Helicobacter pylori VacA induces apoptosis by accumulation of connexin 43 in autophagic vesicles via a Rac1/ERK-dependent pathway.
Helicobacter pylori VacA induces apoptosis by accumulation of connexin 43 in autophagic vesicles via a Rac1/ERK-dependent pathway.
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DOI：
10.1038/cddiscovery.2015.35
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发表时间：
2015
影响因子：
7
通讯作者：
Hirayama T
中科院分区：
文献类型：
--
作者：
Yahiro K;Akazawa Y;Nakano M;Suzuki H;Hisatune J;Isomoto H;Sap J;Noda M;Moss J;Hirayama T
幽门螺杆菌（H.幽门螺杆菌）产生空泡细胞毒素（VacA），一种与胃炎症和溃疡有关的强效蛋白毒素。最近的研究表明，连接蛋白（Cxs），这是负责在间隙连接（GJs）的细胞内通讯以及细胞内稳态，参与VacA诱导的细胞死亡。我们现在证明在AZ-521细胞中，VacA增加细胞质Cx43，伴随着LC 3-II的产生，在时间和剂量依赖性的方式没有诱导Cx43 mRNA的表达。抑制VacA诱导的Rac 1活性阻止ERK磷酸化和Cx43的增加。ERK活性的抑制和N-乙酰半胱氨酸的加入抑制了Cx43和LC 3-II的VacA依赖性增加。DIDS是一种阴离子选择性抑制剂，可抑制Cx43的VacA依赖性增加，表明VacA通道活性参与了该途径。通过共聚焦显微镜，Cx43增加VacA主要定位在富含胆固醇，洗涤剂抗性膜，包括GJ，和Cx43的一部分被纳入内吞囊泡和自噬溶酶体。在H. pylori感染的患者与健康对照组相比，表明病原体在体内引起类似的效果。我们的研究结果表明，VacA介导的自噬作用抑制Cx43的周转，导致细胞质中的水平增加，最终导致凋亡性细胞死亡。
Helicobacter pylori (H. pylori) produces vacuolating cytotoxin (VacA), a potent protein toxin, which is associated with gastric inflammation and ulceration. Recent studies demonstrated that connexins (Cxs), which are responsible for intracellular communication at gap junctions (GJs) as well as cell homeostasis, participate in VacA-induced cell death. We now demonstrate in AZ-521 cells that VacA increased cytoplasmic Cx43, accompanied by LC3-II generation in a time- and dose-dependent manner without induction of Cx43 mRNA expression. Inhibition of VacA-induced Rac1 activity prevented ERK phosphorylation and the increase in Cx43. Suppression of ERK activity and addition of N-acetyl-cysteine inhibited VacA-dependent increase in Cx43 and LC3-II. DIDS, an anion-selective inhibitor, suppressed VacA-dependent increase in Cx43, suggesting that VacA channel activity was involved in this pathway. By confocal microscopy, Cx43 increased by VacA was predominately localized in cholesterol-rich, detergent-resistant membranes including GJs, and a fraction of Cx43 was incorporated in endocytotic vesicles and autophagolysosomes. Accumulation of Cx43 was also observed in gastric mucosa from H. pylori-infected patients compared with healthy controls, suggesting that the pathogen caused a similar effect in vivo. Our findings show that VacA-mediated effects on autophagy inhibits turnover of Cx43, resulting in increased levels in the cytoplasm, leading eventually to apoptotic cell death.