Chapter 8 : Choosing the Right Lifestyle: Regulation of Developmental Pathways by Cyclic Di-GMP

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This chapter provides an understanding of the general developmental concepts associated with cyclic di-GMP (c-di-GMP) regulation. It also provides a broad understanding of the concepts of planktonic and biofilm lifestyles and the modes utilized by bacteria to achieve directed movement. There are distinct advantages to both free-swimming and biofilm lifestyles. In general, free-swimming bacteria may be better suited to identifying new environments, permitting colonization, and proliferation. The authors discuss the two lifestyles by defining the regulatory principles associated with c-di-GMP important for the transition between these lifestyles. They then discuss the biofilm lifestyle and the transition to this lifestyle from a planktonic state. A central concept of c-di-GMP regulation is that its accumulation favors biofilm formation and inhibits motility and the planktonic lifestyle. Importantly, the ability to utilize one or more of the modes of movement can dictate the efficiency of the transition from the planktonic to biofilm lifestyle. Furthermore, it is not only the expression of these systems that is being controlled by c-di-GMP. Another exciting area of exploration is identifying the environmental signals that control c-di-GMP signaling. The fascinating flexibility of c-di-GMP has the net result of a versatile signaling molecule that can induce a gradual response, through changes in gene expression, as well as an immediate response by directly altering the functionality of macromolecular assembly pathways.

Citation: Parsek M, Aldridge P. 2010. Choosing the Right Lifestyle: Regulation of Developmental Pathways by Cyclic Di-GMP, p 99-119. In Wolfe A, Visick K (ed), The Second Messenger Cyclic Di-GMP. ASM Press, Washington, DC. doi: 10.1128/9781555816667.ch8

Key Concept Ranking

Gene Expression and Regulation
Cellular Processes
Type IV Pili
Filament Cap Protein
Outer Membrane Proteins
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Image of Figure 1.
Figure 1.

Influence of c-di-GMP in controlling the transition from the planktonic to biofilm lifestyle and vice versa. DGC enzymes make c-di-GMP, while its breakdown is mediated by either PDE or HD-GYP activity. The observation that many mutants in the genes that encode DGC and PDE of HD-GYP enzymes result in pleiotropic or incomplete phenotypes suggests that cells use c-di-GMP concentrations ([c-di-GMP]) as a means to measure when it is optimal to fully commit to either lifestyle. As a result, there is a level of c-di-GMP where both planktonic and biofilm traits can still be functional (overlapping circles). Only once the thresholds of this intersection have been passed will a cell commit to either lifestyle.

Citation: Parsek M, Aldridge P. 2010. Choosing the Right Lifestyle: Regulation of Developmental Pathways by Cyclic Di-GMP, p 99-119. In Wolfe A, Visick K (ed), The Second Messenger Cyclic Di-GMP. ASM Press, Washington, DC. doi: 10.1128/9781555816667.ch8
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Image of Figure 2.
Figure 2.

Diagram showing the different stages of biofilm formation that are influenced by c-di-GMP (indicated by the stars). Elevated c-di-GMP promotes initial attachment, which involves adherence of free-swimming cells to a surface. In the case of a flat biofilm, cells continue to multiply and move on the surface, forming a confluent, flat mat of cells. In the case of structured biofilms, we present two alternative routes to their formation. The first we call structured biofilm I. Here, dark gray cells represent the immobile stalks of structured biofilms, while light gray cells represent the motile subpopulation that produce the cap. The second we call structured biofilm II. In both cases, the structured biofilms are characterized by EPS production (either Pel, Psl, or alginate, depending upon the strain). EPS production is promoted by elevated c-di-GMP. In this case, small cell aggregates grow clonally, forming large cell aggregates consisting of cells primarily derived from cells in the small cell aggregates. This is indicated by the large aggregate of dark gray cells in the figure, indicating that the cells in the aggregate are progeny derived from the initial small aggregates. Finally, cells can actively leave the biofilm to reinitiate the cycle in a process called dispersion or detachment. c-di-GMP has also been implicated in this step.

Citation: Parsek M, Aldridge P. 2010. Choosing the Right Lifestyle: Regulation of Developmental Pathways by Cyclic Di-GMP, p 99-119. In Wolfe A, Visick K (ed), The Second Messenger Cyclic Di-GMP. ASM Press, Washington, DC. doi: 10.1128/9781555816667.ch8
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Image of Figure 3.
Figure 3.

Schematic diagram depicting the alginate biosynthetic machinery. c-di-GMP binds the PilZ domain of Alg44, acting as a positive allosteric activator. Alg8 and Alg44, inner membrane (I.M.) proteins; K (AlgK), periplasmic lipoprotein with lipid anchor in the outer membrane (O.M.) and a tetratricopeptide repeat scaffold protein; E (AlgE), outer membrane porin-like protein that interacts with AlgK; G (AlgG), periplasmic protein that has a right-handed beta-helical domain and contains the epimerase active site; L (AlgL), a periplasmic protein likely involved in alginate processing; F (AlgF), periplasmic protein; J (AlgJ), periplasmic protein anchored in the inner membrane by a noncleaved signal peptide; I (AlgI), inner membrane protein with seven membrane-spanning domains; X (AlgX), periplasmic protein (function not known yet).

Citation: Parsek M, Aldridge P. 2010. Choosing the Right Lifestyle: Regulation of Developmental Pathways by Cyclic Di-GMP, p 99-119. In Wolfe A, Visick K (ed), The Second Messenger Cyclic Di-GMP. ASM Press, Washington, DC. doi: 10.1128/9781555816667.ch8
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Image of Figure 4.
Figure 4.

c-di-GMP controls Pel production at two levels. (A) Schematic depicting the predicted localization of the Pel bio-synthetic machinery. The localization is primarily based upon bioinformatics, except in the case of PelC, which has been shown experimentally to localize to the outer membrane. c-di-GMP acts as an allosteric activator of PelD. The Pel proteins are predicted to have the following functions. PelA is a large soluble, periplasmic protein with no clear function; it has a large amount of disorder but may adopt a TIM barrel structure. PelB is an outer membrane protein with a large N-terminal TPR domain followed by a C-terminal porin domain. PelC is an outer membrane lipoprotein that may adopt a TolB-like structure. PelD is a cytoplasmic membrane protein with four transmembrane helices (TMs) that has a c-di-GMP binding domain. PelE is a cytoplasmic membrane protein with two TMs followed by a periplasmic region (residues 90 to 320) carrying four or five TPR motifs (residues 155 to 310). Residues 90 to 150 are predicted to carry loops and helices; residues ∼115 to 127 may be disordered. PelF is a cytoplasmic glycosyltransferase. PelG is a cytoplasmic membrane protein with 12 TMs that is a member of the polysaccharide transporter family and also resembles Na/H antiporters. (B) c-di-GMP also binds to the transcriptional repressor FleQ. When c-di-GMP levels are low, FleQ represses expression (left). At elevated levels, c-di-GMP binds FleQ, relieving repression of expression (right).

Citation: Parsek M, Aldridge P. 2010. Choosing the Right Lifestyle: Regulation of Developmental Pathways by Cyclic Di-GMP, p 99-119. In Wolfe A, Visick K (ed), The Second Messenger Cyclic Di-GMP. ASM Press, Washington, DC. doi: 10.1128/9781555816667.ch8
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Image of Figure 5.
Figure 5.

Contrasting patterns of flagellum-mediated swimming and swarming used to exemplify the difference between these two modes of movement. (A) serovar Typhimurium exhibiting flagellum-mediated swimming and swarming; (B) Motility assays for swimming are performed using 0.3% agar while swarming assays utilize between 0.5 and 0.7% agar. On 0.3% agar, the resulting swarm is of a uniform diameter containing characteristic rings embedded within the agar matrix. These rings are waves of bacteria utilizing chemotaxis to alternative nutrients in the medium. In contrast, swarms produced on 0.5 to 0.7% agar are much more spectacular in shape and diameter.

Citation: Parsek M, Aldridge P. 2010. Choosing the Right Lifestyle: Regulation of Developmental Pathways by Cyclic Di-GMP, p 99-119. In Wolfe A, Visick K (ed), The Second Messenger Cyclic Di-GMP. ASM Press, Washington, DC. doi: 10.1128/9781555816667.ch8
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Image of Figure 6.
Figure 6.

Schematic diagram of the type IV pilus and the assembly and retraction cycles mediated by the two ATPases PilB and PilT. The six major components are not all depicted in the structure for clarity, and the inner membrane anchor PilD and the proposed secretin PilQ are labeled. The models for assembly and retraction reflect the proposed models of Craig and Li ( ) (assembly) and Kaiser ( ) (retraction). An underlying principle of both cycles is that pilin subunits can be stored in the inner membrane until needed. The control of pilus assembly by c-di-GMP through PilZ is highlighted. The gray arrows indicate alternative routes for c-di-GMP regulation of the process, most of which require further investigation.

Citation: Parsek M, Aldridge P. 2010. Choosing the Right Lifestyle: Regulation of Developmental Pathways by Cyclic Di-GMP, p 99-119. In Wolfe A, Visick K (ed), The Second Messenger Cyclic Di-GMP. ASM Press, Washington, DC. doi: 10.1128/9781555816667.ch8
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Image of Figure 7.
Figure 7.

Schematic diagram comparing the coordination of flagellar gene expression and assembly when one or two assembly checkpoints are used. The important structural features of the flagellum are highlighted. One checkpoint: this pathway is utilized by the paradigm flagellar systems of and serovar Typhimurium. The master regulator FlhDC activates P in conjunction with This promoter class drives the expression of the HBB structural components and several regulatory proteins, including and FlgM. Upon HBB completion, FlgM is secreted, allowing to initiate transcription from P. Two checkpoints: one example of this pathway is that used by Here proximal basal body gene expression is activated by the atypical σ EBP, FleQ. A second σ EBP recognizes the initiation of T3S/rod assembly, activating transcription of genes that encode the distal HBB structural components and a number of other regulatory components. For HBB completion is sensed in a similar manner to that of that results in σ activating flagellin gene expression. In this checkpoint is coupled to flagellin translation, not transcription, as an added twist. Note, does not utilize a FleQ homologue to activate its transcription hierarchy but does utilize a σ EBP sensory system to coordinate distal basal body gene expression and assembly. Txn., transcription; Tln., translation.

Citation: Parsek M, Aldridge P. 2010. Choosing the Right Lifestyle: Regulation of Developmental Pathways by Cyclic Di-GMP, p 99-119. In Wolfe A, Visick K (ed), The Second Messenger Cyclic Di-GMP. ASM Press, Washington, DC. doi: 10.1128/9781555816667.ch8
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Image of Figure 8.
Figure 8.

Summary of the multiple targets during flagellar assembly influenced by c-di-GMP regulation. (Gray box) Flagellar rotation is negatively controlled by the homologues YcgR and DgrA in (ec) and (cc), respectively. Flagellar assembly in and (vf) is controlled at a posttranscriptional level during HBB assembly, potentially to control the Mot-to-Mot transition. A direct role for c-di-GMP has been shown for the flagellar master regulator FleQ (see Fig. 3 ).

Citation: Parsek M, Aldridge P. 2010. Choosing the Right Lifestyle: Regulation of Developmental Pathways by Cyclic Di-GMP, p 99-119. In Wolfe A, Visick K (ed), The Second Messenger Cyclic Di-GMP. ASM Press, Washington, DC. doi: 10.1128/9781555816667.ch8
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