Novel Subtilase Cytotoxin Produced by Shiga-Toxigenic Escherichia coli Induces Apoptosis in Vero Cells via Mitochondrial Membrane Damage
Novel Subtilase Cytotoxin Produced by Shiga-Toxigenic Escherichia coli Induces Apoptosis in Vero Cells via Mitochondrial Membrane Damage
复制标题
DOI：
10.1128/iai.01510-08
复制
发表时间：
2009-07-01
影响因子：
3.1
通讯作者：
Noda, Masatoshi
中科院分区：
文献类型：
--
作者：
Matsuura, Gen;Morinaga, Naoko;Noda, Masatoshi
枯草酶细胞毒素（SubAB）是由某些产志贺毒素大肠杆菌菌株产生的AB5细胞毒素。A亚基是一种类似枯草酶的丝氨酸蛋白酶，可切割内质网伴侣蛋白BiP，导致蛋白质合成的短暂抑制和细胞周期阻滞在G(1)期。通过DNA断裂和膜联蛋白V结合检测，我们发现SubAB，而非催化失活突变体SubAB(S272A)，诱导Vero细胞凋亡。SubAB诱导caspase-3、-7和-8的激活。caspase-3出现时间早于caspase-8，通过使用特异性的caspase抑制剂，可以确定caspase-3可能位于caspase-8的上游。通过膜联蛋白V结合检测，一种通用的caspase抑制剂阻断了subab诱导的细胞凋亡。SubAB还刺激线粒体释放细胞色素c，而caspase抑制剂不抑制这种释放。在HeLa细胞中，Apaf-1小干扰RNA抑制caspase-3活化，提示细胞色素c可能形成凋亡小体，导致caspase-3活化。这些数据表明，SubAB通过线粒体膜损伤诱导Vero细胞caspase依赖性凋亡。
Subtilase cytotoxin (SubAB) is an AB5 cytotoxin produced by some strains of Shiga-toxigenic Escherichia coli. The A subunit is a subtilase-like serine protease and cleaves an endoplasmic reticulum chaperone, BiP, leading to transient inhibition of protein synthesis and cell cycle arrest at G(1) phase. Here we show that SubAB, but not the catalytically inactive mutant SubAB(S272A), induced apoptosis in Vero cells, as detected by DNA fragmentation and annexin V binding. SubAB induced activation of caspase-3, -7, and -8. Caspase-3 appeared earlier than caspase-8, and by use of specific caspase inhibitors, it was determined that caspase-3 may be upstream of caspase-8. A general caspase inhibitor blocked SubAB-induced apoptosis, detected by annexin V binding. SubAB also stimulated cytochrome c release from mitochondria, which was not suppressed by caspase inhibitors. In HeLa cells, Apaf-1 small interfering RNA inhibited caspase-3 activation, suggesting that cytochrome c might form an apoptosome, leading to activation of caspase-3. These data suggested that SubAB induced caspase-dependent apoptosis in Vero cells through mitochondrial membrane damage.