scholarly journals The effects of polydisperse crowders on the compaction of the Escherichia coli nucleoid

2019 ◽  
Author(s):  
Da Yang ◽  
Jaana Männik ◽  
Scott T. Retterer ◽  
Jaan Männik
Keyword(s):  
Escherichia Coli ◽  
Cell Growth ◽  
Cell Membrane ◽  
Significant Contribution ◽  
Osmotic Shock ◽  
Dominant Role ◽  
Dna Binding Proteins ◽  
Bacterial Dna ◽  
E Coli ◽  
Cytoplasmic Proteins

ABSTRACTDNA binding proteins, supercoiling, macromolecular crowders, and transient DNA attachments to the cell membrane have all been implicated in the organization of the bacterial chromosome. However, it is unclear what role these factors play in compacting the bacterial DNA into a distinct organelle-like entity, the nucleoid. By analyzing the effects of osmotic shock and mechanical squeezing on Escherichia coli, we show that macromolecular crowders play a dominant role in the compaction of the DNA into the nucleoid. We find that a 30% increase in the crowder concentration from physiological levels leads to a 3-fold decrease in the nucleoid’s volume. The compaction is anisotropic, being higher along the long axes of the cell at low crowding levels. At higher crowding levels the compression becomes isotropic, implying that E. coli nucleoids lack a well-defined backbone. We furthermore show that the compressibility of the nucleoid is not significantly affected by cell growth rates and by prior treatment with rifampicin. The latter results point out that in addition to poly-ribosomes, soluble cytoplasmic proteins have a significant contribution in determining the size of the nucleoid.

scholarly journals Modification by an R determinant for streptomycin of hissd phenotype in a mutant strain of Escherichia coli B

1968 ◽  
Vol 12 (2) ◽  
pp. 109-116 ◽  
Author(s):  
A. M. Molina ◽  
L. Calegari ◽  
G. Conte
Keyword(s):  
Escherichia Coli ◽  
Cell Membrane ◽  
Mutant Strain ◽  
Minimal Medium ◽  
Specific Requirement ◽  
Resistance Level ◽  
E Coli ◽  
Level Characteristic ◽  
Escherichia Coli B

When an R determinant for streptomycin is transferred into a conditionally streptomycin-dependent E. coli B mutant—which requires in minimal medium either histidine or streptomycin—the latter behaves like a histidineless strain. This phenotype modification shows that the repairing action of streptomycin is prevented. The specific requirement of the strain is not now replaced even by streptomycin concentrations up to 10000 µg/ml at which the conditionally streptomycin-dependent mutant could originally grow, and which are well beyond the resistance level characteristic of the R determinant itself. These data seem to suggest that a reduction in permeability of the cell membrane cannot be held responsible for the phenomenon observed.

scholarly journals Staphylococcus aureus MazF Specifically Cleaves a Pentad Sequence, UACAU, Which Is Unusually Abundant in the mRNA for Pathogenic Adhesive Factor SraP

2009 ◽  
Vol 191 (10) ◽  
pp. 3248-3255 ◽  
Author(s):  
Ling Zhu ◽  
Koichi Inoue ◽  
Satoshi Yoshizumi ◽  
Hiroshi Kobayashi ◽  
Yonglong Zhang ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Staphylococcus Aureus ◽  
Cell Growth ◽  
Bioinformatics Analysis ◽  
Human Tissues ◽  
Regulatory Relationship ◽  
E Coli ◽  
Pathogenic Factors ◽  
Inhibit Cell Growth ◽  
Adhesive Factor

ABSTRACT Escherichia coli mRNA interferases, such as MazF and ChpBK, are sequence-specific endoribonucleases encoded by toxin-antitoxin (TA) systems present in its genome. A MazF homologue in Staphylococcus aureus (MazFSa) has been shown to inhibit cell growth when induced in E. coli. Here, we determined the cleavage site for MazFSa with the use of phage MS2 RNA as a substrate and CspA, an RNA chaperone, which prevents the formation of secondary structures in the RNA substrate. MazFSa specifically cleaves the RNA at a pentad sequence, U↓ACAU. Bioinformatics analysis revealed that this pentad sequence is significantly abundant in several genes, including the sraP gene in the S. aureus N315 strain. This gene encodes a serine-rich protein, which is known to play an important role in adhesion of the pathogen to human tissues and thus in endovascular infection. We demonstrated that the sraP mRNA became extremely unstable in comparison with the ompA mRNA only when MazFSa was induced in E. coli. Further bioinformatics analysis indicated that the pentad sequence is also significantly abundant in the mRNAs for all the pathogenic factors in S. aureus. This observation suggests a possible regulatory relationship between the MazEFSa TA module and the pathogenicity in S. aureus.

Contribution of SOS Genes to H2O2-induced Apoptosis-like Death in Escherichia Coli

2021 ◽  
Author(s):  
Heesu Kim ◽  
Dong Gun Lee
Keyword(s):  
Oxidative Stress ◽  
Escherichia Coli ◽  
Treatment Strategies ◽  
Sos Response ◽  
Bacterial Dna ◽  
E Coli ◽  
New Treatment ◽  
Induced Apoptosis ◽  
And Function ◽  
The Relationship

Abstract Hydrogen peroxide (H2O2) is a debriding agent that damages the microbial structure and function by generating various reactive oxygen species (ROS). H2O2-produced hydroxyl radical (OH∙) also exert oxidative stress on microorganisms. The spread of antibiotic resistance in bacteria is a serious issue worldwide, and greater efforts are needed to identify and characterize novel antibacterial mechanisms to develop new treatment strategies. Therefore, this study aimed to clarify the relationship between H2O2 and Escherichia coli and to elucidate a novel antibacterial mechanism(s) of H2O2. Following H2O2 exposure, increased levels of 8-hydroxyldeoxyguanosine and malondialdehyde indicated that H2O2 accelerates oxidation of bacterial DNA and lipids in E. coli. As oxidative damage worsened, the SOS response was triggered. Cell division arrest and resulting filamentation were identified in cells, indicating that LexA was involved in DNA replication. It was also verified that RecA, a representative SOS gene, helps self-cleavage of LexA and acts as a bacterial caspase-like protein. Our findings also showed that dinF is essential to preserve E. coli from H2O2-induced ROS, and furthermore, demonstrated that H2O2-induced SOS response and SOS genes participate differently in guarding E. coli from oxidative stress. As an extreme SOS response is considered apoptosis-like death (ALD) in bacteria, additional experiments were performed to examine the characteristics of ALD. DNA fragmentation and membrane depolarization appeared in H2O2-treated cells, suggesting that H2O2 causes ALD in E. coli. In conclusion, our investigations revealed that ALD is a novel antibacterial mode of action(s) of H2O2 with important contributions from SOS genes.

scholarly journals Proteorhodopsin Overproduction Enhances the Long-Term Viability of Escherichia coli

2019 ◽  
Vol 86 (1) ◽  
Author(s):  
Yizhi Song ◽  
Michaël L. Cartron ◽  
Philip J. Jackson ◽  
Paul A. Davison ◽  
Mark J. Dickman ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Raman Spectroscopy ◽  
Cell Membrane ◽  
Single Cell ◽  
Marine Bacteria ◽  
Membrane Area ◽  
Content Type ◽  
E Coli ◽  
Wide Range

ABSTRACT Genes encoding the photoreactive protein proteorhodopsin (PR) have been found in a wide range of marine bacterial species, reflecting the significant contribution that PR makes to energy flux and carbon cycling in ocean ecosystems. PR can also confer advantages to enhance the ability of marine bacteria to survive periods of starvation. Here, we investigate the effect of heterologously produced PR on the viability of Escherichia coli. Quantitative mass spectrometry shows that E. coli, exogenously supplied with the retinal cofactor, assembles as many as 187,000 holo-PR molecules per cell, accounting for approximately 47% of the membrane area; even cells with no retinal synthesize ∼148,000 apo-PR molecules per cell. We show that populations of E. coli cells containing PR exhibit significantly extended viability over many weeks, and we use single-cell Raman spectroscopy (SCRS) to detect holo-PR in 9-month-old cells. SCRS shows that such cells, even incubated in the dark and therefore with inactive PR, maintain cellular levels of DNA and RNA and avoid deterioration of the cytoplasmic membrane, a likely basis for extended viability. The substantial proportion of the E. coli membrane required to accommodate high levels of PR likely fosters extensive intermolecular contacts, suggested to physically stabilize the cell membrane and impart a long-term benefit manifested as extended viability in the dark. We propose that marine bacteria could benefit similarly from a high PR content, with a stabilized cell membrane extending survival when those bacteria experience periods of severe nutrient or light limitation in the oceans. IMPORTANCE Proteorhodopsin (PR) is part of a diverse, abundant, and widespread superfamily of photoreactive proteins, the microbial rhodopsins. PR, a light-driven proton pump, enhances the ability of the marine bacterium Vibrio strain AND4 to survive and recover from periods of starvation, and heterologously produced PR extends the viability of nutrient-limited Shewanella oneidensis. We show that heterologously produced PR enhances the viability of E. coli cultures over long periods of several weeks and use single-cell Raman spectroscopy (SCRS) to detect PR in 9-month-old cells. We identify a densely packed and consequently stabilized cell membrane as the likely basis for extended viability. Similar considerations are suggested to apply to marine bacteria, for which high PR levels represent a significant investment in scarce metabolic resources. PR-stabilized cell membranes in marine bacteria are proposed to keep a population viable during extended periods of light or nutrient limitation, until conditions improve.

scholarly journals Membrane Association of the Escherichia coli Enterobactin Synthase Proteins EntB/G, EntE, and EntF

2000 ◽  
Vol 182 (6) ◽  
pp. 1768-1773 ◽  
Author(s):  
Feras M. Hantash ◽  
Charles F. Earhart
Keyword(s):  
Escherichia Coli ◽  
Final Stage ◽  
Membrane Fraction ◽  
Osmotic Shock ◽  
Buoyant Density ◽  
Membrane Association ◽  
Cytosolic Proteins ◽  
Outer Membranes ◽  
Cytoplasmic Proteins ◽  
A Minor

ABSTRACT The cytosolic proteins EntE, EntF, and EntB/G, which areEscherichia coli enzymes necessary for the final stage of enterobactin synthesis, are released by osmotic shock. Here, consistent with the idea that cytoplasmic proteins found in shockates have an affinity for membranes, a small fraction of each was found in membrane preparations. Two procedures demonstrated that the enzymes were enriched in a minor membrane fraction of buoyant density intermediate between that of cytoplasmic and outer membranes, providing indirect support for the notion that these proteins have a role in enterobactin excretion as well as synthesis.

scholarly journals In Vivo Modulation of a DnaJ Homolog, CbpA, by CbpM

2007 ◽  
Vol 189 (9) ◽  
pp. 3635-3638 ◽  
Author(s):  
Matthew R. Chenoweth ◽  
Nancy Trun ◽  
Sue Wickner
Keyword(s):  
Escherichia Coli ◽  
Cell Growth ◽  
Stationary Phase ◽  
Specific Inhibitor ◽  
Vitro Activity ◽  
In Vitro Activity ◽  
E Coli ◽  
Hsp70 Chaperone

ABSTRACT CbpA, an Escherichia coli DnaJ homolog, can function as a cochaperone for the DnaK/Hsp70 chaperone system, and its in vitro activity can be modulated by CbpM. We discovered that CbpM specifically inhibits the in vivo activity of CbpA, preventing it from functioning in cell growth and division. Furthermore, we have shown that CbpM interacts with CbpA in vivo during stationary phase, suggesting that the inhibition of activity is a result of the interaction. These results reveal that the activity of the E. coli DnaK system can be regulated in vivo by a specific inhibitor.

In Vitro Antibacterial Activity and Mechanism of Monocaprylin against Escherichia coli and Staphylococcus aureus

2018 ◽  
Vol 81 (12) ◽  
pp. 1988-1996 ◽  
Author(s):  
JIANYU WANG ◽  
MAOMAO MA ◽  
JUN YANG ◽  
LONG CHEN ◽  
PING YU ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Escherichia Coli ◽  
Staphylococcus Aureus ◽  
Antibacterial Activity ◽  
Membrane Potential ◽  
Cell Membrane ◽  
Sodium Benzoate ◽  
Rapid Change ◽  
Potassium Sorbate ◽  
E Coli

ABSTRACT In the present study, the antibacterial activity of monocaprylin in comparison with sodium benzoate and potassium sorbate against Staphylococcus aureus and Escherichia coli was assessed by measuring MIC, MBC, effect of pH on MIC, and incubation temperature on bactericidal efficacy. Results showed that monocaprylin exhibited an excellent antibacterial activity against both strains, with the lowest MIC and MBC of 1.28 mg/mL. A MIC of monocaprylin remained unchanged despite the pH values of culture medium, ranging from 5 to 9, unlike that of potassium sorbate or sodium benzoate. Furthermore, monocaprylin at MBC effectively reduced the population of E. coli and S. aureus by >5.5 log CFU/mL at 25°C within 6 h and decreased E. coli by approximately 5.0 log CFU/mL and S. aureus by 2.9 log CFU/mL at 12 h. The underlying mechanism of monocaprylin was then investigated by measuring β-galactosidase activity, membrane potential, release of cellular contents, scanning electron microscopy, and transmission electron microscopy observations. Results indicated that monocaprylin killed E. coli by the rapid change in permeability and integrity of cell membrane, leading to decline of membrane potential, leakage of nucleic acids and proteins, and ultimately cell membrane disintegration and lysis. On the other hand, monocaprylin might exert its antibacterial activity against S. aureus mainly by diffusing across the cell wall, collapsing the cell membrane, and disturbing the order of intracellular contents. These findings indicated that monocaprylin had better antibacterial ability compared with traditional synthetic preservatives and might be a potential antibacterial additive independent of pH.

scholarly journals Stationary-Phase Regulation of RpoS Translation in Escherichia coli

2005 ◽  
Vol 187 (21) ◽  
pp. 7204-7213 ◽  
Author(s):  
Matthew Hirsch ◽  
Thomas Elliott
Keyword(s):  
Escherichia Coli ◽  
Stationary Phase ◽  
Osmotic Shock ◽  
Transcriptional Response ◽  
Enteric Bacteria ◽  
Protein Activity ◽  
E Coli ◽  
Phase Regulation ◽  
Shine Dalgarno Sequence ◽  
Necessary And Sufficient

ABSTRACT In enteric bacteria, adaptation to a number of different stresses is mediated by the RpoS protein, one of several sigma factors that collectively allow a tailored transcriptional response to environmental cues. Stress stimuli including low temperature, osmotic shock, nutrient limitation, and growth to stationary phase (SP) all result in a substantial increase in RpoS abundance and activity. The mechanism of regulation depends on the specific signal but may occur at the level of transcription, translation, protein activity, or targeted proteolysis. In both Escherichia coli and Salmonella enterica, SP induction of RpoS in rich medium is >30 fold and includes effects on both transcription and translation. Recently, we found that SP control of rpoS transcription in S. enterica involves repression of the major rpoS promoter during exponential phase by the global transcription factor Fis. Working primarily with E. coli, we now show that 24 nucleotides of the rpoS ribosome-binding site (RBS) are necessary and sufficient for a large part of the increase in rpoS translation as cells grow to SP. Genetic evidence points to an essential role for the leader nucleotides just upstream of the Shine-Dalgarno sequence but is conflicted on the question of whether sequence or structure is important. SP regulation of rpoS is conserved between E. coli and S. enterica. When combined with an fis mutation to block transcriptional effects, replacement of the rpoS RBS sequence by the lacZ RBS eliminates nearly all SP induction of RpoS.

scholarly journals Possible epigenetic influence on cellular differentiation and lineage segregation in Escherichia coli

2019 ◽  
Author(s):  
Wenfa Ng
Keyword(s):  
Gene Expression ◽  
Escherichia Coli ◽  
Cell Growth ◽  
Single Cell ◽  
Cell Fate ◽  
Cellular Differentiation ◽  
Environmental Stressors ◽  
Epigenetic Markers ◽  
E Coli ◽  
Growth Chambers

Epigenetics provides the critical connection between environmental influence and gene expression, where environmental stressors could modulate expression of specific genes in particular scenarios using molecular markers etched at the genome level. Hence, epigenetics likely play important roles in potentiating the development of specific lineages, cell fate or cellular differentiation. For example, when specific environmental stressor is present, epigenetic markers in the genome receive a signal for either activating or deactivating expression of particular sets of genes, which may be linked to the developmental trajectory of the organism. Using Escherichia coli as model organism, a possible study may investigate the role of epigenetics in influencing cellular differentiation of the bacterium. Specifically, a single E. coli cell would be propagated into a consortium of 12 or more bacterial cells in a microfluidics growth chamber. Genetic material extracted would be sent for single cell genomics, transcriptomics, and chromatin immunoprecipitation sequencing (ChIP-seq). After profiling, the residual population would be diverted by microchannels to 6 different cell growth chambers, where they would be cultivated under identical conditions for understanding possible triggers to cell differentiation. At suitable time points of 2, 4, 6, 8, 10, 12 hours, single cell would be extracted from each growth chamber for profiling single cell genomics, transcriptomics, and epigenetics markers. Optical and confocal laser scanning microscopy would provide readout of cell morphologies. Comparison of the readout between the original clonal population and those of the different growth chambers may provide important points for correlating epigenetic markers with gene expression and phenotypic readout in cell lineage, fate and differentiation. In subsequent experiments, different environmental stressors such as pH, imbalance nutrient composition between carbon and nitrogen, nanoparticles or heavy metals, could be used as triggers for specific cell growth response guided by epigenetic programmes embedded within the epigenome of the bacterium. Collectively, epigenetics hold influence for cellular differentiation in view of specific environmental stressors, where epigenetic markers on the genome communicate specific environmental factor's effect on the organism through altering expression of particular sets of genes, that result in different cell fate, lineage and differentiation. Using modern single cell techniques at the genomics, transcriptomics and epigenomics level, the study hopes to elucidate epigenetic potentiators of cellular differentiation in E. coli with and without environmental stressors such as nutrient deprivation, pH and toxic metals.

scholarly journals Reproducing bench-scale cell growth and productivity

2018 ◽  
Author(s):  
Ariel Hecht ◽  
James Filliben ◽  
Sarah A. Munro ◽  
Marc Salit
Keyword(s):  
Escherichia Coli ◽  
Cell Growth ◽  
Small Molecule ◽  
Fluorescent Protein ◽  
Fractional Factorial ◽  
Red Fluorescent Protein ◽  
Factorial Experiments ◽  
Minimum Information ◽  
E Coli ◽  
Product Titer

Reproducing, exchanging, comparing, and building on each other’s work is foundational to technology advances.1Advancing biotechnology calls for reliable reuse of engineered strains.2Reliable reuse of engineered strains requires reproducible growth and productivity. To demonstrate reproducibility for biotechnology, we identified the experimental factors that have the greatest effect on the growth and productivity of our engineered strains.3–6We present a draft of a Minimum Information Standard for Engineered Organism Experiments (MIEO) based on this method. We evaluated the effect of 22 factors onEscherichia coli(E. coli) engineered to produce the small molecule lycopene, and 18 factors onE. coliengineered to produce red fluorescent protein (RFP). Container geometry and shaking had the greatest effect on product titer and yield. We reproduced our results under two different conditions of reproducibility:7conditions of use (different fractional factorial experiments), and time (48 biological replicates performed on 12 different days over four months).

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