Two-component signal transduction systems (TCSs) represent a major mechanism that bacteria use to sense and respond to their environment. Prototypical TCSs are composed of a membrane-embedded histidine kinase, which senses an environmental stimulus and subsequently phosphorylates a cognate partner protein called a response regulator that regulates gene expression in a phosphorylation-dependent manner. Vibrio cholerae uses the hybrid histidine kinase ChiS to activate the expression of the chitin utilization program, which is critical for the survival of this facultative pathogen in its aquatic reservoir. A cognate response regulator for ChiS has not been identified and the mechanism of ChiS-dependent signal transduction remains unclear. Here, we show that ChiS is a noncanonical membrane-embedded one-component system that can both sense chitin and directly regulate gene expression via a cryptic DNA binding domain. Unlike prototypical TCSs, we find that ChiS DNA binding is diminished, rather than stimulated, by phosphorylation. Finally, we provide evidence that ChiS likely activates gene expression by directly recruiting RNA polymerase. This work addresses the mechanism of action for a major transcription factor in V. cholerae and highlights the versatility of signal transduction systems in bacterial species.

Two-component signal transduction systems (TCSs) represent a major mechanism that bacteria use to sense and respond to their environment. Prototypical TCSs are composed of a membrane-embedded histidine kinase (HK), which senses an environmental stimulus and subsequently phosphorylates a cognate partner protein called a response regulator (RR) that regulates gene expression in a phosphorylationdependent manner. Vibrio cholerae uses the hybrid HK ChiS to activate the expression of the chitin utilization program, which is critical for the survival of this facultative pathogen in its aquatic reservoir. A cognate RR for ChiS has not been identified and the mechanism of ChiS-dependent signal transduction remains unclear. Here, we show that ChiS is a noncanonical membrane-embedded one-component system that can both sense chitin and directly regulate gene expression via a cryptic DNA binding domain. Unlike prototypical TCSs, we find that ChiS DNA binding is diminished, rather than stimulated, by phosphorylation. Finally, we provide evidence that ChiS likely activates gene expression by directly recruiting RNA polymerase. Together, this work addresses the mechanism of action for a major transcription factor in V. cholerae and highlights the versatility of signal transduction systems in bacterial species. Significance StatementFrom bacteria to humans, the ability to properly respond to environmental cues is critical for survival. The cholera pathogen Vibrio cholerae uses one protein, ChiS, to sense chitin in its environmental reservoir to regulate the expression of genes that are critical for the survival and evolution of this pathogen in this niche. Here, we study how the chitin sensor ChiS works, and discover that it regulates gene expression in an unexpected and unorthodox manner. Thus, this study uncovers how the major regulator ChiS works in this important human pathogen and highlights the versatile mechanisms that living systems use to respond to their environment.

Bacteria use extracellular appendages called type IV pili (T4P) for diverse behaviors including DNA uptake, surface sensing, virulence, protein secretion, and twitching motility. Dynamic extension and retraction of T4P is essential for their function, and T4P extension is thought to occur through the action of a single, highly conserved motor, PilB. Here, we develop Acinetobacter baylyi as a model to study T4P by employing a recently developed pilus labeling method. By contrast to previous studies of other bacterial species, we find that T4P synthesis in A. baylyi is dependent not only on PilB but also on an additional, phylogenetically distinct motor, TfpB. Furthermore, we identify a protein (CpiA) that inhibits T4P extension by specifically binding and inhibiting PilB but not TfpB. These results expand our understanding of T4P regulation and highlight how inhibitors might be exploited to disrupt T4P synthesis.

Chitin utilization by the cholera pathogen Vibrio cholerae is required for its persistence and evolution via horizontal gene transfer in the marine environment. Genes involved in the uptake and catabolism of the chitin disaccharide chitobiose are encoded by the chb operon. The orphan sensor kinase ChiS is critical for regulation of this locus, however, the mechanisms downstream of ChiS activation that result in expression of the chb operon are poorly understood. Using an unbiased transposon mutant screen, we uncover that the nucleoid occlusion protein SlmA is a regulator of the chb operon. SlmA has not previously been implicated in gene regulation. Also, SlmA is a member of the TetR family of proteins, which are generally transcriptional repressors. In vitro, we find that SlmA binds directly to the chb operon promoter, and in vivo, we show that this interaction is, surprisingly, required for transcriptional activation of this locus and for chitobiose utilization. Using point mutations that disrupt distinct functions of SlmA, we find that DNA-binding, but not nucleoid occlusion, is critical for transcriptional activation. This study identifies a novel role for SlmA as a transcriptional regulator in V. cholerae in addition to its established role as a cell division licensing factor.AUTHOR SUMMARYThe cholera pathogen Vibrio cholerae is a natural resident of the aquatic environment and causes disease when ingested in the form of contaminated food or drinking water. In the aquatic environment, the shells of marine zooplankton, which are primarily composed of chitin, serve as an important food source for this pathogen. The genes required for the utilization of chitin are tightly regulated in V. cholerae, however, the exact mechanism underlying this regulation is currently unclear. Here, we uncover that a protein involved in regulating cell division is also important for regulating the genes involved in chitin utilization. This is a newly identified property for this cell division protein and the significance of a common regulator for these two disparate activities remains to be understood.

Chitin utilization by the cholera pathogen Vibrio cholerae is required for its persistence and evolution via horizontal gene transfer in the marine environment. Genes involved in the uptake and catabolism of the chitin disaccharide chitobiose are encoded by the chb operon. The orphan sensor kinase ChiS is critical for regulation of this locus, however, the mechanisms downstream of ChiS activation that result in expression of the chb operon are poorly understood. Using an unbiased transposon mutant screen, we uncover that the nucleoid occlusion protein SlmA is a regulator of the chb operon. SlmA has not previously been implicated in gene regulation. Also, SlmA is a member of the TetR family of proteins, which are generally transcriptional repressors. In vitro, we find that SlmA binds directly to the chb operon promoter, and in vivo, we show that this interaction is required for transcriptional activation of this locus and for chitobiose utilization. Using point mutations that disrupt distinct functions of SlmA, we find that DNA-binding, but not nucleoid occlusion, is critical for transcriptional activation. This study identifies a novel role for SlmA as a transcriptional regulator in V. cholerae in addition to its established role as a cell division licensing factor.