Department of Microbiology and Immunology, Virginia Commonwealth University Medical Center, School of Medicine, Richmond, Virginia, Unites States of America.
PLoS Negl Trop Dis. 2021 Apr 15;15(4):e0009339. doi: 10.1371/journal.pntd.0009339. eCollection 2021 Apr.
Scrub typhus is a neglected tropical disease that threatens more than one billion people. If antibiotic therapy is delayed, often due to mis- or late diagnosis, the case fatality rate can increase considerably. Scrub typhus is caused by the obligate intracellular bacterium, Orientia tsutsugamushi, which invades phagocytes and endothelial cells in vivo and diverse tissue culture cell types in vitro. The ability of O. tsutsugamushi to replicate in the cytoplasm indicates that it has evolved to counter eukaryotic host cell immune defense mechanisms. The transcription factor, NF-κB, is a tightly regulated initiator of proinflammatory and antimicrobial responses. Typically, the inhibitory proteins p105 and IκBα sequester the NF-κB p50:p65 heterodimer in the cytoplasm. Canonical activation of NF-κB via TNFα involves IKKβ-mediated serine phosphorylation of IκBα and p105, which leads to their degradation and enables NF-κB nuclear translocation. A portion of p105 is also processed into p50. O. tsutsugamushi impairs NF-κB translocation into the nucleus, but how it does so is incompletely defined.
Western blot, densitometry, and quantitative RT-PCR analyses of O. tsutsugamushi infected host cells were used to determine if the pathogen's ability to inhibit NF-κB is linked to modulation of p105. Results demonstrate that p105 levels are elevated several-fold in O. tsutsugamushi infected HeLa and RF/6A cells with only a nominal increase in p50. The O. tsutsugamushi-stimulated increase in p105 is bacterial dose- and protein synthesis-dependent, but does not occur at the level of host cell transcription. While TNFα-induced phosphorylation of p105 serine 932 proceeds unhindered in infected cells, p105 levels remain elevated and NF-κB p65 is retained in the cytoplasm.
O. tsutsugamushi specifically stabilizes p105 to inhibit the canonical NF-κB pathway, which advances understanding of how it counters host immunity to establish infection.
恙虫病是一种被忽视的热带病，威胁着超过 10 亿人的生命。如果抗生素治疗延迟，通常是由于误诊或延迟诊断，病死率会显著增加。恙虫病是由专性细胞内细菌恙虫东方体引起的，它在体内侵入吞噬细胞和内皮细胞，并在体外侵入多种组织培养细胞类型。恙虫东方体在细胞质中复制的能力表明，它已经进化到可以对抗真核宿主细胞的免疫防御机制。转录因子 NF-κB 是一种严格调控的促炎和抗菌反应的启动子。通常，抑制蛋白 p105 和 IκBα 将 NF-κB p50:p65 异二聚体隔离在细胞质中。通过 TNFα 对 NF-κB 的经典激活涉及 IKKβ 介导的 IκBα 和 p105 的丝氨酸磷酸化，这导致它们的降解，并使 NF-κB 核易位。一部分 p105 也被加工成 p50。恙虫东方体损害 NF-κB 向核内易位，但具体机制尚不完全清楚。
使用 Western blot、密度测定和定量 RT-PCR 分析恙虫东方体感染的宿主细胞，以确定病原体抑制 NF-κB 的能力是否与 p105 的调节有关。结果表明，恙虫东方体感染的 HeLa 和 RF/6A 细胞中 p105 的水平升高了数倍，而 p50 仅略有增加。恙虫东方体刺激的 p105 增加与细菌剂量和蛋白质合成有关，但不发生在宿主细胞转录水平。虽然感染细胞中 TNFα 诱导的 p105 丝氨酸 932 磷酸化不受阻碍，但 p105 水平仍然升高，NF-κB p65 仍滞留在细胞质中。
恙虫东方体特异性稳定 p105 以抑制经典的 NF-κB 途径，这有助于了解它如何对抗宿主免疫以建立感染。
