Repeated triggering of sporulation in Bacillus subtilis selects against a protein that affects the timing of cell division.
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ABSTRACT:
SUBMITTER: Siebring J
PROVIDER: S-EPMC3869009 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:The Gram-positive bacterium Bacillus subtilis can divide via two modes. During vegetative growth, the division septum is formed at the midcell to produce two equal daughter cells. However, during sporulation, the division septum is formed closer to one pole to yield a smaller forespore and a larger mother cell. Using cryo-electron tomography, genetics and fluorescence microscopy, we found that the organization of the division machinery is different in the two septa. While FtsAZ filaments, the major orchestrators of bacterial cell division, are present uniformly around the leading edge of the invaginating vegetative septa, they are only present on the mother cell side of the invaginating sporulation septa. We provide evidence suggesting that the different distribution and number of FtsAZ filaments impact septal thickness, causing vegetative septa to be thicker than sporulation septa already during constriction. Finally, we show that a sporulation-specific protein, SpoIIE, regulates asymmetric divisome localization and septal thickness during sporulation.
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Project description:While vegetative Bacillus subtilis cells and mature spores are both surrounded by a thick layer of peptidoglycan (PG, a polymer of glycan strands cross-linked by peptide bridges), it has remained unclear whether PG surrounds prespores during engulfment. To clarify this issue, we generated a slender ?ponA mutant that enabled high-resolution electron cryotomographic imaging. Three-dimensional reconstructions of whole cells in near-native states revealed a thin PG-like layer extending from the lateral cell wall around the prespore throughout engulfment. Cryotomography of purified sacculi and fluorescent labelling of PG in live cells confirmed that PG surrounds the prespore. The presence of PG throughout engulfment suggests new roles for PG in sporulation, including a new model for how PG synthesis might drive engulfment, and obviates the need to synthesize a PG layer de novo during cortex formation. In addition, it reveals that B. subtilis can synthesize thin, Gram-negative-like PG layers as well as its thick, archetypal Gram-positive cell wall. The continuous transformations from thick to thin and back to thick during sporulation suggest that both forms of PG have the same basic architecture (circumferential). Endopeptidase activity may be the main switch that governs whether a thin or a thick PG layer is assembled.
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Project description:This series of experiments was designed to identify the program of gene transcription for a single differentiating cell type during sporulation in Bacillus subtilis. The mother cell is one of two cell types generated by asymmetric division of sporulating cells approximately two hours after initiation of sporulation. The program is governed by a hierarchical cascade consisting of the transcription factors: sigmaE, sigmaK, GerE, GerR (YlbO) and SpoIIID. The characterization of the sigmaE regulon was reported in Eichenberger et al. (2003), J. Mol. Biol. 327, 945-972. Here we report the data for sigmaK, GerE, GerR and SpoIIID.
2004-09-01 | GSE1620 | GEO
Project description:SpoIIE is a dual-function protein in Bacillus subtilis that contributes to the switch from medial to polar cell division during sporulation and is responsible for activating the cell-specific transcription factor sigma(F). SpoIIE consists of an N-terminal domain with 10 membrane-spanning segments (region I), a C-terminal phosphatase domain (region III), and a central domain (region II) of uncertain function. To investigate the role of SpoIIE in polar division, we took advantage of a system for efficiently producing polar septa during growth in a SpoIIE-dependent manner using cells engineered to produce the sporulation protein in response to an inducer. The results show that regions II and III play a critical role in polar septum formation and that specific amino acid substitutions in those regions affect the abilities of SpoIIE both to promote polar division and to localize to the division machinery. Additionally, we show that neither the phosphatase function of SpoIIE nor the N-terminal, membrane-spanning region is needed for the switch to asymmetric division.
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Project description:Quorum sensing (QS) is often required for the formation of bacterial biofilms and is a popular target of biofilm control strategies. Previous studies implicate the ComQXPA quorum sensing system of Bacillus subtilis as a promoter of biofilm formation. Here, we report that ComX signaling peptide deficient mutants form thicker and more robust pellicle biofilms that contain chains of cells. We confirm that ComX positively affects the transcriptional activity of the PepsA promoter, which controls the synthesis of the major matrix polysaccharide. In contrast, ComX negatively controls the PtapA promoter, which drives the production of TasA, a fibrous matrix protein. Overall, the biomass of the mutant biofilm lacking ComX accumulates more monosaccharide and protein content than the wild type. We conclude that this QS phenotype might be due to extended investment into growth rather than spore development. Consistent with this, the ComX deficient mutant shows a delayed activation of the pre-spore specific promoter, PspoIIQ, and a delayed, more synchronous commitment to sporulation. We conclude that ComX mediated early commitment to sporulation of the wild type slows down biofilm formation and modulates the coexistence of multiple biological states during the early stages of biofilm development.
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Project description:Starving Bacillus subtilis cells execute a gene expression program resulting in the formation of stress-resistant spores. Sporulation master regulator, Spo0A, is activated by a phosphorelay and controls the expression of a multitude of genes, including the forespore-specific sigma factor σ(F) and the mother cell-specific sigma factor σ(E). Identification of the system-level mechanism of the sporulation decision is hindered by a lack of direct control over Spo0A activity. This limitation can be overcome by using a synthetic system in which Spo0A activation is controlled by inducing expression of phosphorelay kinase KinA. This induction results in a switch-like increase in the number of sporulating cells at a threshold of KinA. Using a combination of mathematical modeling and single-cell microscopy, we investigate the origin and physiological significance of this ultrasensitive threshold. The results indicate that the phosphorelay is unable to achieve a sufficiently fast and ultrasensitive response via its positive feedback architecture, suggesting that the sporulation decision is made downstream. In contrast, activation of σ(F) in the forespore and of σ(E) in the mother cell compartments occurs via a cascade of coherent feed-forward loops, and thereby can produce fast and ultrasensitive responses as a result of KinA induction. Unlike σ(F) activation, σ(E) activation in the mother cell compartment only occurs above the KinA threshold, resulting in completion of sporulation. Thus, ultrasensitive σ(E) activation explains the KinA threshold for sporulation induction. We therefore infer that under uncertain conditions, cells initiate sporulation but postpone making the sporulation decision to average stochastic fluctuations and to achieve a robust population response.
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Project description:In microbial communities, bacteria chemically and physically interact with one another. Some of these interactions are mediated by secreted specialized metabolites that act as either intraspecies or interspecies signals to alter gene expression and to change cell physiology. Bacillus subtilis is a well-characterized soil microbe that can differentiate into multiple cell types, including metabolically dormant endospores. We were interested in identifying microbial interactions that affected sporulation in B. subtilis Using a fluorescent transcriptional reporter, we observed that coculturing B. subtilis with Escherichia coli promoted sporulation gene expression via a secreted metabolite. To identify the active compound, we screened the E. coli Keio Collection and identified the sporulation-accelerating cue as the siderophore enterobactin. B. subtilis has multiple iron acquisition systems that are used to take up the B. subtilis-produced siderophore bacillibactin, as well as to pirate exogenous siderophores such as enterobactin. While B. subtilis uses a single substrate binding protein (FeuA) to take up both bacillibactin and enterobactin, we discovered that it requires two distinct genes to sporulate in response to these siderophores (the esterase gene besA for bacillibactin and a putative esterase gene, ybbA, for enterobactin). In addition, we found that siderophores from a variety of other microbial species also promote sporulation in B. subtilis Our results thus demonstrate that siderophores can act not only as bacterial iron acquisition systems but also as interspecies cues that alter cellular development and accelerate sporulation in B. subtilisIMPORTANCE While much is known about the genetic regulation of Bacillus subtilis sporulation, little is understood about how other bacteria influence this process. This work describes an interaction between Escherichia coli and B. subtilis that accelerates sporulation in B. subtilis The interaction is mediated by the E. coli siderophore enterobactin; we show that other species' siderophores also promote sporulation gene expression in B. subtilis These results suggest that siderophores not only may supply bacteria with the mineral nutrient iron but also may play a role in bacterial interspecies signaling, providing a cue for sporulation. Siderophores are produced by many bacterial species and thus potentially play important roles in altering bacterial cell physiology in diverse environments.
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Project description:To facilitate more accurate spore proteomic analysis, the current study focuses on inducing homogeneous sporulation by overexpressing kinA and assesses the effect of synchronized sporulation initiation on spore resistance, structures, the germination behavior at single-spore level and the proteome. The results indicate that, in our set up, the sporulation by overexpressing kinA can generate a spore yield of 70% within 8 h. The procedure increases spore wet heat resistance and thickness of the spore coat and cortex layers, whilst delaying the time to spore phase-darkening and burst after addition of germinant. The proteome analysis reveals that the upregulated proteins in the kinA induced spores, compared to spores without kinA induction, as well as the 'wildtype' spores, are mostly involved in spore formation. The downregulated proteins mostly belong to the categories of coping with stress, carbon and nitrogen metabolism, as well as the regulation of sporulation. Thus, while kinA overexpression enhances synchronicity in sporulation initiation, it also has profound effects on the central equilibrium of spore formation and spore germination, through modulation of the spore molecular composition and stress resistance physiology.
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Project description:This series of experiments was designed to identify the program of gene transcription for a single differentiating cell type during sporulation in Bacillus subtilis. The mother cell is one of two cell types generated by asymmetric division of sporulating cells approximately two hours after initiation of sporulation. The program is governed by a hierarchical cascade consisting of the transcription factors: sigmaE, sigmaK, GerE, GerR (YlbO) and SpoIIID. The characterization of the sigmaE regulon was reported in Eichenberger et al. (2003), J. Mol. Biol. 327, 945-972. Here we report the data for sigmaK, GerE, GerR and SpoIIID.
2010-06-06 | E-GEOD-1620 | biostudies-arrayexpress
Project description:Upon nutrient deprivation, Bacillus subtilis initiates the developmental process of sporulation by integrating environmental and extracellular signals. These signals are channeled into a phosphorelay ultimately activating the key transcriptional regulator of sporulation, Spo0A. Subsequently, phosphorylated Spo0A regulates the expression of genes required for sporulation to initiate. Here we identified a group of genes whose transcription levels are controlled by Spo0A during exponential growth. Among them, three upregulated genes, termed sivA, sivB (bslA), and sivC, encode factors found to inhibit Spo0A activation. We furthermore show that the Siv factors operate by reducing the activity of histidine kinases located at the top of the sporulation phosphorelay, thereby decreasing Spo0A phosphorylation. Thus, we demonstrate the existence of modulators, positively controlled by Spo0A, which inhibit inappropriate entry into the costly process of sporulation, when conditions are favorable for exponential growth.
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