The BgIF sensor recruits the BgIG transcription regulator to the membrane and releases it on stimulation

The Escherichia coli BgIF protein is a sugar-sensor that controls the activity of the transcriptional antiterminator BgIG by reversibly phosphorylating it, depending on β-glucoside availability. BgIF is a membrane-bound protein, whereas BgIG is a soluble protein, and they are both present in the cell in minute amounts. How do BgIF and BgIG find each other to initiate signal transduction efficiently? Using bacterial two-hybrid systems and the Far-Western technique, we demonstrated unequivocally that BgIG binds to BgIF and to its active site-containing domain in vivo and in vitro. Measurements by surface plasmon resonance corroborated that the affinity between these proteins is high enough to enable their stable binding. To visualize the subcellular localization of BgIG, we used fluorescence microscopy. In cells lacking BgIF, the BgIG-GFP fusion protein was evenly distributed throughout the cytoplasm. In contrast, in cells producing BgIF, BgIG-GFP was localized to the membrane. On addition of β-glucoside, BgIG-GFP was released from the membrane, becoming evenly distributed throughout the cell. Using mutant proteins and genetic backgrounds that impede phosphorylation of the BgI proteins, we demonstrated that BgIG-BgIF binding and recruitment of BgIG to the membrane sensor requires phosphorylation but does not depend on the individual phosphorylation sites of the Bgl proteins. We suggest a mechanism for rapid response to environmental changes by preassembly of signaling complexes, which contain transcription regulators recruited by their cognate sensors-kinases, under nonstimulating conditions, and release of the regulators to the cytoplasm on stimulation. This mechanism might be applicable to signaling cascades in prokaryotes and eukaryotes.