Torgersen Maria L, Engedal Nikolai, Pedersen Anne-Mari G, Husebye Harald, Espevik Terje, Sandvig Kirsten
Department of Biochemistry, Institute for Cancer Research, The Norwegian Radium Hospital, Oslo University Hospital, Montebello, Oslo, Norway.
FEMS Immunol Med Microbiol. 2011 Feb;61(1):63-75. doi: 10.1111/j.1574-695X.2010.00749.x. Epub 2010 Nov 3.
Infection with Shiga toxin (Stx)-producing, gram-negative bacteria can induce serious conditions such as dysentery and hemolytic uremic syndrome. In target cells, Stx is internalized by endocytosis, and travels through the Golgi apparatus and the endoplasmic reticulum to reach the cytosol, where it inhibits protein synthesis. Toll-like receptor 4 (TLR4) mediates the recognition of gram-negative bacteria. Here, we have investigated whether the cellular uptake and transport of Stx could involve TLR4. We found that upon small interfering RNA (siRNA)-mediated TLR4 depletion in epithelial colon carcinoma cells, Stx transport to the Golgi was strongly reduced, and this was primarily caused by diminished Stx cellular binding rather than by reduction in toxin uptake or endosome-to-Golgi transport. The reduced cellular binding of Stx upon siRNA-transfection was solely due to TLR4 depletion, because reconstitution of TLR4 expression by the introduction of an siRNA-resistant TLR4 gene completely abolished the TLR4-targeting siRNA-mediated effect. Importantly, the effect of TLR4 depletion was not restricted to cancer cells or epithelial cells, because primary human umbilical vein endothelial cells also displayed reduced Stx binding upon TLR4 depletion. These results indicate that although TLR4 is imperative in innate immunity against gram-negative bacteria, it may be exploited by bacterial toxins, for example Stx, to gain access and entry into cells.
感染产生志贺毒素（Stx）的革兰氏阴性菌可引发诸如痢疾和溶血尿毒综合征等严重病症。在靶细胞中，Stx通过内吞作用被内化，并穿过高尔基体和内质网到达细胞质溶胶，在那里它抑制蛋白质合成。Toll样受体4（TLR4）介导对革兰氏阴性菌的识别。在此，我们研究了Stx的细胞摄取和运输是否涉及TLR4。我们发现，在上皮结肠癌细胞中，通过小干扰RNA（siRNA）介导的TLR4缺失后，Stx向高尔基体的运输显著减少，这主要是由于Stx与细胞的结合减少，而非毒素摄取或内体到高尔基体的运输减少所致。siRNA转染后Stx与细胞的结合减少完全是由于TLR4缺失，因为通过引入抗siRNA的TLR4基因重建TLR4表达完全消除了靶向TLR4的siRNA介导的效应。重要的是，TLR4缺失的影响并不局限于癌细胞或上皮细胞，因为原代人脐静脉内皮细胞在TLR4缺失后也表现出Stx结合减少。这些结果表明，尽管TLR4在针对革兰氏阴性菌的先天免疫中至关重要，但它可能被细菌毒素（如Stx）利用，以进入细胞。
