skip to main content
US FlagAn official website of the United States government
dot gov icon
Official websites use .gov
A .gov website belongs to an official government organization in the United States.
https lock icon
Secure .gov websites use HTTPS
A lock ( lock ) or https:// means you've safely connected to the .gov website. Share sensitive information only on official, secure websites.

Explore Research Products in the PAR
It may take a few hours for recently added research products to appear in PAR search results.


  • NSF Public Access
  • Search Results
  • The Slowdown of Growth Rate Controls the Single-Cell Distribution of Biofilm Matrix Production via an SinI-SinR-SlrR Network
Title: The Slowdown of Growth Rate Controls the Single-Cell Distribution of Biofilm Matrix Production via an SinI-SinR-SlrR Network
ABSTRACT In Bacillus subtilis , master regulator Spo0A controls several cell-differentiation pathways. Under moderate starvation, phosphorylated Spo0A (Spo0A~P) induces biofilm formation by indirectly activating genes controlling matrix production in a subpopulation of cells via an SinI-SinR-SlrR network. Under severe starvation, Spo0A~P induces sporulation by directly and indirectly regulating sporulation gene expression. However, what determines the heterogeneity of individual cell fates is not fully understood. In particular, it is still unclear why, despite being controlled by a single master regulator, biofilm matrix production and sporulation seem mutually exclusive on a single-cell level. In this work, with mathematical modeling, we showed that the fluctuations in the growth rate and the intrinsic noise amplified by the bistability in the SinI-SinR-SlrR network could explain the single-cell distribution of matrix production. Moreover, we predicted an incoherent feed-forward loop; the decrease in the cellular growth rate first activates matrix production by increasing in Spo0A phosphorylation level but then represses it via changing the relative concentrations of SinR and SlrR. Experimental data provide evidence to support model predictions. In particular, we demonstrate how the degree to which matrix production and sporulation appear mutually exclusive is affected by genetic perturbations. IMPORTANCE The mechanisms of cell-fate decisions are fundamental to our understanding of multicellular organisms and bacterial communities. However, even for the best-studied model systems we still lack a complete picture of how phenotypic heterogeneity of genetically identical cells is controlled. Here, using B. subtilis as a model system, we employ a combination of mathematical modeling and experiments to explain the population-level dynamics and single-cell level heterogeneity of matrix gene expression. The results demonstrate how the two cell fates, biofilm matrix production and sporulation, can appear mutually exclusive without explicitly inhibiting one another. Such a mechanism could be used in a wide range of other biological systems.  more » « less
Award ID(s):
2204402
PAR ID:
10409009
Author(s) / Creator(s):
; ; ; ;
Editor(s):
Kovács, Ákos T.
Date Published:
Journal Name:
mSystems
ISSN:
2379-5077
Format(s):
Medium: X
Sponsoring Org:
National Science Foundation
More Like this
  1. ; ; ; ; ; (, mBio)
    Zhulin, Igor B. (Ed.)
    ABSTRACT In Bacillus subtilis , biofilm and sporulation pathways are both controlled by a master regulator, Spo0A, which is activated by phosphorylation via a phosphorelay—a cascade of phosphotransfer reactions commencing with autophosphorylation of histidine kinases KinA, KinB, KinC, KinD, and KinE. However, it is unclear how the kinases, despite acting via the same regulator, Spo0A, differentially regulate downstream pathways, i.e., how KinA mainly activates sporulation genes and KinC mainly activates biofilm genes. In this work, we found that KinC also downregulates sporulation genes, suggesting that KinC has a negative effect on Spo0A activity. To explain this effect, with a mathematical model of the phosphorelay, we revealed that unlike KinA, which always activates Spo0A, KinC has distinct effects on Spo0A at different growth stages: during fast growth, KinC acts as a phosphate source and activates Spo0A, whereas during slow growth, KinC becomes a phosphate sink and contributes to decreasing Spo0A activity. However, under these conditions, KinC can still increase the population-mean biofilm matrix production activity. In a population, individual cells grow at different rates, and KinC would increase the Spo0A activity in the fast-growing cells but reduce the Spo0A activity in the slow-growing cells. This mechanism reduces single-cell heterogeneity of Spo0A activity, thereby increasing the fraction of cells that activate biofilm matrix production. Thus, KinC activates biofilm formation by controlling the fraction of cells activating biofilm gene expression. IMPORTANCE In many bacterial and eukaryotic systems, multiple cell fate decisions are activated by a single master regulator. Typically, the activities of the regulators are controlled posttranslationally in response to different environmental stimuli. The mechanisms underlying the ability of these regulators to control multiple outcomes are not understood in many systems. By investigating the regulation of Bacillus subtilis master regulator Spo0A, we show that sensor kinases can use a novel mechanism to control cell fate decisions. By acting as a phosphate source or sink, kinases can interact with one another and provide accurate regulation of the phosphorylation level. Moreover, this mechanism affects the cell-to-cell heterogeneity of the transcription factor activity and eventually determines the fraction of different cell types in the population. These results demonstrate the importance of intercellular heterogeneity for understanding the effects of genetic perturbations on cell fate decisions. Such effects can be applicable to a wide range of cellular systems. 
    more » « less
  2. ; ; ; ; ; ; ; (, mSystems)
    Methe, Barbara (Ed.)
    ABSTRACT Environmental strains of the soil bacterium Bacillus subtilis have valuable applications in agriculture, industry, and biotechnology; however, environmental strains are genetically less accessible. This reduced accessibility is in sharp contrast to laboratory strains, which are well known for their natural competence, and a limitation in their applications. In this study, we observed that robust biofilm formation by environmental strains of B. subtilis greatly reduced the frequency of competent cells in the biofilm. By using model strain 3610, we revealed a cross-pathway regulation that allows biofilm matrix producers and competence-developing cells to undergo mutually exclusive cell differentiation. We further demonstrated that the competence activator ComK represses the key biofilm regulatory gene sinI by directly binding to the sinI promoter, thus blocking competent cells from simultaneously becoming matrix producers. In parallel, the biofilm activator SlrR represses competence through three distinct mechanisms involving both genetic regulation and cell morphological changes. Finally, we discuss the potential implications of limiting competence in a bacterial biofilm. IMPORTANCE The soil bacterium Bacillus subtilis can form robust biofilms, which are important for its survival in the environment. B. subtilis also exhibits natural competence. By investigating competence development in B. subtilis in situ during biofilm formation, we reveal that robust biofilm formation often greatly reduces the frequency of competent cells within the biofilm. We then characterize a cross-pathway regulation that allows cells in these two developmental events to undergo mutually exclusive cell differentiation during biofilm formation. Finally, we discuss potential biological implications of limiting competence in a bacterial biofilm. 
    more » « less
  3. ; ; ; ; ; ; ; (, Journal of Bacteriology)
    ABSTRACT 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. subtilis . IMPORTANCE 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. 
    more » « less
  4. ; ; ; ; ; ; (, mSystems)
    Svensson, Sarah L (Ed.)
    ABSTRACT In starvingBacillus subtilisbacteria,the initiation of two survival programs—biofilm formation and sporulation—is controlled by the same phosphorylated master regulator, Spo0A~P. Its gene,spo0A,is transcribed from two promoters, Pvand Ps,that are, respectively, regulated by RNA polymerase (RNAP) holoenzymes bearing σAand σH. Notably, transcription is directly autoregulated by Spo0A~P binding sites known as 0A1, 0A2, and 0A3 box, located in between the two promoters. It remains unclear whether, at the onset of starvation, these boxes activate or repressspo0Aexpression, and whether the Spo0A~P transcriptional feedback plays a role in the increase inspo0Aexpression. Based on the experimental data of the promoter activities under systematic perturbation of the promoter architecture, we developed a biophysical model of transcriptional regulation ofspo0Aby Spo0A~P binding to each of the 0A boxes. The model predicts that Spo0A~P binding to its boxes does not affect the RNAP recruitment to the promoters but instead affects the transcriptional initiation rate. Moreover, the effects of Spo0A~P binding to 0A boxes are mainly repressive and saturated early at the onset of starvation. Therefore, the increase inspo0Aexpression is mainly driven by the increase in RNAP holoenzyme levels. Additionally, we reveal that Spo0A~P affinity to 0A boxes is strongest at 0A3 and weakest at 0A2 and that there are attractive forces between the occupied 0A boxes. Our findings, in addition to clarifying how the sporulation master regulator is controlled, offer a framework to predict regulatory outcomes of complex gene-regulatory mechanisms. IMPORTANCECell differentiation is often critical for survival. In bacteria, differentiation decisions are controlled by transcriptional master regulators under transcriptional feedback control. Therefore, understanding how master regulators are transcriptionally regulated is required to understand differentiation. However, in many cases, the underlying regulation is complex, with multiple transcription factor binding sites and multiple promoters, making it challenging to dissect the exact mechanisms. Here, we address this problem for theBacillus subtilismaster regulator Spo0A. Using a biophysical model, we quantitatively characterize the effect of individual transcription factor binding sites on eachspo0Apromoter. Furthermore, the model allows us to identify the specific transcription step that is affected by transcription factor binding. Such a model is promising for the quantitative study of a wide range of master regulators involved in transcriptional feedback. 
    more » « less
  5. ; ; (, Microorganisms)
    Bacillus subtilis is a soil-dwelling, spore-forming Gram-positive bacterium capable of cell differentiation. For decades, B. subtilis has been used as a model organism to study development of specialized cell types. In this minireview, we discuss cell differentiation in B. subtilis, covering both past research and recent progresses, and the role of cell differentiation in biofilm formation and prevalence of this bacterium in the environment. We review B. subtilis as a classic model for studies of endospore formation, and highlight more recent investigations on cell fate determination and generation of multiple cell types during biofilm formation. We present mechanistic details of how cell fate determination and mutually exclusive cell differentiation are regulated during biofilm formation. 
    more » « less
  • Citation Formats
  • MLA
  • APA
  • Chicago
  • BibTeX
  • Save / Share this Record
  • Send to Email