More than Enzymes That Make or Break Cyclic Di-GMP-Local Signaling in the Interactome of GGDEF/EAL Domain Proteins of Escherichia coli.

Olga Sarenko,Franziska M Wilke,Gisela Klauck,Anja M Richter,Volkhard Kaever,Susanne Herbst,Regine Hengge,Vanessa Pfiffer,Howard A Shuman

doi:10.1128/mbio.01639-17

Olga Sarenko, Franziska M Wilke + Show 7 more

Open Access

https://doi.org/10.1128/mbio.01639-17

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Abstract

ABSTRACTThe bacterial second messenger bis-(3′-5′)-cyclic diguanosine monophosphate (c-di-GMP) ubiquitously promotes bacterial biofilm formation. Intracellular pools of c-di-GMP seem to be dynamically negotiated by diguanylate cyclases (DGCs, with GGDEF domains) and specific phosphodiesterases (PDEs, with EAL or HD-GYP domains). Most bacterial species possess multiple DGCs and PDEs, often with surprisingly distinct and specific output functions. One explanation for such specificity is “local” c-di-GMP signaling, which is believed to involve direct interactions between specific DGC/PDE pairs and c-di-GMP-binding effector/target systems. Here we present a systematic analysis of direct protein interactions among all 29 GGDEF/EAL domain proteins of Escherichia coli. Since the effects of interactions depend on coexpression and stoichiometries, cellular levels of all GGDEF/EAL domain proteins were also quantified and found to vary dynamically along the growth cycle. Instead of detecting specific pairs of interacting DGCs and PDEs, we discovered a tightly interconnected protein network of a specific subset or “supermodule” of DGCs and PDEs with a coregulated core of five hyperconnected hub proteins. These include the DGC/PDE proteins representing the c-di-GMP switch that turns on biofilm matrix production in E. coli. Mutants lacking these core hub proteins show drastic biofilm-related phenotypes but no changes in cellular c-di-GMP levels. Overall, our results provide the basis for a novel model of local c-di-GMP signaling in which a single strongly expressed master PDE, PdeH, dynamically eradicates global effects of several DGCs by strongly draining the global c-di-GMP pool and thereby restricting these DGCs to serving as local c-di-GMP sources that activate specific colocalized effector/target systems.

Highlights

The bacterial second messenger bis-(3=-5=)-cyclic diguanosine monophosphate (c-di-GMP) ubiquitously promotes bacterial biofilm formation
The nucleotide second messenger bis-(3=-5=)-cyclic diguanosine monophosphate (c-di-GMP) activates genes involved in biofilm formation or directly stimulates the biosynthesis and secretion of the exopolysaccharides found in the extracellular matrix of bacterial biofilms
Cellular c-di-GMP is controlled by antagonistic enzymes, i.e., diguanylate cyclases (DGCs [with GGDEF domains]) and specific phosphodiesterases (PDEs [which can feature either EAL or HD-GYP domains])

Summary

Introduction

The bacterial second messenger bis-(3=-5=)-cyclic diguanosine monophosphate (c-di-GMP) ubiquitously promotes bacterial biofilm formation. Most bacterial species possess multiple DGCs and PDEs, often with surprisingly distinct and specific output functions One explanation for such specificity is “local” c-di-GMP signaling, which is believed to involve direct interactions between specific DGC/PDE pairs and c-di-GMP-binding effector/target systems. Instead of detecting specific pairs of interacting DGCs and PDEs, we discovered a tightly interconnected protein network of a specific subset or “supermodule” of DGCs and PDEs with a coregulated core of five hyperconnected hub proteins These include the DGC/PDE proteins representing the c-di-GMP switch that turns on biofilm matrix production in E. coli. Escherichia coli K-12 has 29 GGDEF/EAL domain proteins, including 12 DGCs and 13 PDEs [5, 6] Despite this multiplicity, single distinct DGCs and/or PDEs can generate distinct and highly specific outputs suggesting localized operation in conjunction with specific targets [7,8,9]. This releases DgcM and MlrA from direct inhibition by PdeR, which allows DgcM/MlrA-driven expression of the biofilm regulator CsgD [13, 14, 17]

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