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Local control: a hub-based model for the c-di-GMP network.

ABSTRACT: The genome of Pseudomonas fluorescens encodes >50 proteins predicted to play a role in bis-(3'-5')-cyclic dimeric guanosine monophosphate (c-di-GMP)-mediated biofilm formation. We built a network representation of protein-protein interactions and extracted key information via multidimensional scaling (i.e., principal component analysis) of node centrality measures, which measure features of proteins in a network. Proteins of different domain types (diguanylate cyclase, dual domain, phosphodiesterase, PilZ) exhibit unique network behavior and can be accurately classified by their network centrality values (i.e., roles in the network). The predictive power of protein-protein interactions in biofilm formation indicates the possibility of localized pools of c-di-GMP. A regression model showed a statistically significant impact of protein-protein interactions on the extent of biofilm formation in various environments. These results highlight the importance of a localized c-di-GMP signaling, extend our understanding of signaling by this second messenger beyond the current "Bow-tie Model," support a newly proposed "Hub Model," and suggest future avenues of investigation.

SUBMITTER: Vasenina A

PROVIDER: S-EPMC11237430 | biostudies-literature | 2024 May

REPOSITORIES: biostudies-literature

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Local control: a hub-based model for the c-di-GMP network.

Vasenina Anna A Fu Yu Y O'Toole George A GA Mucha Peter J PJ

mSphere 20240409 5

The genome of <i>Pseudomonas fluorescens</i> encodes >50 proteins predicted to play a role in bis-(3'-5')-cyclic dimeric guanosine monophosphate (c-di-GMP)-mediated biofilm formation. We built a network representation of protein-protein interactions and extracted key information via multidimensional scaling (i.e., principal component analysis) of node centrality measures, which measure features of proteins in a network. Proteins of different domain types (diguanylate cyclase, dual domain, phosph ...[more]

PMID: 38591888

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Project description:UnlabelledOver the course of the last 3 decades the role of the second messenger cyclic di-GMP (c-di-GMP) as a master regulator of bacterial physiology was determined. Although the control over c-di-GMP levels via synthesis and breakdown and the allosteric regulation of c-di-GMP over receptor proteins (effectors) and riboswitches have been extensively studied, relatively few effectors have been identified and most are of unknown functions. The obligate predatory bacterium Bdellovibrio bacteriovorus has a peculiar dimorphic life cycle, in which a phenotypic transition from a free-living attack phase (AP) to a sessile, intracellular predatory growth phase (GP) is tightly regulated by specific c-di-GMP diguanylate cyclases. B. bacteriovorus also bears one of the largest complement of defined effectors, almost none of known functions, suggesting that additional proteins may be involved in c-di-GMP signaling. In order to uncover novel c-di-GMP effectors, a c-di-GMP capture-compound mass-spectroscopy experiment was performed on wild-type AP and host-independent (HI) mutant cultures, the latter serving as a proxy for wild-type GP cells. Eighty-four proteins were identified as candidate c-di-GMP binders. Of these proteins, 65 did not include any recognized c-di-GMP binding site, and 3 carried known unorthodox binding sites. Putative functions could be assigned to 59 proteins. These proteins are included in metabolic pathways, regulatory circuits, cell transport, and motility, thereby creating a potentially large c-di-GMP network. False candidate effectors may include members of protein complexes, as well as proteins binding nucleotides or other cofactors that were, respectively, carried over or unspecifically interacted with the capture compound during the pulldown. Of the 84 candidates, 62 were found to specifically bind the c-di-GMP capture compound in AP or in HI cultures, suggesting c-di-GMP control over the whole-cell cycle of the bacterium. High affinity and specificity to c-di-GMP binding were confirmed using microscale thermophoresis with a hypothetical protein bearing a PilZ domain, an acyl coenzyme A dehydrogenase, and a two-component system response regulator, indicating that additional c-di-GMP binding candidates may be bona fide novel effectors.ImportanceIn this study, 84 putative c-di-GMP binding proteins were identified in B. bacteriovorus, an obligate predatory bacterium whose lifestyle and reproduction are dependent on c-di-GMP signaling, using a c-di-GMP capture compound precipitation approach. This predicted complement covers metabolic, energy, transport, motility and regulatory pathways, and most of it is phase specific, i.e., 62 candidates bind the capture compound at defined modes of B. bacteriovorus lifestyle. Three of the putative binders further demonstrated specificity and high affinity to c-di-GMP via microscale thermophoresis, lending support for the presence of additional bona fide c-di-GMP effectors among the pulled-down protein repertoire.

Project description:Bacterial chemotaxis receptors provide the sensory inputs that inform the direction of navigation in changing environments. Recently, we described the bacterial second messenger cyclic di-GMP (c-di-GMP) as a novel regulator of a subclass of chemotaxis receptors. In Azospirillum brasilense, c-di-GMP binds to a chemotaxis receptor, Tlp1, and modulates its signaling function during aerotaxis. Here, we further characterize the role of c-di-GMP in aerotaxis using a novel dichromatic optogenetic system engineered for manipulating intracellular c-di-GMP levels in real time. This system comprises a red/near-infrared-light-regulated diguanylate cyclase and a blue-light-regulated c-di-GMP phosphodiesterase. It allows the generation of transient changes in intracellular c-di-GMP concentrations within seconds of irradiation with appropriate light, which is compatible with the time scale of chemotaxis signaling. We provide experimental evidence that binding of c-di-GMP to the Tlp1 receptor activates its signaling function during aerotaxis, which supports the role of transient changes in c-di-GMP levels as a means of adjusting the response of A. brasilense to oxygen gradients. We also show that intracellular c-di-GMP levels in A. brasilense change with carbon metabolism. Our data support a model whereby c-di-GMP functions to imprint chemotaxis receptors with a record of recent metabolic experience, to adjust their contribution to the signaling output, thus allowing the cells to continually fine-tune chemotaxis sensory perception to their metabolic state.IMPORTANCE Motile bacteria use chemotaxis to change swimming direction in response to changes in environmental conditions. Chemotaxis receptors sense environmental signals and relay sensory information to the chemotaxis machinery, which ultimately controls the swimming pattern of cells. In bacteria studied to date, differential methylation has been known as a mechanism to control the activity of chemotaxis receptors and modulates their contribution to the overall chemotaxis response. Here, we used an optogenetic system to perturb intracellular concentrations of the bacterial second messenger c-di-GMP to show that in some chemotaxis receptors, c-di-GMP functions in a similar feedback loop to connect the metabolic status of the cells to the sensory activity of chemotaxis receptors.

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Local control: a hub-based model for the c-di-GMP network.

Publications

Local control: a hub-based model for the c-di-GMP network.

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