Legionella pneumophila is an intracellular pathogen found in aquatic environments where it replicates in protozoan hosts. My objectives were to identify molecular mechanisms that control virulence and multidrug resistance in L. pneumophila, and especially to explore the role of proteins “GGDEF/EAL” in virulence. GGDEF and EAL domains are found in enzymes able to synthesize (diguanylate cyclase, DGC) or degrade (phosphodiesterase, PDE) c-di-GMP, respectively. C-di-GMP is a bacterial second messenger which plays a key role in regulating main functions including motility, and virulence. L. pneumophila Lens contains 22 genes encoding GGDEF/EAL proteins and most of them are expressed simultaneously with genes encoding virulence factors. The enzymatic activities of the 22 GGDEF/EAL proteins of L. pneumophila Lens were assayed in vitro. Among the 10 proteins purified, 6 showed a DGC activity and 2 contained both activities. The role of the GGDEF/EAL proteins of L. pneumophila Lens on virulence was investigated. Inactivation of 5 genes and overexpression of 2 other genes led to a significant decrease in virulence. Moreover, DGC activity of at least two of these proteins is required for bacterial virulence. Finally, an original two-component system was identified comprising Lpl0330, a histidine kinase able to autophosphorylate on a new HisKA domain, and Lpl 0329, a protein with dual in vitro DGC/PDE activity. Phosphorylation of Lpl0329 led to a decrease in its DGC activity only, giving the first example of a bifunctional enzyme which modulates synthesis and turnover of c-di-GMP in response to phosphorylation (Levet-Paulo et al., 2011).
