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Mechanical stimuli activate gene expression via a cell envelope stress sensing pathway.

ABSTRACT: Mechanosensitive mechanisms are often used to sense damage to tissue structure, stimulating matrix synthesis and repair. While this kind of mechanoregulatory process is well recognized in eukaryotic systems, it is not known whether such a process occurs in bacteria. In Vibrio cholerae, antibiotic-induced damage to the load-bearing cell wall promotes increased signaling by the two-component system VxrAB, which stimulates cell wall synthesis. Here we show that changes in mechanical stress within the cell envelope are sufficient to stimulate VxrAB signaling in the absence of antibiotics. We applied mechanical forces to individual bacteria using three distinct loading modalities: extrusion loading within a microfluidic device, direct compression and hydrostatic pressure. In all cases, VxrAB signaling, as indicated by a fluorescent protein reporter, was increased in cells submitted to greater magnitudes of mechanical loading, hence diverse forms of mechanical stimuli activate VxrAB signaling. Reduction in cell envelope stiffness following removal of the endopeptidase ShyA led to large increases in cell envelope deformation and substantially increased VxrAB response, further supporting the responsiveness of VxrAB. Our findings demonstrate a mechanosensitive gene regulatory system in bacteria and suggest that mechanical signals may contribute to the regulation of cell wall homeostasis.

SUBMITTER: Harper CE

PROVIDER: S-EPMC10460444 | biostudies-literature | 2023 Aug

REPOSITORIES: biostudies-literature

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Mechanical stimuli activate gene expression via a cell envelope stress sensing pathway.

Harper Christine E CE Zhang Wenyao W Lee Junsung J Shin Jung-Ho JH Keller Megan R MR van Wijngaarden Ellen E Chou Emily E Wang Zhaohong Z Dörr Tobias T Chen Peng P Hernandez Christopher J CJ

Scientific reports 20230826 1

Mechanosensitive mechanisms are often used to sense damage to tissue structure, stimulating matrix synthesis and repair. While this kind of mechanoregulatory process is well recognized in eukaryotic systems, it is not known whether such a process occurs in bacteria. In Vibrio cholerae, antibiotic-induced damage to the load-bearing cell wall promotes increased signaling by the two-component system VxrAB, which stimulates cell wall synthesis. Here we show that changes in mechanical stress within t ...[more]

PMID: 37633922

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Project description:S. aureus possesses 16 two-component systems (TCS), two of which (GraRS & NsaRS) belong to the intramembrane-sensing histidine kinase (IM-HK) family, conserved within the firmicutes. NsaRS has recently been documented as being important for nisin-resistance in S. aureus. In this study we present a detailed characterization of NsaRS and reveal that, as with other IM-HK TCS, it responds to disruptions in cell-wall stability. Analysis using a lacZ reporter-gene fusion demonstrated that nsaRS expression is upregulated by a variety of cell-wall damaging antibiotics, including Phosphomycin, Ampicillin, Nisin and Penicillin G. Additionally; we reveal that NsaRS regulates a downstream transporter, NsaAB, during nisin-induced stress. NsaS mutants also display a 200-fold decreased ability to develop resistance to the cell-wall targetting antibiotic bacitracin. Microarray analysis reveals the transcription of 245 genes is altered in a nsaS mutant, with the vast majority down-regulated. Included within this list are genes involved in transport, drug-resistance, cell-envelope synthesis, transcriptional regulation, amino acid metabolism and virulence. Using ICP-MS we observed a decrease in intracellular divalent metal ions in an nsaS mutant when grown under low affinity conditions. Characterization of cells using electron microscopy reveals that nsaS mutants have alterations in both cell-wall and cell-envelope structure. Finally, a variety of virulence related phenotypes are impaired in nsaS mutants, including biofilm formation, resistance to killing by human macrophages and survival in whole human blood. Thus NsaRS is important in sensing cell-wall instability in S. aureus, and functions to reprogram gene expression to modify cell-envelope architecture, facilitating adaptation and survival. We did four hybridizations for this experiment, including a biological replicate and a dye-swap experiment for each replicate to account for dye-bias. Spots flagged as empty or bad were excluded and the raw data from each slide was normalized using LOWESS method, with background correction. The data from the replicates were combined (using the median value) and a one-sample t-test was performed. The volcano plot was used with a fold change cut-off of >= 2 and a p-value of <0.05, to filter the genes that were differentially expressed.

Project description:While the green production and application of 2D functional nanomaterials, such as graphene flakes, in films for stretchable and wearable technologies is a promising platform for advanced technologies, there are still challenges involved in the processing of the deposited material to improve properties such as electrical conductivity. In applications such as wearable biomedical and flexible energy devices, the widely used flexible and stretchable substrate materials are incompatible with high-temperature processing traditionally employed to improve the electrical properties, which necessitates alternative manufacturing approaches and new steps for enhancing the film functionality. We hypothesize that a mechanical stimulus, in the form of substrate straining, may provide such a low-energy approach for modifying deposited film properties through increased flake packing and reorientation. To this end, graphene flakes were exfoliated using an unexplored combination of ethanol and cellulose acetate butyrate for morphological and percolative electrical characterization prior to application on polydimethylsiloxane (PDMS) substrates as a flexible and stretchable electrically conductive platform. The deposited percolative free-standing films on PDMS were characterized via in situ resistance strain monitoring and surface morphology measurements over numerous strain cycles, with parameters extracted describing the dynamic modulation of the film's electrical properties. A reduction in the film resistance and strain gauge factor was found to correlate with the surface roughness and densification of a sample's (sub)surface and the applied strain. High surface roughness samples exhibited enhanced reduction in resistance as well as increased sensitivity to strain compared to samples with low surface roughness, corresponding to surface smoothing, which is related to the dynamic settling of graphene flakes on the substrate surface. This procedure of incorporating strain as a mechanical stimulus may find application as a manufacturing tool/step for the routine fabrication of stretchable and wearable devices, as a low energy and compatible approach, for enhancing the properties of such devices for either high sensitivity or low sensitivity of electrical resistance to substrate strain.

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Mechanical stimuli activate gene expression via a cell envelope stress sensing pathway.

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Mechanical stimuli activate gene expression via a cell envelope stress sensing pathway.

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