孢子形成是人类肠致病性艰难梭菌的环境生存和传播所必需的。在所有细菌孢子形成物中,孢子形成是通过激活主反应调节剂来调节的,Spo0A.然而,直接调节艰难梭菌Spo0A活性的因素和机制未定义。在经过充分研究的芽孢杆菌属物种中,spo0A被spo0E直接灭活,一个小的磷酸酶。要了解C.difficile中的Spo0E功能,我们创建了spo0E直系同源的无效突变,并评估了孢子形成和生理学.spo0E突变体产生明显更多的孢子,显示Spo0E抑制艰难梭菌孢子形成。出乎意料的是,spo0E突变体还表现出增加的运动性和毒素产生,并增强动物感染的毒力。我们发现spo0E与spo0A和毒素和运动调节剂相互作用,Rsta.Spo0A之间的直接相互作用,Spo0E,和RstA构成一个以前未知的分子开关,协调孢子形成的运动和毒素的产生。对B.subtilis的Spo0E功能的重新研究显示,Spo0E诱导的运动性,证明Spo0E调节不同物种之间的运动性和孢子形成。Further,Spo0E的3D结构分析揭示了Spo0E与艰难梭菌和枯草芽孢杆菌中的结合配偶体之间的特异性和排他性相互作用,其提供了对该调节机制在不同物种之间的保守性的洞察。
Spore formation is required for environmental survival and transmission of the human enteropathogenic Clostridioides difficile. In all bacterial spore formers, sporulation is regulated through activation of the master response regulator, Spo0A. However, the factors and mechanisms that directly regulate C. difficile Spo0A activity are not defined. In the well-studied Bacillus species, Spo0A is directly inactivated by Spo0E, a small phosphatase. To understand Spo0E function in C. difficile, we created a null mutation of the spo0E ortholog and assessed sporulation and physiology. The spo0E mutant produced significantly more spores, demonstrating Spo0E represses C. difficile sporulation. Unexpectedly, the spo0E mutant also exhibited increased motility and toxin production, and enhanced virulence in animal infections. We uncovered that Spo0E interacts with both Spo0A and the toxin and motility regulator, RstA. Direct interactions between Spo0A, Spo0E, and RstA constitute a previously unknown molecular switch that coordinates sporulation with motility and toxin production. Reinvestigation of Spo0E function in B. subtilis revealed that Spo0E induced motility, demonstrating Spo0E regulation of motility and sporulation among divergent species. Further, 3D structural analyses of Spo0E revealed specific and exclusive interactions between Spo0E and binding partners in C. difficile and B. subtilis that provide insight into the conservation of this regulatory mechanism among different species.