Streptococcal autolysin promotes dysfunction of swine tracheal epithelium by interacting with vimentin.
Streptococcal autolysin promotes dysfunction of swine tracheal epithelium by interacting with vimentin.
复制标题
链球菌自脂蛋白通过与波形蛋白相互作用促进猪气管上皮的功能障碍。
DOI：
10.1371/journal.ppat.1010765
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发表时间：
2022-08
期刊：
影响因子：
6.7
通讯作者：
中科院分区：
文献类型：
--
作者：
猪链球菌2型（Streptococcussuisserotype 2，SS 2）是一种重要的人畜共患病原体，可引起感染性休克和肺炎。上呼吸道通常被认为是猪中SS 2的主要定植和进入部位，但其穿透呼吸屏障的机制仍不清楚。本研究从SS 2的TnYLB-1转座子插入突变体库中筛选到一个对猪气管上皮细胞（STECs）具有低侵袭潜力的突变体，经鉴定该突变体为自溶素（atl）。与野生型（WT）SS 2相比，Δatl突变体在小鼠模型中表现出较低的穿透气管上皮屏障的能力。纯化的Atl还增强了SS 2在Transwell插入物中跨STEC单层的易位。此外，Atl通过肌球蛋白轻链激酶（MLCK）信号转导重新分布STECs中的紧密连接（TJ），这导致屏障通透性增加。使用质谱、免疫共沉淀（co-IP）、下拉、细菌双杂交和饱和结合实验，我们表明Atl直接与波形蛋白结合。STECs（Vim KO STECs）中波形蛋白的CRISPR/Cas9靶向缺失消除了SS 2跨单层易位的能力、SS 2诱导的肌球蛋白II调节轻链（MLC）磷酸化和MLCK转录，表明波形蛋白对于MLCK活化是不可或缺的。一致地，波形蛋白敲除小鼠免受SS 2感染，并表现出减少的气管和肺损伤。因此，MLCK介导的上皮屏障开放所造成的β-波形蛋白的相互作用被认为是可能的关键机制，SS 2渗透气管上皮。猪链球菌2型（SS 2）是一种新发现的人畜共患病病原体，可破坏猪的呼吸道屏障，引起猪的侵袭性疾病。在这里，我们确定了自溶素Atl在SS 2穿透呼吸屏障及其全身传播中的新作用，并确定了其结合伴侣波形蛋白（一种III型中间丝蛋白）。Atl有助于MLCK触发的紧密连接的重新分布，以打开气管上皮屏障。波形蛋白的敲除消除了SS 2穿透单层屏障的能力和MLCK的活化。此外，鼻内施用SS 2保护波形蛋白缺失小鼠免受感染。本研究首次证明Atl的GBS Bsp样结构域与波形蛋白之间的相互作用促进MLCK介导的上皮屏障功能障碍，这可能为预防和/或治疗由类似呼吸道病原体引起的疾病提供理论信息。
Streptococcus suis serotype 2 (SS2) is a major zoonotic pathogen resulting in manifestations as pneumonia and septic shock. The upper respiratory tract is typically thought to be the main colonization and entry site of SS2 in pigs, but the mechanism through which it penetrates the respiratory barrier is still unclear. In this study, a mutant with low invasive potential to swine tracheal epithelial cells (STECs) was screened from the TnYLB-1 transposon insertion mutant library of SS2, and the interrupted gene was identified as autolysin (atl). Compared to wild-type (WT) SS2, Δatl mutant exhibited lower ability to penetrate the tracheal epithelial barrier in a mouse model. Purified Atl also enhanced SS2 translocation across STEC monolayers in Transwell inserts. Furthermore, Atl redistributed the tight junctions (TJs) in STECs through myosin light chain kinase (MLCK) signaling, which led to increased barrier permeability. Using mass spectrometry, co-immunoprecipitation (co-IP), pull-down, bacterial two-hybrid and saturation binding experiments, we showed that Atl binds directly to vimentin. CRISPR/Cas9-targeted deletion of vimentin in STECs (VIM KO STECs) abrogated the capacity of SS2 to translocate across the monolayers, SS2-induced phosphorylation of myosin II regulatory light chain (MLC) and MLCK transcription, indicating that vimentin is indispensable for MLCK activation. Consistently, vimentin null mice were protected from SS2 infection and exhibited reduced tracheal and lung injury. Thus, MLCK-mediated epithelial barrier opening caused by the Atl-vimentin interaction is found to be likely the key mechanism by which SS2 penetrates the tracheal epithelium. Streptococcus suis serotype 2 (SS2), an emerging zoonotic agent, can breach the respiratory barrier and cause invasive disease in pigs. Here, we identified the novel role of autolysin Atl in penetration of the respiratory barrier by SS2 and its systemic dissemination and identified its binding partner, vimentin, a type III intermediate filament protein. Atl contributed to the MLCK-triggered redistribution of tight junctions to open the tracheal epithelial barrier. Knockout of vimentin abolished the ability of SS2 to penetrate the monolayer barrier and the activation of MLCK. Furthermore, vimentin null mice were protected from infection by intranasally administered SS2. This study is the first to demonstrate that the interaction between the GBS Bsp-like domain of Atl and vimentin promotes MLCK-mediated dysfunction of the epithelial barrier, which may provide theoretical information for prophylactic and/or therapeutic treatments against diseases caused by similar respiratory pathogens.
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