Department of NanoBiotechnology, NanoGlycobiology Unit, Universität für Bodenkultur Wien, Vienna, Austria.
Department of Chemistry, Institute of Biochemistry, Universität für Bodenkultur Wien, Vienna, Austria.
Front Cell Infect Microbiol. 2022 Feb 10;12:835509. doi: 10.3389/fcimb.2022.835509. eCollection 2022.
and target distinct virulence factors bearing a structurally conserved C-terminal domain (CTD) to the type IX protein secretion system (T9SS). The T9SS comprises an outer membrane translocation complex which works in concert with a signal peptidase for CTD cleavage. Among prominent T9SS cargo linked to periodontal diseases are the TfsA and TfsB components of cell surface (S-) layer, the bacterium's BspA surface antigen and a set of cysteine proteinases (gingipains) from . To assess the overall role of the bacterial T9SS in the host response, human macrophages and human gingival fibroblasts were stimulated with and wild-type bacteria and T9SS signal peptidase-deficient mutants defective in protein secretion, respectively. The immunostimulatory potential of these bacteria was compared by analyzing the mRNA expression levels of the pro-inflammatory mediators IL-6, IL-8, MCP-1 and TNF-α by qPCR and by measuring the production of the corresponding proteins by ELISA. Shot-gun proteomics analysis of and outer membrane preparations confirmed that several CTD-bearing virulence factors which interact with the human immune system were depleted from the signal peptidase mutants, supportive of effective T9SS shut-down. Three and, more profoundly, 16 hours post stimulation, the T9SS mutant induced significantly less production of cytokines and the chemokine in human cells compared to the corresponding parent strain, while the opposite was observed for the T9SS mutant. Our data indicate that T9SS shut-down translates into an altered inflammatory response in periodontal pathogens. Thus, the T9SS as a potential novel target for periodontal therapy needs further evaluation.
并靶向具有结构保守 C 端结构域 (CTD) 的独特毒力因子到九型蛋白分泌系统 (T9SS)。T9SS 包括一个外膜易位复合物，与信号肽酶协同作用进行 CTD 切割。与牙周病相关的主要 T9SS 货物包括细胞表面 (S-) 层的 TfsA 和 TfsB 成分、细菌的 BspA 表面抗原和一组半胱氨酸蛋白酶 (gingipains)。为了评估细菌 T9SS 在宿主反应中的整体作用，用 和 野生型细菌和 T9SS 信号肽酶缺陷突变体分别刺激人巨噬细胞和人牙龈成纤维细胞进行蛋白质分泌。通过 qPCR 分析促炎介质 IL-6、IL-8、MCP-1 和 TNF-α 的 mRNA 表达水平，并通过 ELISA 测量相应蛋白质的产生，比较这些细菌的免疫刺激潜力。和 外膜制剂的鸟枪法蛋白质组学分析证实，与人类免疫系统相互作用的几种携带 CTD 的毒力因子从信号肽酶突变体中耗尽，支持有效的 T9SS 关闭。在刺激后 3 小时和 16 小时，与相应的亲本菌株相比，T9SS 突变体在人类细胞中诱导产生的细胞因子和趋化因子明显减少，而 T9SS 突变体则相反。我们的数据表明，T9SS 关闭会导致牙周病原体的炎症反应改变。因此，T9SS 作为牙周治疗的潜在新靶点需要进一步评估。
