Dataset Information

Pulcherrimin formation controls growth arrest of the Bacillus subtilis biofilm.

ABSTRACT: Biofilm formation by Bacillus subtilis is a communal process that culminates in the formation of architecturally complex multicellular communities. Here we reveal that the transition of the biofilm into a nonexpanding phase constitutes a distinct step in the process of biofilm development. Using genetic analysis we show that B. subtilis strains lacking the ability to synthesize pulcherriminic acid form biofilms that sustain the expansion phase, thereby linking pulcherriminic acid to growth arrest. However, production of pulcherriminic acid is not sufficient to block expansion of the biofilm. It needs to be secreted into the extracellular environment where it chelates Fe3+ from the growth medium in a nonenzymatic reaction. Utilizing mathematical modeling and a series of experimental methodologies we show that when the level of freely available iron in the environment drops below a critical threshold, expansion of the biofilm stops. Bioinformatics analysis allows us to identify the genes required for pulcherriminic acid synthesis in other Firmicutes but the patchwork presence both within and across closely related species suggests loss of these genes through multiple independent recombination events. The seemingly counterintuitive self-restriction of growth led us to explore if there were any benefits associated with pulcherriminic acid production. We identified that pulcherriminic acid producers can prevent invasion by neighboring communities through the generation of an "iron-free" zone, thereby addressing the paradox of pulcherriminic acid production by B. subtilis.

SUBMITTER: Arnaouteli S

PROVIDER: S-EPMC6613138 | biostudies-literature | 2019 Jul

REPOSITORIES: biostudies-literature

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Pulcherrimin formation controls growth arrest of the Bacillus subtilis biofilm.

Arnaouteli Sofia S Matoz-Fernandez D A DA Porter Michael M Kalamara Margarita M Abbott James J MacPhee Cait E CE Davidson Fordyce A FA Stanley-Wall Nicola R NR

Proceedings of the National Academy of Sciences of the United States of America 20190619 27

Biofilm formation by Bacillus subtilis is a communal process that culminates in the formation of architecturally complex multicellular communities. Here we reveal that the transition of the biofilm into a nonexpanding phase constitutes a distinct step in the process of biofilm development. Using genetic analysis we show that B. subtilis strains lacking the ability to synthesize pulcherriminic acid form biofilms that sustain the expansion phase, thereby linking pulcherriminic acid t ...[more]

PMID: 31217292

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Project description:Biofilm development in Bacillus subtilis is regulated at multiple levels. While a number of known signals that trigger biofilm formation do so through the activation of one or more sensory histidine kinases, it was discovered that biofilm activation is also coordinated by sensing intracellular metabolic signals, including serine starvation. Serine starvation causes ribosomes to pause on specific serine codons, leading to a decrease in the translation rate of sinR, which encodes a master repressor for biofilm matrix genes and ultimately triggers biofilm induction. How serine levels change in different growth stages, how B. subtilis regulates intracellular serine levels, and how serine starvation triggers ribosomes to pause on selective serine codons remain unknown. Here, we show that serine levels decrease as cells enter stationary phase and that unlike most other amino acid biosynthesis genes, expression of serine biosynthesis genes decreases upon the transition into stationary phase. The deletion of the gene for a serine deaminase responsible for converting serine to pyruvate led to a delay in biofilm formation, further supporting the idea that serine levels are a critical intracellular signal for biofilm activation. Finally, we show that levels of all five serine tRNA isoacceptors are decreased in stationary phase compared with exponential phase. However, the three isoacceptors recognizing UCN serine codons are reduced to a much greater extent than the two that recognize AGC and AGU serine codons. Our findings provide evidence for a link between serine homeostasis and biofilm development in B. subtilisIMPORTANCE In Bacillus subtilis, biofilm formation is triggered in response to environmental and cellular signals. It was proposed that serine limitation acts as a proxy for nutrient status and triggers biofilm formation at the onset of biofilm entry through a novel signaling mechanism caused by global ribosome pausing on selective serine codons. In this study, we reveal that serine levels decrease at the biofilm entry due to catabolite control and a serine shunt mechanism. We also show that levels of five serine tRNA isoacceptors are differentially decreased in stationary phase compared with exponential phase; three isoacceptors recognizing UCN serine codons are reduced much more than the two recognizing AGC and AGU codons. This finding indicates a possible mechanism for selective ribosome pausing.

Project description:Bacillus subtilis forms robust biofilms in the presence of large amounts of carbon sources, such as glycerol. However, little is known about the importance of the metabolic systems, or the relationship between metabolic systems and regulatory systems, involved in biofilm formation. Glutamate synthase, encoded by gltAB, is an enzyme that converts 2-ketoglutarate (a tricarboxylic acid [TCA] cycle intermediate) and glutamine into glutamate, which is a general amino group donor in metabolism. Here, we show that a ?gltA mutant exhibited early arrest of biofilm formation in complex medium containing glycerol. This phenotype was not due to glutamate auxotrophy. Consistent with its biofilm formation phenotype, the ?gltA mutant exhibited an early decrease in expression of the epsA and tapA operons, which are responsible for production of biofilm matrix polymers. This resulted from decreased activity of their regulator, Spo0A, as evidenced by reduced expression of other Spo0A-regulated genes in the ?gltA mutant. The ?gltA mutation prevented biofilm formation only in the presence of large amounts of glycerol. Moreover, limited expression of citrate synthase (but not other TCA enzymes) restored biofilm-forming ability to the ?gltA mutant. These results indicate that the ?gltA mutant accumulates an inhibitory intermediate (citrate) in the TCA cycle in the presence of large amounts of glycerol. The ?gltA mutant formed biofilms when excess iron was added to the medium. Taken together, the data suggest that accumulation of citrate ions by the ?gltA mutant causes iron shortage due to chelation, which prevents activation of Spo0A and causes defective biofilm formation.IMPORTANCE Bacillus subtilis, a model organism for bacterial biofilm formation, forms robust biofilms in a medium-dependent manner. Although the regulatory network that controls biofilm formation has been well studied, the importance of the underlying metabolic systems remains to be elucidated. The present study demonstrates that a metabolic disorder in a well-conserved metabolic system causes accumulation of an inhibitory metabolic intermediate that prevents activation of the system that regulates biofilm formation. These findings increase our understanding of the coordination between cellular metabolic status and the regulatory networks governing biofilm formation.

Dataset Information

Pulcherrimin formation controls growth arrest of the Bacillus subtilis biofilm.

Publications

Pulcherrimin formation controls growth arrest of the <i>Bacillus subtilis</i> biofilm.

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