Abstract
The marine bacterium Vibrio fischeri uses a biofilm to promote colonization of its eukaryotic host Euprymna scolopes. This biofilm depends on the symbiosis polysaccharide (syp) locus, which is transcriptionally regulated by the RscS-SypG two-component regulatory system. An additional response regulator (RR), SypE, exerts both positive and negative control over biofilm formation. SypE is a novel RR protein, with its three putative domains arranged in a unique configuration: a central phosphorylation receiver (REC) domain flanked by two effector domains with putative enzymatic activities (serine kinase and serine phosphatase). To determine how SypE regulates biofilm formation and host colonization, we generated a library of SypE domain mutants. Our results indicate that the N-terminus inhibits biofilm formation, while the C-terminus plays a positive role. The phosphorylation state of SypE appears to regulate these opposing activities, as disruption of the putative site of phosphorylation results in a protein that constitutively inhibits biofilm formation. Furthermore, SypE restricts host colonization: (i) sypE mutants with constitutive inhibitory activity fail to efficiently initiate host colonization and (ii) loss of sypE partially alleviates the colonization defect of an rscS mutant. We conclude that SypE must be inactivated to promote symbiotic colonization by V. fischeri.