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

Abstract

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.

The chemoreceptor controlling the Wsp-like transduction pathway inHalomonas titanicaeKHS3 binds purine derivatives

“…The low identity value between the periplasmic domains of these two receptors was expected, as their LBDs do not share folding type. However, big differences between their cytoplasmic domains are rather surprising taking into account that Ht Che2 pathway seems to be functionally equivalent to the Wsp [4]. The fact that some other components show important differences between pathways (for example regarding coupling proteins and domains in the associated diguanylated cyclase) highlights the need of more studies to characterize the signalling behavior of the proteins involved in these cases and also in related pathways in other microorganisms.…”

“…The main physiological role of the Ht Che2 transduction pathway could be still considered an open question. Even though the removal of the cognate methyltransferase gene and the consequent increase in receptor methylation generates an hyperbiofilm phenotype that is comparable to the equivalent mutant in the Pseudomonas Wsp pathway [4], it is likely that this only reflects an increase in the activity of the controlled diguanylate cyclase and not necessarily reveals the main function of the pathway. The strong conservation of the Htc10-like cytoplasmic domain, its divergence compared to WspA-like receptors as well as additional differences in gene and/or domain composition of the predicted signalling components, leave open the question about how related both pathways are in terms of signalling.…”

“…HtChe2 pathway seems to be functionally equivalent to the Wsp [4]. The fact that some other components show important differences between pathways (for example regarding coupling proteins and domains in the associated diguanylated cyclase) highlights the need of more studies to characterize the signalling behavior of the proteins involved in these cases and also in related pathways in other microorganisms.…”

“…Its genome sequence revealed the presence of two chemotaxis-related gene clusters [3]. Cluster Che1 ( Ht Che1) resembles the Che cluster from Escherichia coli and controls the chemotaxis behaviour [4]. Cluster Che2 ( Ht Che2), instead, contains a diguanylate cyclase gene encoding a protein with N-terminal phosphorylatable domains ( Ht DGC, locus RO22_21180), resembling the Wsp pathway of Pseudomonas [3] (see below).…”

“…Cluster Che2 ( Ht Che2), instead, contains a diguanylate cyclase gene encoding a protein with N-terminal phosphorylatable domains ( Ht DGC, locus RO22_21180), resembling the Wsp pathway of Pseudomonas [3] (see below). Moreover, gene cluster Ht Che2 seems to be functionally equivalent to the Wsp cluster as a deletion mutant in the methylesterase gene cheB2 , predicted to cause a constitutive activation of the pathway, resulted in a wrinkly colony phenotype and in increased biofilm formation ability [4].…”