C-di-GMP Regulates Motile to Sessile Transition by Modulating MshA Pili Biogenesis and Near-Surface Motility Behavior in Vibrio cholerae

In many bacteria, including Vibrio cholerae, cyclic dimeric guanosine monophosphate (c-di-GMP) controls the motile to biofilm life style switch. Yet, little is known about how this occurs. In this study, we report that changes in c-di-GMP concentration impact the biosynthesis of the MshA pili, resulting in altered motility and biofilm phenotypes in V. cholerae. Previously, we reported that cdgJ encodes a c-di-GMP phosphodiesterase and a ΔcdgJ mutant has reduced motility and enhanced biofilm formation. Here we show that loss of the genes required for the mannose-sensitive hemagglutinin (MshA) pilus biogenesis restores motility in the ΔcdgJ mutant. Mutations of the predicted ATPase proteins mshE or pilT, responsible for polymerizing and depolymerizing MshA pili, impair near surface motility behavior and initial surface attachment dynamics. A ΔcdgJ mutant has enhanced surface attachment, while the ΔcdgJmshA mutant phenocopies the high motility and low attachment phenotypes observed in a ΔmshA strain. Elevated concentrations of c-di-GMP enhance surface MshA pilus production. MshE, but not PilT binds c-di-GMP directly, establishing a mechanism for c-di-GMP signaling input in MshA pilus production. Collectively, our results suggest that the dynamic nature of the MshA pilus established by the assembly and disassembly of pilin subunits is essential for transition from the motile to sessile lifestyle and that c-di-GMP affects MshA pilus assembly and function through direct interactions with the MshE ATPase.在包括霍乱弧菌（Vibrio cholerae）在内的许多细菌中，环状双鸟苷单磷酸（cyclic dimeric guanosine monophosphate, c-di-GMP）调控着运动型到生物膜型的生活方式转换。然而，人们对这一过程的具体机制仍知之甚少。本研究发现，c-di-GMP浓度的变化会影响MshA菌毛的生物合成，进而改变霍乱弧菌的运动能力与生物膜表型。此前我们曾报道，cdgJ基因编码一种c-di-GMP磷酸二酯酶，ΔcdgJ突变体的运动能力下降且生物膜形成增强。本研究进一步证实，缺失甘露糖敏感血凝素（mannose-sensitive hemagglutinin, MshA）菌毛生物合成所需基因，可恢复ΔcdgJ突变体的运动能力。负责聚合与解聚MshA菌毛的预测ATP酶蛋白mshE或pilT发生突变后，会损害细菌的近表面运动行为与初始表面附着动态。ΔcdgJ突变体的表面附着能力增强，而ΔcdgJmshA双突变体的表型与ΔmshA菌株观察到的高运动性、低附着表型一致。高浓度的c-di-GMP会促进表面MshA菌毛的产生。MshE可直接结合c-di-GMP，而PilT则不能，这为c-di-GMP信号通路调控MshA菌毛生成提供了机制依据。综上，本研究结果表明，由菌毛亚基组装与解聚所确立的MshA菌毛动态变化，对于细菌从运动型到固着型生活方式的转变至关重要；同时c-di-GMP可通过直接与MshE ATP酶相互作用，影响MshA菌毛的组装与功能。