Sandu Pamela, Crepin Valerie F, Drechsler Hauke, McAinsh Andrew D, Frankel Gad, Berger Cedric N
MRC Centre for Molecular Bacteriology and Infection, Department of Life Sciences, Imperial College London, London, United Kingdom.
Centre for Mechanochemical Cell Biology, Division of Biomedical Cell Biology, Warwick Medical School, University of Warwick, Coventry, United Kingdom.
Infect Immun. 2017 Aug 18;85(9). doi: 10.1128/IAI.00244-17. Print 2017 Sep.
Enterohemorrhagic (EHEC) is a diarrheagenic pathogen that colonizes the gut mucosa and induces attaching-and-effacing lesions. EHEC employs a type III secretion system (T3SS) to translocate 50 effector proteins that hijack and manipulate host cell signaling pathways, which allow bacterial colonization and subversion of immune responses and disease progression. The aim of this study was to characterize the T3SS effector EspW. We found in the sequenced O157:H7 and non-O157 EHEC strains as well as in Furthermore, a truncated version of EspW, containing the first 206 residues, is present in EPEC strains belonging to serotype O55:H7. Screening a collection of clinical EPEC isolates revealed that is present in 52% of the tested strains. We report that EspW modulates actin dynamics in a Rac1-dependent manner. Ectopic expression of EspW results in formation of unique membrane protrusions. Infection of Swiss cells with an EHEC deletion mutant induces a cell shrinkage phenotype that could be rescued by Rac1 activation via expression of the bacterial guanine nucleotide exchange factor, EspT. Furthermore, using a yeast two-hybrid screen, we identified the motor protein Kif15 as a potential interacting partner of EspW. Kif15 and EspW colocalized in cotransfected cells, while ectopically expressed Kif15 localized to the actin pedestals following EHEC infection. The data suggest that Kif15 recruits EspW to the site of bacterial attachment, which in turn activates Rac1, resulting in modifications of the actin cytoskeleton that are essential to maintain cell shape during infection.
肠出血性大肠杆菌（EHEC）是一种致泻性病原体，它定殖于肠道黏膜并诱导黏附-脱落性损伤。EHEC利用III型分泌系统（T3SS）转运50种效应蛋白，这些蛋白劫持并操纵宿主细胞信号通路，从而实现细菌定殖、颠覆免疫反应以及疾病进展。本研究的目的是对T3SS效应蛋白EspW进行特性分析。我们发现在已测序的O157:H7和非O157 EHEC菌株中以及在属于血清型O55:H7的肠致病性大肠杆菌（EPEC）菌株中存在EspW的截短版本，其包含前206个氨基酸残基。对一系列临床EPEC分离株进行筛选发现，52%的受试菌株中存在EspW。我们报告EspW以Rac1依赖的方式调节肌动蛋白动力学。EspW的异位表达导致独特的膜突出形成。用EHEC缺失突变体感染瑞士细胞会诱导细胞收缩表型，通过表达细菌鸟嘌呤核苷酸交换因子EspT激活Rac1可挽救该表型。此外，通过酵母双杂交筛选，我们鉴定出动力蛋白Kif15是EspW的潜在相互作用伴侣。Kif15和EspW在共转染细胞中共定位，而异位表达的Kif15在EHEC感染后定位于肌动蛋白菌毛。数据表明Kif15将EspW招募到细菌附着位点，进而激活Rac1，导致肌动蛋白细胞骨架发生改变，这对于在感染期间维持细胞形态至关重要。
