Scaffold proteins are ubiquitous chaperones that bind to proteins and facilitate the physical interaction of the components of signal transduction pathways or multi-enzymic complexes. In this study, we used a biochemical approach to dissect the molecular mechanism of a membrane-associated scaffold protein, FloT, a flotillin-homologue protein that is localized in functional membrane microdomains of the bacterium Bacillus subtilis. This study provides unambiguous evidence that FloT physically binds to and interacts with the membrane-bound sensor kinase KinC. This sensor kinase activates biofilm formation in B. subtilis in response to the presence of the self-produced signal surfactin. Furthermore, we have characterized the mechanism by which the interaction of FloT with KinC benefits the activity of KinC. Two separate and synergistic effects constitute this mechanism: first, the scaffold activity of FloT promotes more efficient self-interaction of KinC and facilitates dimerization into its active form. Second, the selective binding of FloT to KinC prevents the occurrence of unspecific aggregation between KinC and other proteins that may generate dead-end intermediates that could titrate the activity of KinC. Flotillin proteins appear to play an important role in prokaryotes in promoting effective binding of signalling proteins with their correct protein partners.
RsmA is a post-transcriptional RNA-binding protein that acts as a pleiotropic global regulator of mRNAs in the opportunistic pathogen Pseudomonas aeruginosa. Upon binding to its target, RsmA impedes the translation of the mRNA by the ribosome. The RsmA regulon affects over 500 genes, many of which have been identified as important in the pathogenicity of P. aeruginosa. Whilst the regulatory function of RsmA is relatively well characterized, the genetic regulation of rsmA itself at the transcriptional and translational levels remains poorly understood. Here, we show that RsmA is capable of self-regulation through an unorthodox mechanism. This regulation occurs via direct interaction of the protein with an RsmA-binding site located in the early portion of its coding sequence. To the best of our knowledge this is the first report of such an unusual regulation in pseudomonads.