Rippo M R, Malisan F, Ravagnan L, Tomassini B, Condo I, Costantini P, Susin S A, Rufini A, Todaro M, Kroemer G, Testi R
Laboratory of Signal Transduction, Department of Experimental Medicine and Biochemical Sciences, University of Rome 'Tor Vergata', 00133 Rome, Italy.
FASEB J. 2000 Oct;14(13):2047-54. doi: 10.1096/fj.99-1028com.
Lipid and glycolipid diffusible mediators are involved in the intracellular progression and amplification of apoptotic signals. GD3 ganglioside is rapidly synthesized from accumulated ceramide after the clustering of death-inducing receptors and triggers apoptosis. Here we show that GD3 induces dissipation of DeltaPsim and swelling of isolated mitochondria, which results in the mitochondrial release of cytochrome c, apoptosis inducing factor, and caspase 9. Soluble factors released from GD3-treated mitochondria are sufficient to trigger DNA fragmentation in isolated nuclei. All these effects can be blocked by cyclosporin A, suggesting that GD3 is acting at the level of the permeability transition pore complex. We found that endogenous GD3 accumulates within mitochondria of cells undergoing apoptosis after ceramide exposure. Accordingly, suppression of GD3 synthase (ST8) expression in intact cells substantially prevents ceramide-induced DeltaPsim dissipation, indicating that endogenously synthesized GD3 induces mitochondrial changes in vivo. Finally, enforced expression of bcl-2 significantly prevents GD3-induced mitochondrial changes, caspase 9 activation, and apoptosis. These results show that mitochondria are a key destination for apoptogenic GD3 ganglioside along the lipid pathway to programmed cell death and indicate that relevant GD3 targets are under bcl-2 control.
脂质和糖脂类可扩散介质参与凋亡信号的细胞内进展和放大。死亡诱导受体聚集后，神经节苷脂GD3由积累的神经酰胺快速合成并触发凋亡。在此我们表明，GD3诱导线粒体跨膜电位（ΔΨm）的消散和分离线粒体的肿胀，这导致细胞色素c、凋亡诱导因子和半胱天冬酶9从线粒体释放。GD3处理的线粒体释放的可溶性因子足以触发分离细胞核中的DNA片段化。所有这些效应均可被环孢菌素A阻断，提示GD3作用于通透性转换孔复合体水平。我们发现，神经酰胺暴露后，内源性GD3在经历凋亡的细胞线粒体内积累。因此，在完整细胞中抑制GD3合酶（ST8）的表达可显著防止神经酰胺诱导的ΔΨm消散，表明内源性合成的GD3在体内诱导线粒体变化。最后，强制表达bcl-2可显著防止GD3诱导的线粒体变化、半胱天冬酶9激活和凋亡。这些结果表明，线粒体是沿脂质途径走向程序性细胞死亡的凋亡原性神经节苷脂GD3的关键作用靶点，并表明相关的GD3靶点受bcl-2调控。
