Proteomic Profiling Reveals RisR as a Novel Regulatory Element in the Virulence Network of Bordetella pertussis

Bordetella pertussis, the causative agent of whooping cough, regulates its virulence through the BvgAS two-component system, which controls the transition between virulent (Bvg+) and avirulent (Bvg–) phases. In the virulent phase, virulence-activated genes (vags) are expressed, while in the avirulent phase, these are repressed. Virulence-repressed genes (vrgs) are induced under the control of the response regulator RisA. Phosphorylation of RisA by the nonoperonic kinase RisK is essential for vrgs expression. Adjacent to risK lies risR, a gene encoding a putative response regulator, whose function remained unexplored. To investigate the RisR’s role in B. pertussis biology, we performed comparative proteomic analyses between wild-type and an isogenic risR-deficient strain, grown under both virulent and avirulent conditions. Our data show that RisR modulates the abundance of proteins encoded by vags and vrgs, suggesting a previously unrecognized interplay between RisR and the BvgAS–RisAK regulatory network. Moreover, RisR regulates the abundance of proteins important for bacterial fitness, including those involved in iron acquisition and intracellular adaptation. Functional assays further support the role of RisR in promoting intracellular survival. This study provides the first proteome-wide characterization of the RisR regulon and identifies it as a novel regulator of B. pertussis virulence and adaptation.

百日咳博德特菌（Bordetella pertussis）是引发百日咳的致病菌，其通过BvgAS双组分系统（two-component system）调控毒力，该系统可控制致病菌在毒力相（Bvg+）与非毒力相（Bvg–）之间的表型转换。在毒力相状态下，毒力激活基因（virulence-activated genes，vags）得以表达；而在非毒力相状态下，这类基因会被抑制。毒力抑制基因（virulence-repressed genes，vrgs）的表达受应答调控因子（response regulator）RisA的诱导。由非操纵子激酶（nonoperonic kinase）RisK对RisA进行磷酸化，是vrgs表达的必要条件。risK的相邻基因为risR，其编码一种推定的应答调控因子，此前其功能尚未被探索。为探究RisR在百日咳博德特菌生物学中的作用，我们分别在毒力相和非毒力相培养条件下，对野生型菌株与同基因risR缺陷菌株开展了比较蛋白质组学分析。研究数据显示，RisR可调控vags与vrgs所编码蛋白的丰度，这表明RisR与BvgAS–RisAK调控网络之间存在此前未被认知的相互作用。此外，RisR还可调控对细菌适应性至关重要的蛋白丰度，其中包括参与铁获取与胞内适应过程的蛋白。功能实验进一步证实了RisR在促进细菌胞内存活中的作用。本研究首次完成了RisR调控子（regulon）的全蛋白质组表征，并将其鉴定为百日咳博德特菌毒力与适应性的新型调控因子。