A Symphony of Cyclases: Specificity in Diguanylate Cyclase Signaling

Abstract

Cyclic diguanylate (c-di-GMP) is a near universal signaling molecule produced by diguanylate cyclases that can direct a variety of bacterial behaviors. A major area of research over the last several years has been aimed at understanding how a cell with dozens of diguanylate cyclases can deploy a given subset of them to produce a desired phenotypic outcome without undesired cross talk between c-di-GMP-dependent systems. Several models have been put forward to address this question, including specificity of cyclase activation, tuned binding constants of effector proteins, and physical interaction between cyclases and effectors. Additionally, recent evidence has suggested that there may be a link between the catalytic state of a cyclase and its physical contact with an effector. This review highlights several key studies, examines the proposed global and local models of c-di-GMP signaling specificity in bacteria, and attempts to identify the most fruitful steps that can be taken to better understand how dynamic networks of sibling cyclases and effector proteins result in sensible outputs that govern cellular behavior.

Keywords: biofilms; c-di-GMP; cyclic diguanylate; diguanylate cyclase; signaling specificity.

Figure 1

Modes of c-di-GMP signaling specificity. A variety of models have been proposed to explain the signaling specificity of a given DGC for a specific cellular process. (a) In a global model of signaling specificity, different DGCs produce varying amounts of c-di-GMP that contribute to a global pool, and PDEs can reduce the amount of the global pool. Effectors with the lowest dissociation constant are activated first. As the global pool of c-di-GMP increases, effectors with higher dissociation constants are activated. (b) Local models of c-di-GMP signaling can operate on several different principles. DGCs that are in proximity of—but not necessarily interacting with—effectors can create a local pool of c-di-GMP that may activate target proteins, whereas PDEs may be responsible for keeping the local c-di-GMP pool from affecting other effectors (left). Alternatively, direct interaction between DGCs, effectors, and PDEs can result in functional complexes. The state of the effector depends on the activation state of the DGC or PDE enzymes. Additionally, the DGC catalytic rate may be affected by physical interaction with its partners (right).

Figure 2

Components of GGDEF domain of DGCs. Crystal structure of the GcbC GGDEF domain. The amino acid backbone is shown as a ribbon and is overlaid on the electrostatic surface. So-called GGDEF domains (which can also have a GGEEF motif) catalyze the cyclization of two molecules of GTP into c-di-GMP using the catalytically active pocket (GGEEF residues shown in yellow, left). The GGDEF domain makes contact with its effector LapD using the N-terminal portion of the α5^GGDEF helix shown in cyan (left). The primary I-site, shown at right in purple, is used to quench catalytic activity when c-di-GMP binds to the I-site pocket. GcbC has an RRxxD motif as its primary I-site, and the R366 residue makes up the secondary I-site (left), which works in conjunction with the primary I-site to bind c-di-GMP.

Figure 3

The trigger enzyme system of Escherichia coli. The global c-di-GMP pool is partially controlled by the DGC YegE and the PDE YhjH. When the c-di-GMP pool is low, the trigger enzyme and PDE YciR bind the DGC YdaM and its target, the transcription factor MlrA (bold arrows), preventing YdaM activation of MlrA. When the c-di-GMP pool is high, YciR degrades c-di-GMP and releases YdaM and MlrA in the process, leading to YdaM activation of MlrA and further contribution to the global c-di-GMP pool (blue arrow). Adapted from Lindenberg et al. (38).

Figure 4

The Lap system of Pseudomonas fluorescens. (a) A simplified model of the activated Lap system. The large adhesin LapA is translocated through the type I secretion apparatus composed of LapBCE. As long as LapD remains c-di-GMP bound, it sequesters the periplasmic protease LapG. (b) This LapD-LapG binding prevents LapG cleavage of the N terminus of LapA, and biofilm formation may commence via LapA adherence to a substratum. Low c-di-GMP levels result in a LapD protein unable to sequester LapG, LapG-mediated cleaving of the N-terminal 107 amino acids of LapA, loss of LapA from the cell surface, and a reduction in biofilm formation. From Boyd & O’Toole (6).

Figure 5

GcbC physically delivers c-di-GMP to LapD. The inner membrane DGC requires physical contact to fully deliver its c-di-GMP signal to LapD. The interaction α5^GGDEF helix of GcbC is depicted in blue. The inhibitory site of GcbC has also been shown to be a necessary element for interaction to occur and may help facilitate contact between these two proteins through an as yet undefined mechanism. c-di-GMP is depicted as purple pentagons located at the GcbC inhibitory site and bound at the LapD active site. Each protein functions as a dimer. GcbC has a cache domain that presumably allows enzymatic activity in response to an unidentified signal. The HAMP, GGDEF, EAL, and periplasmic domains of LapD are also shown. When LapD is activated by c-di-GMP, the periplasmic protease LapG is sequestered and the large adhesin LapA accumulates on the surface of the cell, allowing biofilm formation to commence. Adapted from Navarro et al. (45).