Cyclic Di-GMP-Mediated Repression of Swarming Motility by Pseudomonas aeruginosa PA14 Requires the MotAB Stator

The second messenger cyclic diguanylate (c-di-GMP) plays a critical role in the regulation of motility. In Pseudomonas aerugi-nosa PA14, c-di-GMP inversely controls biofilm formation and surface swarmingmotility, with high levels of this dinucleotide signal stimulating biofilm formation and repressing swarming. P. aeruginosa encodes two stator complexes, MotAB and MotCD, that participate in the function of its single polar flagellum. Here we show that the repression of swarmingmotility re-quires a functional MotAB stator complex. Mutating themotAB genes restores swarming motility to a strain with artificially ele-vated levels of c-di-GMP as well as stimulates swarming in the wild-type strain, while overexpression of MotA from a plasmid represses swarming motility. Using point mutations inMotA and the FliG rotor protein of the motor supports the conclusion that MotA-FliG interactions are critical for c-di-GMP-mediated swarming inhibition. Finally, we show that high c-di-GMP levels affect the localization of a green fluorescent protein (GFP)-MotD fusion, indicating a mechanism whereby this secondmessenger has an impact onMotCD function.We propose that when c-di-GMP level is high, the MotAB stator can displace MotCD from the motor, thereby affecting motor function. Our data suggest a newly identified means of c-di-GMP-mediated control of surface motility, perhaps conserved among Pseudomonas, Xanthomonas, and other organisms that encode two stator systems. Since its discovery in 1987 as an allosteric activator of bacterialcellulose synthesis (1), cyclic diguanylate (c-di-GMP)has been shown to be a remarkably important signaling molecule across diverse bacterial species, controlling a multitude of behaviors an