Department of Oral Diagnosis, Piracicaba Dental School, State University of Campinas, Piracicaba, SP, Brazil.
Department of Oral Biology, University of Florida College of Dentistry, Gainesville, Florida, USA.
J Bacteriol. 2018 Nov 6;200(23). doi: 10.1128/JB.00141-18. Print 2018 Dec 1.
Cnm is a surface-associated protein present in a subset of strains that mediates binding to extracellular matrices, intracellular invasion, and virulence. Here, we showed that transcription is controlled by the global regulators CovR and VicRKX. analysis identified multiple putative CovR- and VicR-binding motifs in the regulatory region of as well as in the downstream gene , which is associated with the posttranslational modification of Cnm. Electrophoretic mobility shift assays revealed that CovR and VicR specifically and independently bind to the and promoter regions. Quantitative real-time PCR and Western blot analyses of Δ and Δ strains as well as of a strain overexpressing revealed that CovR functions as a positive regulator of , whereas VicRKX acts as a negative regulator. In agreement with the role of VicRKX as a repressor, the Δ strain showed enhanced binding to collagen and laminin and higher intracellular invasion rates. Overexpression of was associated with decreased rates of intracellular invasion but did not affect collagen or lamin binding activities, suggesting that this system controls additional genes involved in binding to these extracellular matrix proteins. As expected, based on the role of CovR in regulation, the Δ strain showed decreased intracellular invasion rates, but, unexpectedly collagen and laminin binding activities were increased in this mutant strain. Collectively, the results presented here expand the repertoire of virulence-related genes regulated by CovR and VicRKX to include the core gene and the noncore gene is a major pathogen associated with dental caries and also implicated in systemic infections, in particular, infective endocarditis. The Cnm adhesin of is an important virulence factor associated with systemic infections and caries severity. Despite its role in virulence, the regulatory mechanisms governing expression are poorly understood. Here, we describe the identification of two independent regulatory systems controlling the transcription of and the downstream operon. A better understanding of the mechanisms controlling expression of virulence factors like Cnm can facilitate the development of new strategies to treat bacterial infections.
Cnm 是一种存在于某些 菌株中的表面相关蛋白，介导与细胞外基质的结合、细胞内入侵和毒力。在这里，我们表明 转录受全局调控因子 CovR 和 VicRKX 控制。 分析鉴定了 调控区和下游基因 中的多个推定 CovR 和 VicR 结合基序，该基因与 Cnm 的翻译后修饰有关。电泳迁移率变动分析显示 CovR 和 VicR 特异性和独立地结合到 和 启动子区域。Δ 和 Δ 菌株以及过表达 的菌株的定量实时 PCR 和 Western blot 分析表明，CovR 作为 的正调控因子发挥作用，而 VicRKX 作为负调控因子发挥作用。与 VicRKX 作为抑制剂的作用一致，Δ 菌株显示出与胶原蛋白和层粘连蛋白更强的结合和更高的细胞内入侵率。 的过表达与细胞内入侵率降低相关，但不影响胶原蛋白或层粘连蛋白结合活性，表明该系统控制与这些细胞外基质蛋白结合的其他基因。不出所料，基于 CovR 在 调节中的作用，Δ 菌株显示出降低的细胞内入侵率，但令人惊讶的是，在该突变菌株中胶原蛋白和层粘连蛋白结合活性增加。总的来说，这里呈现的结果扩展了 CovR 和 VicRKX 调控的与毒力相关基因的范围，包括核心基因 和非核心基因 是一种与龋齿密切相关的主要病原体，也与全身感染有关，特别是感染性心内膜炎。 的 Cnm 黏附素是与全身感染和龋齿严重程度相关的重要毒力因子。尽管它在毒力中起作用，但控制 表达的调节机制知之甚少。在这里，我们描述了两个独立的调控系统的鉴定，这些系统控制着 和下游 操纵子的转录。更好地了解控制像 Cnm 这样的毒力因子表达的机制可以促进治疗细菌感染的新策略的发展。
