The apoptosis/autophagy paradox:: autophagic vacuolization before apoptotic death
The apoptosis/autophagy paradox:: autophagic vacuolization before apoptotic death
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DOI：
10.1242/jcs.02447
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发表时间：
2005-07-15
影响因子：
4
通讯作者：
Kroemer, G
中科院分区：
文献类型：
--
作者：
González-Polo, RA;Boya, P;Kroemer, G
自噬细胞死亡的形态学特征是自噬液泡的积累。在这里，我们发现通过RNA干扰或同源重组使LAMP2失活会导致营养耗尽的细胞中发生自噬空泡化。缺乏 LAMP2 表达的细胞显示携带标记 LC3 的液泡积累增加，但 LC3 和溶酶体的共定位减少，表明自噬液泡和溶酶体之间的融合受到抑制。虽然表达饥饿的 LAMP2 细胞的一小部分线粒体与自噬溶酶体内的溶酶体标记物共定位，但从实验系统中去除 LAMP2 后没有发现这种共定位。值得注意的是，LAMP1 缺失没有这样的影响，并且不会加重 LAMP2 特异性小干扰 RNA 诱导的表型。血清和氨基酸饥饿的 LAMP2 阴性细胞表现出自噬空泡的积累，然后导致细胞死亡，并具有细胞凋亡的特征，例如线粒体跨膜电位丧失、半胱天冬酶激活和染色质浓缩。虽然半胱天冬酶抑制可延缓细胞死亡，但对线粒体没有保护作用。然而，通过过度表达 Bcl-2 或线粒体靶向的巨细胞病毒蛋白 vMIA 来稳定线粒体，可以阻止所有细胞凋亡的迹象。 Caspase 抑制和线粒体稳定均不能拮抗 LAMP2 缺陷细胞中的自噬空泡化。总而言之，这些数据表明自噬液泡的积累可能先于细胞凋亡。这些发现反对 1 型（细胞凋亡）和 2 型（自噬）细胞死亡之间的明确区别。
Autophagic cell death is morphologically characterized by an accumulation of autophagic vacuoles. Here, we show that inactivation of LAMP2 by RNA interference or by homologous recombination leads to autophagic vacuolization in nutrient-depleted cells. Cells that lack LAMP2 expression showed an enhanced accumulation of vacuoles carrying the marker LC3, yet a decreased colocalization of LC3 and lysosomes, suggesting that the fusion between autophagic vacuoles and lysosomes was inhibited. While a fraction of mitochondria from starved LAMP2-expressing cells colocalized with lysosomal markers, within autophagolysosomes, no such colocalization was found on removal of LAMP2 from the experimental system. Of note, LAMP1 depletion had no such effects and did not aggravate the phenotype induced by LAMP2-specific small interfering RNA. Serum and amino acid-starved LAMP2-negative cells exhibited an accumulation of autophagic vacuoles and then succumbed to cell death with hallmarks of apoptosis such as loss of the mitochondrial transmembrane potential, caspase activation and chromatin condensation. While caspase inhibition retarded cell death, it had no protective effect on mitochondria. Stabilization of mitochondria by overexpression of Bcl-2 or the mitochondrion-targeted cytomegalovirus protein vMIA, however, blocked all signs of apoptosis. Neither caspase inhibition nor mitochondrial stabilization antagonized autophagic vacuolization in LAMP2-deficient cells. Altogether, these data indicate that accumulation of autophagic vacuoles can precede apoptotic cell death. These findings argue against the clear-cut distinction between type 1 (apoptotic) and type 2 (autophagic) cell death.