Department of Molecular Genetics and Microbiology, Stony Brook University, Stony Brook, New York, USA.
Infect Immun. 2014 May;82(5):2068-78. doi: 10.1128/IAI.00044-14. Epub 2014 Mar 10.
Francisella tularensis is a facultative intracellular, Gram-negative pathogen and the causative agent of tularemia. We previously identified TolC as a virulence factor of the F. tularensis live vaccine strain (LVS) and demonstrated that a ΔtolC mutant exhibits increased cytotoxicity toward host cells and elicits increased proinflammatory responses compared to those of the wild-type (WT) strain. TolC is the outer membrane channel component used by the type I secretion pathway to export toxins and other bacterial virulence factors. Here, we show that the LVS delays activation of the intrinsic apoptotic pathway in a TolC-dependent manner, both during infection of primary macrophages and during organ colonization in mice. The TolC-dependent delay in host cell death is required for F. tularensis to preserve its intracellular replicative niche. We demonstrate that TolC-mediated inhibition of apoptosis is an active process and not due to defects in the structural integrity of the ΔtolC mutant. These findings support a model wherein the immunomodulatory capacity of F. tularensis relies, at least in part, on TolC-secreted effectors. Finally, mice vaccinated with the ΔtolC LVS are protected from lethal challenge and clear challenge doses faster than WT-vaccinated mice, demonstrating that the altered host responses to primary infection with the ΔtolC mutant led to altered adaptive immune responses. Taken together, our data demonstrate that TolC is required for temporal modulation of host cell death during infection by F. tularensis and highlight how shifts in the magnitude and timing of host innate immune responses may lead to dramatic changes in the outcome of infection.
土拉弗朗西斯菌是一种兼性细胞内革兰氏阴性病原体，也是土拉热的病原体。我们之前确定 TolC 是土拉弗朗西斯菌活疫苗株（LVS）的毒力因子，并证明与野生型（WT）菌株相比，ΔtolC 突变体能增加对宿主细胞的细胞毒性，并引发更强的促炎反应。TolC 是 I 型分泌途径的外膜通道组件，用于输出毒素和其他细菌毒力因子。在这里，我们表明 LVS 以 TolC 依赖的方式延迟内在凋亡途径的激活，无论是在原代巨噬细胞感染期间还是在小鼠器官定植期间。宿主细胞死亡的 TolC 依赖性延迟对于土拉弗朗西斯菌在细胞内复制龛中存活是必需的。我们证明 TolC 介导的凋亡抑制是一个主动过程，而不是由于 ΔtolC 突变体的结构完整性缺陷所致。这些发现支持了这样一种模型，即土拉弗朗西斯菌的免疫调节能力至少部分依赖于 TolC 分泌的效应物。最后，用 ΔtolC LVS 接种的小鼠能够抵抗致死性挑战并比 WT 接种的小鼠更快地清除挑战剂量，表明对 ΔtolC 突变体原发性感染的宿主反应改变导致了适应性免疫反应的改变。总之，我们的数据表明 TolC 是土拉弗朗西斯菌感染期间宿主细胞死亡的时间调节所必需的，并强调了宿主先天免疫反应的幅度和时间的变化如何导致感染结局的显著变化。
