Comprehensive Transcriptional Profiling of the Cell Envelope Stress Response of Bacillus subtilis
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ABSTRACT:
ORGANISM(S): Bacillus subtilis
PROVIDER: GSE160345 | GEO | 2023/10/28
REPOSITORIES: GEO
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Project description:The integrity of the bacterial cell envelope is essential to sustain life by countering the high turgor pressure of the cell and providing a barrier against chemical insults. In Bacillus subtilis, synthesis of both peptidoglycan and wall teichoic acids requires a common C55 lipid carrier, undecaprenyl-pyrophosphate (UPP), to ferry precursors across the cytoplasmic membrane. The synthesis and recycling of UPP requires a phosphatase to generate the monophosphate form Und-P, which is the substrate for peptidoglycan and wall teichoic acid synthases. Using an optimized CRISPR-dCas9 based transcriptional repression system (CRISPRi), we demonstrate that B. subtilis requires either of two UPP phosphatases, UppP or BcrC, for viability. We show that a third predicted lipid phosphatase (YodM), with homology to diacylglycerol pyrophosphatases, can also support growth when overexpressed. Depletion of UPP phosphatase activity leads to morphological defects consistent with a failure of cell envelope synthesis and strongly activates the M-dependent cell envelope stress response, including bcrC which encodes one of the two UPP phosphatases. These results highlight the utility of an optimized CRISPRi system for investigation of synthetic lethal gene pairs, clarify the nature of the B. subtilis UPP-Pase enzymes, and provide further evidence linking the M regulon to cell envelope homeostasis pathways.
2016-08-12 | GSE85492 | GEO
Project description:The bacterial cell wall has been a celebrated target for antibiotics and holds real promise as a target for the discovery of new chemical matter to surmount pervasive multi-drug resistance among pathogenic bacteria. While the walls of Gram-negative bacteria are composed primarily of peptidoglycan, those of Gram-positives are more substantial and contain, in addition, large amounts of the polymer teichoic acid, covalently attached to peptidoglycan. Wall teichoic acids are a diverse group of phosphate-rich, extracellular polysaccharides that have been largely regarded as ancillary cell surface components. Recently, wall teichoic acid was shown to be essential to the proper rod-shaped cell morphology of the prototype Gram-positive bacterium Bacillus subtilis and an important virulence factor for the human pathogen Staphylococcus aureus. Thus wall teichoic acid synthesis is an intriguing target for the development of new cell wall-active antibiotics. Nevertheless, recent studies have shown that the dispensability of genes encoding teichoic acid biosynthetic enzymes in both B. subtilis and S. aureus is paradoxical and complex. Here, we report here on the discovery of a promoter (PywaC), which is sensitive to lesions in teichoic acid synthesis. Using this promoter we developed a luminescent, cell-based, reporter system to take a chemical-genetic approach to understanding the complexity of wall teichoic acid biogenesis using a large collection of antibiotics of well characterized biological activity. Our results reveal surprising interactions among undecaprenol, peptidoglycan and teichoic acid biosynthesis that help explain the complexity of teichoic acid gene dispensability. Furthermore, the new reporter assay represents an exciting avenue for the discovery of novel antibacterial molecules that impinge broadly on Gram-positive bacterial cell wall biogenesis. Keywords: comparison between depleted and repleted tagD mutant
2008-03-01 | GSE7373 | GEO
Project description:Transcriptional profiling of Bacillus subtilis to lethal heat stress by systematic sampling of bacterial cell suspensions exposed to gradually increasing temperatures. Keywords: Stress response
2008-12-08 | GSE12372 | GEO
Project description:Transcriptional profiling of Bacillus subtilis to lethal heat stress by systematic sampling of bacterial cell suspensions exposed to gradually increasing temperatures. Keywords: Stress response Bacterial cell suspensions exposed to increasing temperatures ranging from 30-62ºC (Cy5 channel) versus 30 degrees C reference (Cy 3 channel), by transfer of the bacterial cell suspension from a 30ºC to a 65ºC water bath; sampling and quenching at 30, 40, 50, 57, 58, 59, 60 and 62ºC; total time to reach 62ºC is ~12 minutes . Biological replicates: 2, independently grown cultures. One replicate per array. Strains B subtilis 168 1A1; B subtilis A163 (food product isolate); B. subtilis MC85 (food product isolate).
2008-12-07 | E-GEOD-12372 | biostudies-arrayexpress
Project description:Human Peptidoglycan Recognition Proteins (PGRPs) kill bacteria, likely by over-activating stress responses in bacteria. To gain insight into the mechanism of PGRP killing of Bacillus subtilis and bacterial defense against PGRP killing, gene expression in B. subtilis treated with a control protein (bovine serum albumin, BSA), human recombinant PGRP (PGLYRP4), gentamicin (aminoglycoside antibiotic), and CCCP (membrane potential decoupler) were compared. Each treatment induced unique and somewhat overlapping pattern of gene expression. PGRP highly increased expression of genes for oxidative and disulfide stress, detoxification and efflux of Cu, As, and Zn, several transporters, repair of damaged proteins and DNA, energy generation, histidine and cysteine synthesis, envelope lysis and remodeling, and other stress responses. PGRP also caused marked decrease in the expression of genes for phosphate uptake and utilization, Fe uptake, and motility. Gene expression microarray in B. subtilis exposed to a human bactericidal innate immunity protein, PGRP, showed induction of oxidative stress response and defense genes, with different expression pattern than B. subtilis exposed to an aminoglycoside antibiotic and a membrane potential decoupler.
2014-07-25 | GSE44212 | GEO
Project description:The general assumption is that when bacteria run out of nutrients they become dormant or form spores. Here we show, using a new technique, that under deep starvation conditions non-sporulating Bacillus subtilis cells do not become dormant but continue to grow. B. subtilis can form (endo)spores and this has been regarded as the principal mechanism through which it survives long periods of nutrient depletion. However, in this study we demonstrate that non-sporulating B. subtilis cells can survive deep starvation conditions for many months. During this period, cells adopt an almost coccoid shape and become tolerant to antibiotics and oxidative stress. Interestingly, these cells appeared to be metabolically active, and transcriptome analyses indicated that their gene-expression profile differs substantially from both stationary phase cells, and exponentially growing cells. Surprisingly, using an inhibitor for cell division, we discovered that these coccoid-like B. subtilis cells are not dormant but actually grow and divide, albeit with a doubling time of ~4 days. It emerged that secreted proteases, allowing acquisition of nutrients from lysed brethren, are essential for this growth mode. In fact, nutrient levels comparable to 10,000 times diluted LB (Lysogeny broth) appeared to be sufficient to sustain this growth. The very slow growth provides an alternative strategy for B. subtilis to survive nutrient depletion and environmental stresses. We propose to call this the oligotrophic growth state. This state might be common among bacterial species to survive deep starvation conditions.
2018-08-01 | GSE102140 | GEO
Project description:Human Peptidoglycan Recognition Proteins (PGRPs) kill bacteria, likely by over-activating stress responses in bacteria. To gain insight into the mechanism of PGRP killing of Bacillus subtilis and bacterial defense against PGRP killing, gene expression in B. subtilis treated with a control protein (bovine serum albumin, BSA), human recombinant PGRP (PGLYRP4), gentamicin (aminoglycoside antibiotic), and CCCP (membrane potential decoupler) were compared. Each treatment induced unique and somewhat overlapping pattern of gene expression. PGRP highly increased expression of genes for oxidative and disulfide stress, detoxification and efflux of Cu, As, and Zn, several transporters, repair of damaged proteins and DNA, energy generation, histidine and cysteine synthesis, envelope lysis and remodeling, and other stress responses. PGRP also caused marked decrease in the expression of genes for phosphate uptake and utilization, Fe uptake, and motility. Gene expression microarray in B. subtilis exposed to a human bactericidal innate immunity protein, PGRP, showed induction of oxidative stress response and defense genes, with different expression pattern than B. subtilis exposed to an aminoglycoside antibiotic and a membrane potential decoupler. B. subtilis was treated with PGRP (human recombinant PGLYRP4), gentamicin, or CCCP (carbonyl cyanide 3-chlorophenylhydrazone). RNA was obtained from each culture and assayed for gene expression using custom whole genome Affymetrix 900513 GeneChip B. subtilis Genome Array. Each experiment was repeated 3 times.
2014-07-25 | E-GEOD-44212 | biostudies-arrayexpress
Project description:Investigation of the whole genome expression level changes in phosphate limited Bacillus subtilis wild-type and delta-phoPR cells Investigation of the whole genome expression level changes of wild-type and delta-phoPR Bacills subtilis cells comparing high and low phosphate medium
2011-07-13 | GSE23164 | GEO
Project description:Abstract of associated manuscript: Daptomycin is the first of a new class of cyclic lipopeptide antibiotics used against multidrug-resistant Gram-positive pathogens. The proposed mechanism of action involves disruption of the functional integrity of the bacterial membrane in a Ca2+-dependent manner. We have used transcriptional profiling to demonstrate that treatment of Bacillus subtilis with daptomycin strongly induces the lia operon including the autoregulatory LiaRS two-component system (homologous to Staphylococcus aureus VraSR). The lia operon protects against daptomycin and deletion of liaH, encoding a phage shock protein A (PspA)-like protein, leads to 3-fold increased susceptibility. Since daptomycin interacts with the membrane, we tested mutants with altered membrane composition for effects on susceptibility. Deletion mutations of mprF (lacking lysyl-phosphatidylglycerol) or des (lipid desaturase) increased daptomycin susceptibility, whereas overexpression of MprF decreased susceptibility. Conversely, depletion of the cell for the anionic lipid phosphatidylglycerol led to increased resistance. Fluorescently-labeled daptomycin localized to the septa and in a helical pattern around the cell envelope and was delocalized upon depletion of phosphatidylglycerol. Together, these results indicate that the daptomycin-Ca2+ complex interacts preferentially with regions enriched in anionic phospholipids and leads to membrane stresses that can be ameliorated by PspA family proteins.
2008-12-11 | GSE13900 | GEO
Project description:Bacillus subtilis encodes seven extracytoplasmic function (ECF) sigma factors. Three (sigma M, sigma W and simga X) mediate responses to cell envelope active antibiotics. The functions of sigma Y, sigma Z, sigma V, and YlaC remain largely unknown, and strong inducers of these sigma factors and their regulons have yet to be defined. Here, we define transcriptomic and phenotypic differences under non-stress conditions between strains carrying deletions in all seven ECF sigma factor genes (Δ7ECF), a sigMWX triple mutant (∆MWX), and the parental 168 strain. Our results identify >80 genes as at least partially dependent on ECF sigma factors and, as expected, most of these are dependent on sigma M, sigma W or sigma X which are active at a significant basal level during growth. Several genes, including the eps operon encoding enzymes for exopolysaccharide (EPS) production, were decreased in expression in Δ7ECF but affected little if at all in ΔMWX. Consistent with this observation, Δ7ECF (but not ∆MWX) showed reduced biofilm formation. Extending previous observations, we also note that ∆MWX is sensitive to a variety of antibiotics and Δ7ECF is either as sensitive as, or slightly more sensitive than, the ΔMWX strain to these stressors. These findings emphasize the overlapping nature of the seven ECF s factor regulons in B. subtilis, confirm that three of these (sigma M, W or X) play the dominant role in conferring intrinsic resistance to antibiotics, and provide initial insights into the roles of the remaining ECF sigma factors.
2010-07-21 | GSE22930 | GEO