Uptake of Shiga-toxigenic Escherichia coli SubAB by HeLa cells requires an actin- and lipid raft-dependent pathway.
Uptake of Shiga-toxigenic Escherichia coli SubAB by HeLa cells requires an actin- and lipid raft-dependent pathway.
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DOI：
10.1111/cmi.12315
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发表时间：
2014-10
影响因子：
3.4
通讯作者：
Yahiro K
中科院分区：
文献类型：
--
作者：
Nagasawa S;Ogura K;Tsutsuki H;Saitoh H;Moss J;Iwase H;Noda M;Yahiro K
新型细胞毒性因子枯草杆菌酶细胞毒素 (SubAB) 主要由非 O157 志贺毒素大肠杆菌 (STEC) 产生。 SubAB 裂解内质网 (ER) 中的分子伴侣 BiP/GRP78，导致 RNA 依赖性蛋白激酶 (PKR) 样 ER 激酶 (PERK) 的激活，随后导致 caspase 依赖性细胞死亡。然而，HeLa 细胞中 SubAB 的摄取机制尚不清楚。在这项研究中，采用了多种抑制剂和 siRNA 来表征 SubAB 摄取过程。 SubAB 诱导的 BiP 裂解被高浓度的 Dynasore、甲基-β-环糊精 (mβCD) 和 Filipin III 抑制，但在网格蛋白、动力 I/II、caveolin1 和 Caveolin2 敲低细胞中不受抑制。我们观察到 SubAB 处理导致显着的肌动蛋白重排，例如质膜泡的形成，同时液体摄取显着增加。共聚焦显微镜分析表明，SubAB 摄取需要肌动蛋白细胞骨架重塑和脂筏胆固醇。此外，在细胞内的去污剂抗性结构域（DRM）结构中发现了内化的 SubAB。有趣的是，IPA-3 是丝氨酸/苏氨酸激酶 p21 激活激酶 (PAK1)（巨胞饮作用的重要蛋白）的抑制剂，可直接抑制 SubAB 介导的 BiP 裂解和 SubAB 内化。因此，我们的研究结果表明，SubAB 使用脂筏和肌动蛋白依赖性途径，而不是网格蛋白、小窝蛋白和动力依赖性途径作为其主要内吞易位途径。
The novel cytotoxic factor Subtilase cytotoxin (SubAB) is produced mainly by non-O157 Shiga-toxigenic Escherichia coli (STEC). SubAB cleaves the molecular chaperone BiP/GRP78 in the endoplasmic reticulum (ER), leading to activation of RNA-dependent protein kinase (PKR)-like ER kinase (PERK), followed by caspase-dependent cell death. However, the SubAB uptake mechanism in HeLa cells is unknown. In this study, a variety of inhibitors and siRNAs were employed to characterize the SubAB uptake process. SubAB-induced BiP cleavage was inhibited by high concentrations of Dynasore, and methyl-β-cyclodextrin (mβCD) and Filipin III, but not suppressed in clathrin-, dynamin I/II-, caveolin1- and caveolin2-knockdown cells. We observed that SubAB treatment led to dramatic actin rearrangements, e.g., formation of plasma membrane blebs, with a significant increase in fluid uptake. Confocal microscopy analysis showed that SubAB uptake required actin cytoskeleton remodeling and lipid raft cholesterol. Furthermore, internalized SubAB in cells was found in the detergent-resistant domain (DRM) structure. Interestingly, IPA-3, an inhibitor of serine/threonine kinase p21-activated kinase (PAK1), an important protein of macropinocytosis, directly inhibited SubAB-mediated BiP cleavage and SubAB internalization. Thus, our findings suggest that SubAB uses lipid raft- and actin-dependent, but not clathrin-, caveolin- and dynamin-dependent pathways as its major endocytic translocation route.
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影响因子：
5.4
作者：
Kaelin, Stefan;Amstutz, Beat;Greber, Urs F.
通讯作者：
Greber, Urs F.
内吞作用未插入：多种进入细胞的方法。
影响因子：
44.1
作者：
Kumari S;Mg S;Mayor S
通讯作者：
Mayor S
影响因子：
11.8
作者：
Macia, Eric;Ehrlich, Marcelo;Kirchhausen, Tomas
通讯作者：
Kirchhausen, Tomas
影响因子：
5.3
作者：
Jiang, Min;Chen, Gong
通讯作者：
Chen, Gong
影响因子：
64.8
作者：
Byres, Emma;Paton, Adrienne W.;Paton, James C.;Lofling, Jonas C.;Smith, David F.;Wilce, Matthew C. J.;Talbot, Ursula M.;Chong, Damien C.;Yu, Hai;Huang, Shengshu;Chen, Xi;Varki, Nissi M.;Varki, Ajit;Rossjohn, Jamie;Beddoe, Travis
通讯作者：
Beddoe, Travis