scholarly journals Quantitation and Comparison of Phenotypic Heterogeneity Among Single Cells of Monoclonal Microbial Populations

2019 ◽  
Vol 10 ◽  
Author(s):  
Federica Calabrese ◽  
Iryna Voloshynovska ◽  
Florin Musat ◽  
Martin Thullner ◽  
Michael Schlömann ◽  
...  
Keyword(s):  
Single Cells ◽  
Phenotypic Heterogeneity ◽  
Microbial Populations

scholarly journals Quantitative profiling of integrin αvβ3 on single cells with quantum dot labeling to reveal the phenotypic heterogeneity of glioblastoma

Nanoscale ◽  
2019 ◽  
Vol 11 (39) ◽  
pp. 18224-18231 ◽  
Author(s):  
Tingting Wang ◽  
Guang Li ◽  
Dianbing Wang ◽  
Feng Li ◽  
Dong Men ◽  
...  
Keyword(s):  
Quantum Dot ◽  
Single Cells ◽  
Integrin Αvβ3 ◽  
Phenotypic Heterogeneity ◽  
Specific Receptors ◽  
Cell Phenotypes ◽  
Quantitative Profiling

The distribution, localization and density of individual molecules (e.g. drug-specific receptors) on single cells can offer profound information about cell phenotypes.

scholarly journals On the Ecological Significance of Phenotypic Heterogeneity in Microbial Populations Undergoing Starvation

2022 ◽  
Author(s):  
Monika Opalek ◽  
Bogna Smug ◽  
Michael Doebeli ◽  
Dominika Wloch-Salamon
Keyword(s):  
Phenotypic Heterogeneity ◽  
Microbial Populations ◽  
Cell Heterogeneity ◽  
Ecological Significance

Nongenetic cell heterogeneity is present in glucose-starved yeast populations in the form of quiescent (Q) and nonquiescent (NQ) phenotypes. There is evidence that Q cells help the population survive long starvation.

scholarly journals Generation of genetic tools for gauging multiple gene expression at single cell level

2021 ◽  
Author(s):  
Marta Mellini ◽  
Massimiliano Lucidi ◽  
Francesco Imperi ◽  
Paolo Visca ◽  
Livia Leoni ◽  
...  
Keyword(s):  
Gene Expression ◽  
Image Processing ◽  
Single Cell ◽  
Fluorescent Protein ◽  
Single Cells ◽  
Phenotypic Heterogeneity ◽  
Single Cell Level ◽  
Cell Level ◽  
Multiple Gene ◽  
Genetic Tools

Key microbial processes in many bacterial species are heterogeneously expressed in single cells of bacterial populations. However, the paucity of adequate molecular tools for live, real-time monitoring of multiple gene expression at the single cell level has limited the understanding of phenotypic heterogeneity. In order to investigate phenotypic heterogeneity in the ubiquitous opportunistic pathogen Pseudomonas aeruginosa, a genetic tool that allows gauging multiple gene expression at the single cell level has been generated. This tool, named pRGC, consists in a promoter-probe vector for transcriptional fusions that carries three reporter genes coding for the fluorescent proteins mCherry, green fluorescent protein (GFP) and cyan fluorescent protein (CFP). The pRGC vector has been characterized and validated via single cell gene expression analysis of both constitutive and iron-regulated promoters, showing clear discrimination of the three fluorescence signals in single cells of a P. aeruginosa population, without the need of image-processing for spectral crosstalk correction. In addition, two pRGC variants have been generated for either i) integration of the reporter gene cassette into a single neutral site of P. aeruginosa chromosome, that is suitable for long-term experiments in the absence of antibiotic selection, or ii) replication in bacterial genera other than Pseudomonas. The easy-to-use genetic tools generated in this study will allow rapid and cost-effective investigation of multiple gene expression in populations of environmental and pathogenic bacteria, hopefully advancing the understanding of microbial phenotypic heterogeneity. IMPORTANCE Within a bacterial population single cells can differently express some genes, even though they are genetically identical and experience the same chemical and physical stimuli. This phenomenon, known as phenotypic heterogeneity, is mainly driven by gene expression noise and results in the emergence of bacterial sub-populations with distinct phenotypes. The analysis of gene expression at the single cell level has shown that phenotypic heterogeneity is associated with key bacterial processes, including competence, sporulation and persistence. In this study, new genetic tools have been generated that allow easy cloning of up to three promoters upstream of distinct fluorescent genes, making it possible to gauge multiple gene expression at the single cell level by fluorescent microscopy, without the need of advanced image-processing procedures. A proof of concept has been provided by investigating iron-uptake and iron-storage gene expression in response to iron availability in P. aeruginosa.

scholarly journals Nanoscale resolution of microbial fiber degradation in action

2021 ◽  
Author(s):  
Meltem Tatlı ◽  
Sarah Moraïs ◽  
Omar E. Tovar-Herrera ◽  
Yannick Bomble ◽  
Edward A. Bayer ◽  
...  
Keyword(s):  
Bacterial Population ◽  
Single Cells ◽  
Phenotypic Heterogeneity ◽  
Plant Cell Walls ◽  
Plant Fibers ◽  
Fiber Degradation ◽  
Dynamic Growth ◽  
Global Carbon ◽  
Nanoscale Level

AbstractDeconstruction of plant cell walls is imperative to global carbon cycling and sustainability efforts. Selected microbes degrade plant fibers using extremely efficient multi-enzymatic cellulosomes assemblies. Organization of cellulosomes on the bacterial cell surface and their ecological regulation remain elusive. By combining structural methodologies with molecular and biochemical approaches on the canonical Clostridium thermocellum system, we provide an unprecedented view into the in-situ structure and distribution of cellulosomal enzymes while interacting with their cellulosic substrate during fiber degradation. Structural exploration of growing cultures revealed isogenic phenotypic heterogeneity of cellulosome organization on single cells across the bacterial population, suggesting a division-of labor strategy driven by product-dependent dynamics. This study demonstrates how structural biology under near-physiological conditions can be employed to develop ecological hypotheses to understand microbial plant-fiber degradation at the single-cell nanoscale level.One Sentence SummaryThis study contributes critical insights into the in-situ organization of cellulosomes and their cellulosic substrates and provides evidence for phenotypic heterogeneity, with dynamic, growth phase-dependent organization of the fiber-degrading machinery.

scholarly journals Phenotypic heterogeneity is adaptive for microbial populations under starvation

2021 ◽  
Author(s):  
Monika Opalek ◽  
Bogna Smug ◽  
Michael Doebeli ◽  
Dominika Magdalena Wloch-Salamon
Keyword(s):  
Nutrient Recycling ◽  
Phenotypic Heterogeneity ◽  
Microbial Populations ◽  
Lag Phase ◽  
Starvation Period ◽  
Quiescent State ◽  
Bet Hedging ◽  
Quiescent Cells ◽  
Variable Environments ◽  
Over Time

To persist in variable environments populations of microorganisms have to survive periods of starvation and be able to restart cell division in nutrient-rich conditions. Typically, starvation signals initiate a transition to a quiescent state in a fraction of individual cells, while the rest of the cells remain non-quiescent. It is widely believed that, while quiescent cells (Q) help the population to survive long starvation, the non-quiescent cells (NQ) are a side effect of imperfect transition. We analysed regrowth of starved monocultures of Q and NQ cells compared to mixed, heterogeneous cultures in simple and complex starvation environments. Our experiments, as well as mathematical modelling, demonstrate that Q monocultures benefit from better survival during long starvation, and from a shorter lag phase after resupply of rich medium. However, when the starvation period is very short, the NQ monocultures outperform Q and mixed cultures, due to their short lag phase. In addition, only NQ monocultures benefit from complex starvation environments, where nutrient recycling is possible. Our study suggests that phenotypic heterogeneity in starved populations could be a form of bet hedging, which is adaptive when environmental determinants, such as the length of the starvation period, the length of the regrowth phase, and the complexity of the starvation environment vary over time.

scholarly journals Ecological feedback in quorum-sensing microbial populations can induce heterogeneous production of autoinducers

eLife ◽  
2017 ◽  
Vol 6 ◽  
Author(s):  
Matthias Bauer ◽  
Johannes Knebel ◽  
Matthias Lechner ◽  
Peter Pickl ◽  
Erwin Frey
Keyword(s):  
Gene Expression ◽  
Quorum Sensing ◽  
Promoter Activity ◽  
Phenotypic Heterogeneity ◽  
Microbial Populations ◽  
Alternative Mechanism ◽  
Stochastic Gene Expression ◽  
Regulatory Circuits ◽  
Gene Regulatory ◽  
Gene Regulatory Circuits

Autoinducers are small signaling molecules that mediate intercellular communication in microbial populations and trigger coordinated gene expression via ‘quorum sensing’. Elucidating the mechanisms that control autoinducer production is, thus, pertinent to understanding collective microbial behavior, such as virulence and bioluminescence. Recent experiments have shown a heterogeneous promoter activity of autoinducer synthase genes, suggesting that some of the isogenic cells in a population might produce autoinducers, whereas others might not. However, the mechanism underlying this phenotypic heterogeneity in quorum-sensing microbial populations has remained elusive. In our theoretical model, cells synthesize and secrete autoinducers into the environment, up-regulate their production in this self-shaped environment, and non-producers replicate faster than producers. We show that the coupling between ecological and population dynamics through quorum sensing can induce phenotypic heterogeneity in microbial populations, suggesting an alternative mechanism to stochastic gene expression in bistable gene regulatory circuits.

scholarly journals Direct Observation of the Dynamics of Single-Cell Metabolic Activity during Microbial Diauxic Growth

mBio ◽  
2020 ◽  
Vol 11 (2) ◽  
Author(s):  
H. L. O. McClelland ◽  
C. Jones ◽  
L. M. Chubiz ◽  
D. A. Fike ◽  
A. S. Bradley
Keyword(s):  
Single Cell ◽  
Metabolic Activity ◽  
Single Cells ◽  
Indirect Evidence ◽  
Biomedical Science ◽  
Microbial Populations ◽  
Diurnal Changes ◽  
Diauxic Growth ◽  
Content Type ◽  
Regulatory Cascade

ABSTRACT Population-level analyses are rapidly becoming inadequate to answer many of biomedical science and microbial ecology’s most pressing questions. The role of microbial populations within ecosystems and the evolutionary selective pressure on individuals depend fundamentally on the metabolic activity of single cells. Yet, many existing single-cell technologies provide only indirect evidence of metabolic specialization because they rely on correlations between transcription and phenotype established at the level of the population to infer activity. In this study, we take a top-down approach using isotope labels and secondary ion mass spectrometry to track the uptake of carbon and nitrogen atoms from different sources into biomass and directly observe dynamic changes in anabolic specialization at the level of single cells. We investigate the classic microbiological phenomenon of diauxic growth at the single-cell level in the model methylotroph Methylobacterium extorquens. In nature, this organism inhabits the phyllosphere, where it experiences diurnal changes in the available carbon substrates, necessitating an overhaul of central carbon metabolism. We show that the population exhibits a unimodal response to the changing availability of viable substrates, a conclusion that supports the canonical model but has thus far been supported by only indirect evidence. We anticipate that the ability to monitor the dynamics of anabolism in individual cells directly will have important applications across the fields of ecology, medicine, and biogeochemistry, especially where regulation downstream of transcription has the potential to manifest as heterogeneity that would be undetectable with other existing single-cell approaches. IMPORTANCE Understanding how genetic information is realized as the behavior of individual cells is a long-term goal of biology but represents a significant technological challenge. In clonal microbial populations, variation in gene regulation is often interpreted as metabolic heterogeneity. This follows the central dogma of biology, in which information flows from DNA to RNA to protein and ultimately manifests as activity. At present, DNA and RNA can be characterized in single cells, but the abundance and activity of proteins cannot. Inferences about metabolic activity usually therefore rely on the assumption that transcription reflects activity. By tracking the atoms from which they build their biomass, we make direct observations of growth rate and substrate specialization in individual cells throughout a period of growth in a changing environment. This approach allows the flow of information from DNA to be constrained from the distal end of the regulatory cascade and will become an essential tool in the rapidly advancing field of single-cell metabolism.

scholarly journals Crosstalk between guanosine nucleotides regulates cellular heterogeneity in protein synthesis during nutrient limitation

2021 ◽  
Author(s):  
Simon Diez ◽  
Molly Hydorn ◽  
Abigail Whalen ◽  
Jonathan Dworkin
Keyword(s):  
Protein Synthesis ◽  
Bacillus Subtilis ◽  
Nutrient Limitation ◽  
Dynamic Environments ◽  
Phenotypic Heterogeneity ◽  
Cellular Heterogeneity ◽  
Microbial Populations ◽  
Low Protein ◽  
Synthesis Activity

Phenotypic heterogeneity of microbial populations can facilitate survival in dynamic environments by generating sub-populations of cells that may have differential fitness in a future environment. Bacillus subtilis cultures experiencing nutrient limitation contain distinct sub-populations of cells exhibiting either comparatively high or low protein synthesis activity. This heterogeneity requires the production of phosphorylated guanosine nucleotides (pp)ppGpp by three synthases: SasA, SasB, and RelA. Here we show that these enzymes differentially affect this bimodality: RelA and SasB are necessary to generate the sub-population of cells exhibiting low protein synthesis whereas SasA is necessary to generate cells exhibiting comparatively higher protein synthesis. The RelA product (pppGpp) allosterically activates SasB and we find, in contrast, that the SasA product (pGpp) competitively inhibits this activation. Finally, we provide in vivo evidence that this antagonistic interaction mediates the observed heterogeneity in protein synthesis. This work therefore identifies the mechanism underlying phenotypic heterogeneity in the central physiological process of protein synthesis.

Sign in / Sign up

Export Citation Format

Share Document