Primordial mimicry induces morphological change in Escherichia coli.
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ABSTRACT:
SUBMITTER: Lu H
PROVIDER: S-EPMC8752768 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:BACKGROUND:Thymol is a phenolic compound used for its wide spectrum antimicrobial activity. There is a limited understanding of the antimicrobial mechanisms underlying thymol activity. To investigate this, E. coli strain JM109 was exposed to thymol at sub-lethal concentrations and after 16 rounds of exposure, isolates with a 2-fold increased minimal inhibitory concentration (MIC) were recovered (JM109-Thyr). The phenotype was stable after multiple sub-cultures without thymol. RESULTS:Cell morphology studies by scanning electron microscopy (SEM) suggest that thymol renders bacterial cell membranes permeable and disrupts cellular integrity. 1H Nuclear magnetic resonance (NMR) data showed an increase in lactate and the lactic acid family amino acids in the wild type and JM109-Thyr in the presence of thymol, indicating a shift from aerobic respiration to fermentation. Sequencing of JM109-Thyr defined multiple mutations including a stop mutation in the acrR gene resulting in a truncation of the repressor of the AcrAB efflux pump. AcrAB is a multiprotein complex traversing the cytoplasmic and outer membrane, and is involved in antibiotic clearance. CONCLUSIONS:Our data suggests that thymol tolerance in E. coli induces morphological, metabolic and genetic changes to adapt to thymol antimicrobial activity.
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Project description:Uropathogenic Escherichia coli proceed through a complex intracellular developmental pathway that includes multiple morphological changes. During intracellular growth within Toll-like receptor 4-activated superficial bladder epithelial cells, a subpopulation of uropathogenic E. coli initiates SulA-mediated filamentation. In this study, we directly investigated the role of bacterial morphology in the survival of uropathogenic E. coli from killing by phagocytes. We initially determined that both polymorphonuclear neutrophils and macrophages are recruited to murine bladder epithelium at times coincident with extracellular bacillary and filamentous uropathogenic E. coli. We further determined that bacillary uropathogenic E. coli were preferentially destroyed when mixed uropathogenic E. coli populations were challenged with cultured murine macrophages in vitro. Consistent with studies using elliptical-shaped polymers, the initial point of contact between the phagocyte and filamentous uropathogenic E. coli influenced the efficacy of internalization. These findings demonstrate that filamentous morphology provides a selective advantage for uropathogenic E. coli evasion of killing by phagocytes and defines a mechanism for the essential role for SulA during bacterial cystitis. Thus, morphological plasticity can be viewed as a distinct class of mechanism used by bacterial pathogens to subvert host immunity.
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Project description:The peptidoglycan exoskeleton shapes bacteria and protects them against osmotic forces, making its synthesis the target of many current antibiotics. Peptidoglycan precursors are attached to a lipid carrier and flipped from the cytoplasm into the periplasm to be incorporated into the cell wall. In Escherichia coli, this carrier is undecaprenyl phosphate (Und-P), which is synthesized as a diphosphate by the enzyme undecaprenyl pyrophosphate synthase (UppS). E. coli MG1655 exhibits wild-type morphology at all temperatures, but one of our laboratory strains (CS109) was highly aberrant when grown at 42°C. This strain contained mutations affecting the Und-P synthetic pathway genes uppS, ispH, and idi Normal morphology was restored by overexpressing uppS or by replacing the mutant (uppS31) with the wild-type allele. Importantly, moving uppS31 into MG1655 was lethal even at 30°C, indicating that the altered enzyme was highly deleterious, but growth was restored by adding the CS109 versions of ispH and idi Purified UppSW31R was enzymatically defective at all temperatures, suggesting that it could not supply enough Und-P during rapid growth unless suppressor mutations were present. We conclude that cell wall synthesis is profoundly sensitive to changes in the pool of polyisoprenoids and that isoprenoid homeostasis exerts a particularly strong evolutionary pressure.IMPORTANCE Bacterial morphology is determined primarily by the overall structure of the semirigid macromolecule peptidoglycan. Not only does peptidoglycan contribute to cell shape, but it also protects cells against lysis caused by excess osmotic pressure. Because it is critical for bacterial survival, it is no surprise that many antibiotics target peptidoglycan biosynthesis. However, important gaps remain in our understanding about how this process is affected by peptidoglycan precursor availability. Here, we report that a mutation altering the enzyme that synthesizes Und-P prevents cells from growing at high temperatures and that compensatory mutations in enzymes functioning upstream of uppS can reverse this phenotype. The results highlight the importance of Und-P metabolism for maintaining normal cell wall synthesis and shape.
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Project description:Using a synthetic biosensor to couple production of a specific metabolite with cell growth, we spontaneously evolved cells under the selective condition toward the acquisition of genotypes that optimally reallocated cellular resources. Using 3-hydroxypropionic acid (3-HP) production from glycerol in Escherichia coli as a model system, we determined that spontaneous mutations in the conserved regions of proteins involved in global transcriptional regulation altered the expression of several genes associated with central carbon metabolism. Our study provides a new perspective on adaptive laboratory evolution (ALE) using synthetic biosensors, thereby supporting future efforts in metabolic pathway optimization.
2021-08-23 | GSE162594 | GEO
Project description:One of the adaptive strategies for the constantly changing conditions of the environment utilized in bacterial cells involves the condensation of DNA in complex with the DNA-binding protein, Dps. With the use of electron microscopy and electron tomography, we observed several morphologically different types of DNA condensation in dormant Escherichia coli cells, namely: nanocrystalline, liquid crystalline, and the folded nucleosome-like. We confirmed the presence of both Dps and DNA in all of the ordered structures using EDX analysis. The comparison of EDX spectra obtained for the three different ordered structures revealed that in nanocrystalline formation the majority of the Dps protein is tightly bound to nucleoid DNA. The dps-null cells contained only one type of condensed DNA structure, liquid crystalline, thus, differing from those with Dps. The results obtained here shed some light on the phenomenon of DNA condensation in dormant prokaryotic cells and on the general problem of developing a response to stress. We demonstrated that the population of dormant cells is structurally heterogeneous, allowing them to respond flexibly to environmental changes. It increases the ability of the whole bacterial population to survive under extreme stress conditions.
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Project description:Escherichia coli is not considered naturally competent, yet it has homologues of the genes that most competent bacteria use for DNA uptake and processing. In Haemophilus influenzae and Vibrio cholerae, these genes are regulated by the Sxy and cyclic AMP receptor (CRP) proteins. We used microarrays to find out whether similar regulation occurs in E. coli. Expression of sxy strongly induced 63 transcriptional units, 34 of which required CRP for transcriptional activation and had promoter sites resembling the Sxy- and CRP-dependent CRP-S motif previously characterized in H. influenzae. As previously reported, sxy expression also induced the sigma-H regulon. Flagellar operons were downregulated by sxy expression, although motility remained unaffected. The CRP-S regulon included all of E. coli's known competence gene homologues, so we investigated Sxy's effect on competence-associated phenotypes. A sxy knockout reduced both "natural" plasmid transformation and competitive fitness in long-term culture. In addition, expression of plasmid-borne sxy led to production of type IV pilin, the main subunit of the DNA uptake machinery of most bacteria. Although H. influenzae Sxy only weakly activated the E. coli Sxy regulon, induction was dramatically improved when it was coexpressed with its cognate CRP, suggesting that intimate interactions between Sxy and CRP are required for transcriptional activation at CRP-S sites.
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Project description:To cope with fluctuations in their environment, bacteria have evolved multiple adaptive stress responses. One such response is the nitrogen regulation stress response, which allows bacteria, such as Escherichia coli, to cope with and overcome conditions of nitrogen limitation. This response is directed by the two-component system NtrBC, where NtrC acts as the major transcriptional regulator to activate the expression of genes to mount the response. Recently, my colleagues and I showed that NtrC directly regulates the expression of the relA gene, the major (p)ppGpp synthetase in E. coli, coupling the nitrogen regulation stress and stringent responses. As elevated levels of (p)ppGpp have been implicated in the formation of persister cells, here, I investigated whether nitrogen starvation promotes their formation and whether the NtrC-RelA regulatory cascade plays a role. The results reveal that nitrogen-starved E. coli synthesizes (p)ppGpp and forms a higher percentage of persister cells than nonstarved cells and that both NtrC and RelA are important for these processes. This study provides novel insights into how the formation of persisters can be promoted in response to a nutritional stress.IMPORTANCE Bacteria often reside in environments where nutrient availability is scarce; therefore, they have evolved adaptive responses to rapidly cope with conditions of feast and famine. Understanding the mechanisms that underpin the regulation of how bacteria cope with this stress is a fundamentally important question in the wider context of understanding the biology of the bacterial cell and bacterial pathogenesis. Two major adaptive mechanisms to cope with starvation are the nitrogen regulation (ntr) stress and stringent responses. Here, I describe how these bacterial stress responses are coordinated under conditions of nitrogen starvation to promote the formation of antibiotic-tolerant persister cells by elevating levels of the secondary messenger (p)ppGpp.
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