Second messenger regulation of biofilm formation: breakthroughs in understanding c-di-GMP effector systems

Abstract

The second messenger bis-(3'-5')-cyclic dimeric guanosine monophosphate (c-di-GMP) has emerged as a broadly conserved intracellular signaling molecule. This soluble molecule is important for controlling biofilm formation, adhesion, motility, virulence, and cell morphogenesis in diverse bacterial species. But how is the typical bacterial cell able to coordinate the actions of upward of 50 proteins involved in synthesizing, degrading, and binding c-di-GMP? Understanding the specificity of c-di-GMP signaling in the context of so many enzymes involved in making, breaking, and binding the second messenger will be possible only through mechanistic studies of its output systems. Here we discuss three newly characterized c-di-GMP effector systems that are best understood in terms of molecular and structural detail. As they are conserved across many bacterial species, they likely will serve as central paradigms for c-di-GMP output systems and contribute to our understanding of how bacteria control critical aspects of their biology.

Figure 1

c-di-GMP synthesis and degradation. Diguanylate cyclases (DGCs) containing the GGDEF domain synthesize c-di-GMP (shown center) from two molecules of GTP. Phosphodiesterases (PDEs) containing the EAL domain or HD-GYP domain degrade c-di-GMP to pGpG or GMP, respectively. Illustration courtesy of William Scavone, MA, CMI, Kestrel Illustration Studio, LLC. Abbreviations: GMP, guanosine monophosphate; pGpG, 5′-phosphoguanylyl-(3′-5′)-guanosine.

Figure 2

LapD binds c-di-GMP. c-di-GMP binding to LapD induces a conformational change in the protein. (a) Autoinhibited state. Intramolecular interactions among the EAL domain, the GGDEF domain, and the S-helix prevent c-di-GMP from binding to the EAL domain. Structure-guided mutagenesis studies suggest that the S-helix stabilizes the autoinhibited state of LapD, whereas the positioning of the GGDEF domain blocks c-di-GMP from accessing the c-di-GMP binding pocket with the EAL domain (Navarro et al. 2011). (b) c-di-GMP activated state. Upon c-di-GMP binding to the EAL domain, LapD undergoes a conformational change as the S-helix and GGDEF domain are displaced (Navarro et al. 2011; Illustration courtesy of William Scavone, MA, CMI, Kestrel Illustration Studio, LLC). Abbreviations: GMP, guanosine monophosphate.

Figure 3

c-di-GMP effector system in Pseudomonas fluorescens. This diagram depicts a summary of the current model for the c-di-GMP effector system in P. fluorescens, which impacts the motile-to-sessile transition. The LapA protein, a predicted cell-surface adhesion, is transported to the cell surface through the ABC transporter, comprised of the LapBCE proteins. LapD binds c-di-GMP, and through an inside-out signaling mechanism, the periplasmic domain of LapD binds LapG. Thus, LapG is prevented from cleaving and releasing LapA from the cell surface, thereby promoting biofilm formation (Newell et al. 2011a; Illustration courtesy of William Scavone, MA, CMI, Kestrel Illustration Studio, LLC). Abbreviations: GMP, guanosine monophosphate; IM, inner membrane; OM, outer membrane.

Figure 4

c-di-GMP effector system in Escherichia coli. This diagram depicts a summary of the current models of the c-di-GMP effector system in E. coli, which impacts the motile-to-sessile transition. YcgR bound to c-di-GMP stimulates YcgR to interact with one or more components of the flagellar motor, reducing or altering motor function and thus promoting the motile-to-sessile transition. (a) Fang & Gomelsky (2010) suggest that c-di-GMP-bound YcgR interacts with FliG and disrupts the FliG-FliM interaction. An increased bias in counterclockwise flagellar rotation ensues, which leads to poor migration and thus facilitates the motile-to-sessile transition. (b) Paul et al. (2010) suggest that c-di-GMP-bound YcgR binds to FliM, disrupting the FliG-FliM interaction and thereby promoting a counterclockwise rotational bias. They also propose that YcgR also interacts with FliG, which leads to a disruption in the FliG-MotA interactions and a reduction in torque generation. (c) Boehm et al. (2010) suggest that c-di-GMP-bound YcgR interacts with MotA to reduce flagellar motor function. Illustration courtesy of William Scavone, MA, CMI, Kestrel Illustration Studio, LLC. Abbreviations: GMP, guanosine monophosphate; IM, inner membrane; OM, outer membrane.

Figure 5

c-di-GMP effector system in Vibrio cholerae. This diagram depicts a summary of the current model of the c-di-GMP effector system in V. cholerae, which impacts the motile-to-sessile transition. The transcriptional regulator VpsT binds c-di-GMP to positively regulate the transcription of the vps genes encoding the proteins needed for VPS production. The VPS polysaccharide is required for biofilm formation (Krasteva et al. 2010; Illustration courtesy of William Scavone, MA, CMI, Kestrel Illustration Studio, LLC).