activation and its association with CD95. Under these conditions, however, no tyrosine phosphorylation of CD95 occurred; instead, CD95 was tyrosine-nitrated. When ONOO(-) was added after induction of CD95-tyrosine phosphorylation by CD95L or hyperosmolarity, tyrosine nitration of CD95 was largely prevented and death-inducing signaling complex formation occurred. CD95-tyrosine nitration abolished the hyperosmotic sensitization of hepatocytes toward CD95L-induced apoptosis. Additionally, in CD95-yellow fluorescent protein-transfected Huh7-hepatoma cells, ONOO(-) induced CD95 Tyr nitration and prevented CD95L-induced Tyr phosphorylation and apoptosis. Tyrosine-nitrated CD95 was also found in rat livers derived from an in vivo model of endotoxinemia. The data suggest that CD95-tyrosine nitration prevents CD95 activation by inhibiting CD95-tyrosine phosphorylation. Apparently, CD95-tyrosine phosphorylation and nitration are mutually exclusive. The data identify critical tyrosine residues of CD95 as another target of the anti-apoptotic action of NO.
表皮生长因子受体依赖性CD95酪氨酸磷酸化最近被确定为通过CD95系统诱导细胞凋亡的早期步骤（Reinehr, R., Schliess, F., and Häussinger, D. (2003) FASEB J. 17, 731 - 733）。研究了过氧亚硝酸盐（ONOO(-)）对高渗和CD95配体（CD95L）诱导的CD95激活过程的调节作用。用ONOO(-)预处理肝细胞可抑制CD95L和高渗诱导的CD95膜转运以及死亡诱导信号复合物的形成，但不影响表皮生长因子受体的激活及其与CD95的结合。然而，在这些条件下，CD95未发生酪氨酸磷酸化；相反，CD95被酪氨酸硝化。当在CD95L或高渗诱导CD95酪氨酸磷酸化后加入ONOO(-)时，CD95的酪氨酸硝化在很大程度上被阻止，并且死亡诱导信号复合物形成。CD95酪氨酸硝化消除了肝细胞对CD95L诱导的细胞凋亡的高渗敏感性。此外，在转染了CD95 - 黄色荧光蛋白的Huh7肝癌细胞中，ONOO(-)诱导CD95酪氨酸硝化并阻止CD95L诱导的酪氨酸磷酸化和细胞凋亡。在源自内毒素血症体内模型的大鼠肝脏中也发现了酪氨酸硝化的CD95。数据表明，CD95酪氨酸硝化通过抑制CD95酪氨酸磷酸化来阻止CD95激活。显然，CD95酪氨酸磷酸化和硝化是相互排斥的。这些数据确定CD95的关键酪氨酸残基是NO抗凋亡作用的另一个靶点。
