Expression of the EspB Protein of Enteropathogenic Escherichia coli within HeLa Cells Affects Stress Fibers and Cellular Morphology

The EspB protein of enteropathogenic Escherichia coli (EPEC) is essential for the signaling events that lead to the accumulation of actin beneath intimately attached bacteria, a process that is known as the attaching and effacing effect. EspB is targeted to the host cell cytoplasm by a type III secretion apparatus. To determine the effect of intracellular EspB on the host cell cytoskeleton, we transfected HeLa cells with a plasmid containing the espB gene under the control of an inducible eukaryotic promoter. A HeLa cell clone that expressed espB mRNA and EspB protein after induction was selected for further study. The expression of EspB in these cells caused a dramatic change in cell morphology and a marked reduction in actin stress fibers. Cells expressing EspB were significantly impaired in their ability to support invasion by EPEC and Salmonella typhimurium. However, the expression of EspB within host cells could not compensate for the lack of EspB expression by an espB mutant strain of EPEC to restore attaching and effacing activity. These studies suggest that EspB is a cytoskeletal toxin that is translocated to the host cell cytoplasm, where it causes a redistribution of actin.

肠致病性大肠杆菌（enteropathogenic Escherichia coli, EPEC）的EspB蛋白，对于介导紧密黏附细菌下方肌动蛋白聚集的信号级联反应至关重要，这一过程被称为黏附消除效应（attaching and effacing effect）。EspB可通过Ⅲ型分泌系统（type III secretion apparatus）被靶向递送至宿主细胞细胞质。为探究胞内EspB对宿主细胞细胞骨架的影响，我们将携带有受诱导型真核启动子（inducible eukaryotic promoter）调控的espB基因的质粒转染海拉细胞（HeLa cells）。我们筛选出经诱导后可稳定表达espB mRNA与EspB蛋白的海拉细胞克隆，用于后续实验研究。此类细胞中EspB的表达会引发细胞形态的显著改变，并使肌动蛋白应力纤维（actin stress fibers）明显减少。表达EspB的细胞，其支持EPEC与鼠伤寒沙门氏菌（Salmonella typhimurium）侵袭的能力显著受损。然而，宿主细胞内的EspB表达无法弥补EPEC的espB突变株缺失EspB表达的缺陷，以恢复黏附消除活性。上述研究表明，EspB是一种被转运至宿主细胞细胞质的细胞骨架毒素，可在该处引发肌动蛋白的重分布。