Chemosensory pathways of Halomonas titanicae KHS3 control chemotaxis behaviour and biofilm formation.

Halomonas titanicae KHS3 is a marine bacterium whose genome codes for two different chemosensory pathways. Chemosensory gene cluster 1 is very similar to the canonical Che cluster from Escherichia coli. Chemosensory cluster 2 includes a gene coding for a diguanylate cyclase with receiver domains, suggesting that it belongs to the functional group that regulates alternative cellular functions other than chemotaxis. In this work we assess the functional roles of both chemosensory pathways through approaches that include the heterologous expression of Halomonas proteins in E. coli strains and phenotypic analyses of Halomonas mutants. Our results confirm that chemosensory cluster 1 is indeed involved in chemotaxis behaviour, and only proteins from this cluster complement E. coli defects. We present evidence suggesting that chemosensory cluster 2 resembles the Wsp pathway from Pseudomonas, since the corresponding methylesterase mutant shows an increased methylation level of the cognate receptor and develops a wrinkly colony morphology correlated with an increased ability to form biofilm. Consistently, mutational interruption of this gene cluster correlates with low levels of biofilm. Our results suggest that the proteins from each pathway assemble and function independently. However, the phenotypic characteristics of the mutants show functional connections between the pathways controlled by each chemosensory system.

Direct binding of benzoate derivatives to two chemoreceptors with Cache sensor domains in Halomonas titanicae KHS3.

Halomonas titanicae KHS3, isolated from a hydrocarbon-contaminated sea harbor in Argentina, is able to grow on aromatic hydrocarbons and displays chemotaxis toward those compounds. This behavior might contribute to the efficiency of its degradation capacity. Using high throughput screening, we identified two chemoreceptors (Htc1 and Htc2) that bind benzoate derivatives and other organic acids. Whereas Htc1 has a high affinity for benzoate (Kd 112 µM) and 2-hydroxybenzoate (Kd 83 µM), Htc2 binds 2-hydroxybenzoate with low affinity (Kd 3.25 mM), and also C3/C4 dicarboxylates. Both chemoreceptors are able to trigger a chemotactic response of E. coli cells to the specific ligands. A H. titanicae htc1 mutant has reduced chemotaxis toward benzoate, and is complemented upon expression of the corresponding receptor. Both chemoreceptors have a Cache-type sensor domain, double (Htc1) or single (Htc2), and their ability to bind aromatic compounds is reported here for the first time.