A c-di-GMP Signaling Cascade Controls Motility, Biofilm Formation, and Virulence in Burkholderia thailandensis.

As a key bacterial second messenger, cyclic di-GMP (c-di-GMP) regulates various physiological processes, such as motility, biofilm formation, and virulence. Cellular c-di-GMP levels are regulated by the opposing activities of diguanylate cyclases (DGCs) and phosphodiesterases (PDEs). Beyond that, the enzymatic activities of c-di-GMP metabolizing proteins are controlled by a variety of extracellular signals and intracellular physiological conditions. Here, we report that pdcA (BTH_II2363), pdcB (BTH_II2364), and pdcC (BTH_II2365) are cotranscribed in the same operon and are involved in a regulatory cascade controlling the cellular level of c-di-GMP in Burkholderia thailandensis. The GGDEF domain-containing protein PdcA was found to be a DGC that modulates biofilm formation, motility, and virulence in B. thailandensis. Moreover, the DGC activity of PdcA was inhibited by phosphorylated PdcC, a single-domain response regulator composed of only the phosphoryl-accepting REC domain. The phosphatase PdcB affects the function of PdcA by dephosphorylating PdcC. The observation that homologous operons of pdcABC are widespread among betaproteobacteria and gammaproteobacteria suggests a general mechanism by which the intracellular concentration of c-di-GMP is modulated to coordinate bacterial behavior and virulence. IMPORTANCE The transition from planktonic cells to biofilm cells is a successful strategy adopted by bacteria to survive in diverse environments, while the second messenger c-di-GMP plays an important role in this process. Cellular c-di-GMP levels are mainly controlled by modulating the activity of c-di-GMP-metabolizing proteins via the sensory domains adjacent to their enzymatic domains. However, in most cases how c-di-GMP-metabolizing enzymes are modulated by their sensory domains remains unclear. Here, we reveal a new c-di-GMP signaling cascade that regulates motility, biofilm formation, and virulence in B. thailandensis. While pdcA, pdcB, and pdcC constitute an operon, the phosphorylated PdcC binds the PAS sensory domain of PdcA to inhibit its DGC activity, with PdcB dephosphorylating PdcC to derepress the activity of PdcA. We also show this c-di-GMP regulatory model is widespread in the phylum Proteobacteria. Our study expands the current knowledge of how bacteria regulate intracellular c-di-GMP levels.

Sensing of autoinducer-2 by functionally distinct receptors in prokaryotes.

Autoinducer-2 (AI-2) is a quorum sensing signal that mediates communication within and between many bacterial species. However, its known receptors (LuxP and LsrB families) are not found in all the bacteria capable of responding to this signaling molecule. Here, we identify a third type of AI-2 receptor, consisting of a dCACHE domain. AI-2 binds to the dCACHE domain of chemoreceptors PctA and TlpQ of Pseudomonas aeruginosa, thus inducing chemotaxis and biofilm formation. Boron-free AI-2 is the preferred ligand for PctA and TlpQ. AI-2 also binds to the dCACHE domains of histidine kinase KinD from Bacillus subtilis and diguanylate cyclase rpHK1S-Z16 from Rhodopseudomonas palustris, enhancing their enzymatic activities. dCACHE domains (especially those belonging to a subfamily that includes the AI-2 receptors identified in the present work) are present in a large number of bacterial and archaeal proteins. Our results support the idea that AI-2 serves as a widely used signaling molecule in the coordination of cell behavior among prokaryotic species.