ABSTRACTGlucose-specific enzyme IIA (EIIAGlc) is a central regulator of bacterial metabolism and an intermediate in the phosphoenolpyruvate phosphotransferase system (PTS), a conserved phosphotransfer cascade that controls carbohydrate transport. We previously reported that EIIAGlcactivates transcription of the genes required forVibrio choleraebiofilm formation. While EIIAGlcmodulates the function of many proteins through a direct interaction, none of the known regulatory binding partners of EIIAGlcactivates biofilm formation. Therefore, we used tandem affinity purification (TAP) to compare binding partners of EIIAGlcin both planktonic and biofilm cells. A surprising number of novel EIIAGlcbinding partners were identified predominantly under one condition or the other. Studies of planktonic cells revealed established partners of EIIAGlc, such as adenylate cyclase and glycerol kinase. In biofilms, MshH, a homolog ofEscherichia coliCsrD, was found to be a dominant binding partner of EIIAGlc. Further studies revealed that MshH inhibits biofilm formation. This function was independent of the Carbon storage regulator (Csr) pathway and dependent on EIIAGlc. To explore the existence of multiprotein complexes centered on EIIAGlc, we also affinity purified the binding partners of adenylate cyclase from biofilm cells. In addition to EIIAGlc, this analysis yielded many of the same proteins that copurified with EIIAGlc. We hypothesize that EIIAGlcserves as a hub for multiprotein complexes and furthermore that these complexes may provide a mechanism for competitive and cooperative interactions between binding partners.IMPORTANCEEIIAGlcis a global regulator of microbial physiology that acts through direct interactions with other proteins. This work represents the first demonstration that the protein partners of EIIAGlcare distinct in the microbial biofilm. Furthermore, it provides the first evidence that EIIAGlcmay exist in multiprotein complexes with its partners, setting the stage for an investigation of how the multiple partners of EIIAGlcinfluence one another. Last, it provides a connection between the phosphoenolpyruvate phosphotransferase (PTS) and Csr (Carbon storage regulator) regulatory systems. This work increases our understanding of the complexity of regulation by EIIAGlcand provides a link between the PTS and Csr networks, two global regulatory cascades that influence microbial physiology.
Correction: Engineering microbial physiology with synthetic polymers: cationic polymers induce biofilm formation inVibrio choleraeand downregulate the expression of virulence genes
