(A) Overview of LapD-mediated signaling. The primary structure and domain organization of the LapD receptor is shown (top panel). The cartoon (bottom panel) summarizes the current model of LapD-mediated regulation of biofilm formation in P. fluorescens by the differential recruitment of the periplasmic protease LapG in response to exogenous inorganic phosphate (Pi) availability (reviewed in Boyd and O'Toole, 2012). Low bioavailability of Pi induces the expression of phosphodiesterases (PDEs) via the Pho system, which results in low intra-cellular concentrations of c-di-GMP and LapD adopting an autoinhibited state. Auto-inhibited LapD cannot sequester the periplasmic protease LapG, allowing the LapG enzyme to cleave the cell-surface adhesin LapA, releasing it from the cell surface. Conversely, when environmental Pi is present in sufficient concentration, a subset of diguanylate cyclases (DGCs) produce intra-cellular c-di-GMP to levels sufficient for binding to the EAL domain of LapD, thus activating this receptor. Activated LapD binds to and sequesters LapG in the periplasm, which promotes biofilm formation by preventing cleavage of LapA. (B) Crystal structures of the periplasmic output domain from P. fluorescens LapD and L. pneumophila CdgS9. Two orthogonal views of each output domain structure are shown in ribbon representation, with the two protomers colored pink and gray, respectively (P. fluorescens LapDOutput, PDB 3PJV; Navarro et al., 2011). Distances between the tryptophan residues in the conserved GWxQ motif of each protomer are indicated. The relative position of the inner cell membrane (gray bar) and connection to the flanking transmembrane (TM) helices are indicated. (C) Surface conservation of the output domain. The sequences of 18 orthologs of LapD were aligned and the sequence conservation was mapped on to the solvent-accessible surface of CdgS9Output. The surface is colored according to the degree of conservation.