scholarly journals Fine-Tuning of the Cpx Envelope Stress Response Is Required for Cell Wall Homeostasis inEscherichia coli

mBio ◽  
2016 ◽  
Vol 7 (1) ◽  
Author(s):  
Antoine Delhaye ◽  
Jean-François Collet ◽  
Géraldine Laloux
Keyword(s):  
Quality Control ◽  
Cell Wall ◽  
Control System ◽  
Stress Response ◽  
Cell Shape ◽  
Protein Quality ◽  
Fine Tuning ◽  
Gram Negative Bacteria ◽  
Gram Negative ◽  
Envelope Stress

ABSTRACTThe envelope of Gram-negative bacteria is an essential compartment that constitutes a protective and permeability barrier between the cell and its environment. The envelope also hosts the cell wall, a mesh-like structure made of peptidoglycan (PG) that determines cell shape and provides osmotic protection. Since the PG must grow and divide in a cell-cycle-synchronized manner, its synthesis and remodeling are tightly regulated. Here, we discovered that PG homeostasis is intimately linked to the levels of activation of the Cpx system, an envelope stress response system traditionally viewed as being involved in protein quality control in the envelope. We first show that Cpx is activated when PG integrity is challenged and that this activation provides protection to cells exposed to antibiotics inhibiting PG synthesis. By rerouting the outer membrane lipoprotein NlpE, a known Cpx activator, to a different envelope subcompartment, we managed to manipulate Cpx activation levels. We found that Cpx overactivation leads to aberrant cellular morphologies, to an increased sensitivity to β-lactams, and to dramatic division and growth defects, consistent with a loss of PG homeostasis. Remarkably, these phenotypes were largely abrogated by the deletion ofldtD, a Cpx-induced gene involved in noncanonical PG cross-linkage, suggesting that this transpeptidase is an important link between PG homeostasis and the Cpx system.Altogether our data show that fine-tuning of an envelope quality control system constitutes an important layer of regulation of the highly organized cell wall structure.IMPORTANCEThe envelope of Gram-negative bacteria is essential for viability. First, it includes the cell wall, a continuous polymer of peptidoglycan (PG) that determines cell morphology and protects against osmotic stress. Moreover, the envelope constitutes a protective barrier between the cell interior and the environment. Therefore, mechanisms called envelope stress response systems (ESRS) exist to monitor and defend envelope integrity against harmful conditions. Cpx is a major ESRS that detects and manages the accumulation of misfolded proteins in the envelope ofEscherichia coli. We found that this protein quality control system also plays a fundamental role in the regulation of PG assembly. Strikingly, the level of Cpx response is critical, as an excessive activation leads to phenotypes associated with a loss of cell wall integrity. Thus, by contributing to PG homeostasis, the Cpx system lies at the crossroads between key processes of bacterial life, including cell shape, growth, division, and antibiotic resistance.

scholarly journals Cell shape and cell-wall organization in Gram-negative bacteria

2008 ◽  
Vol 105 (49) ◽  
pp. 19282-19287 ◽  
Author(s):  
K. C. Huang ◽  
R. Mukhopadhyay ◽  
B. Wen ◽  
Z. Gitai ◽  
N. S. Wingreen
Keyword(s):  
Cell Wall ◽  
Cell Shape ◽  
Gram Negative Bacteria ◽  
Gram Negative ◽  
Cell Wall Organization

scholarly journals Metabolism of long-chain fatty acids affects disulfide bond formation in Escherichia coli and activates envelope stress response pathways as a combat strategy

2020 ◽  
Author(s):  
Kanchan Jaswal ◽  
Megha Shrivastava ◽  
Deeptodeep Roy ◽  
Shashank Agrawal ◽  
Rachna Chaba
Keyword(s):  
Escherichia Coli ◽  
Stress Response ◽  
Disulfide Bond ◽  
Bond Formation ◽  
Redox Status ◽  
Disulfide Bond Formation ◽  
Gram Negative Bacteria ◽  
Gram Negative ◽  
Envelope Stress ◽  
Long Chain

AbstractThe envelope of gram-negative bacteria serves as the first line of defense against environmental insults. Therefore, its integrity is continuously monitored and maintained by several envelope stress response (ESR) systems. Due to its oxidizing environment, the envelope represents an important site for disulfide bond formation. In Escherichia coli, the periplasmic oxidoreductase, DsbA introduces disulfide bonds in substrate proteins and transfers electrons to the inner membrane oxidoreductase, DsbB. Under aerobic conditions, the reduced form of DsbB is re-oxidized by ubiquinone, an electron carrier in the electron transport chain (ETC). Given the critical role of ubiquinone in transferring electrons derived from the oxidation of reduced cofactors, we were intrigued whether metabolic conditions that generate a large number of reduced cofactors render ubiquinone unavailable for disulfide bond formation. To test this, here we investigated the influence of metabolism of long-chain fatty acid (LCFA), an energy-rich carbon source, on the redox state of the envelope. We show that LCFA degradation increases electron flow in the ETC. Further, we find that whereas cells metabolizing LCFAs exhibit several characteristics of insufficient disulfide bond formation, these hallmarks are averted in cells exogenously provided with ubiquinone. Importantly, the ESR pathways, Cpx and σE, are activated by envelope signals generated during LCFA metabolism, and these systems maintain proper disulfide bond formation. We find that σE downregulation hampers disulfide bond formation only in the absence of Cpx, and amongst the two ESR systems, only Cpx senses redox-dependent signal and is induced to a greater extent by LCFAs. Therefore, we argue that Cpx is the primary ESR that senses and maintains envelope redox homeostasis. Taken together, our results demonstrate an intricate relationship between cellular metabolism and disulfide bond formation dictated by ETC and ESR, and provide the basis for examining whether similar mechanisms control envelope redox status in other gram-negative bacteria.Author summaryDisulfide bonds contribute to the folding and stability of many extracytoplasmic proteins in all domains of life. In gram-negative bacteria, including Escherichia coli, disulfide bond formation occurs in the oxidizing environment of the periplasmic space enclosed within the outer and inner membrane layers of the envelope. Because disulfide-bonded proteins are involved in diverse biological processes, bacteria must monitor the envelope redox status and elicit an appropriate response when perturbations occur; however, these mechanisms are not well elucidated. Here, we demonstrated that the metabolism of an energy-rich carbon source, long-chain fatty acid (LCFA) hampers disulfide bond formation in E. coli. An envelope stress response (ESR) system, Cpx, senses this redox imbalance and maintains proper disulfide bond formation. The σE pathway, another ESR system, plays an ancillary role in maintaining redox homeostasis. LCFA metabolism, disulfide bond formation, and ESR systems have independently been implicated in the pathogenesis of several gram-negative bacteria. The present study sets the basis to explore whether LCFA metabolism impacts the virulence of these bacteria by influencing the redox status of their envelope and activation of ESR pathways.

scholarly journals Fine-Tuning of σEActivation Suppresses Multiple Assembly-Defective Mutations inEscherichia coli

2019 ◽  
Vol 201 (11) ◽  
Author(s):  
Elizabeth M. Hart ◽  
Aileen O’Connell ◽  
Kimberly Tang ◽  
Joseph S. Wzorek ◽  
Marcin Grabowicz ◽  
...  
Keyword(s):  
Stress Response ◽  
Cell Viability ◽  
Outer Membrane ◽  
Stress Responses ◽  
Expression Profiles ◽  
Outer Membrane Proteins ◽  
Fine Tuning ◽  
Permeability Barrier ◽  
Gram Negative ◽  
Envelope Stress

ABSTRACTThe Gram-negative outer membrane (OM) is a selectively permeable asymmetric bilayer that allows vital nutrients to diffuse into the cell but prevents toxins and hydrophobic molecules from entering. Functionally and structurally diverse β-barrel outer membrane proteins (OMPs) build and maintain the permeability barrier, making the assembly of OMPs crucial for cell viability. In this work, we characterize an assembly-defective mutant of the maltoporin LamB, LamBG439D. We show that the folding defect of LamBG439Dresults in an accumulation of unfolded substrate that is toxic to the cell when the periplasmic protease DegP is removed. Selection for suppressors of this toxicity identified the novel mutantdegSA323Eallele. The mutant DegSA323Eprotein contains an amino acid substitution at the PDZ/protease domain interface that results in a partially activated conformation of this protein. This activation increases basal levels of downstream σEstress response signaling. Furthermore, the enhanced σEactivity of DegSA323Esuppresses a number of other assembly-defective conditions without exhibiting the toxicity associated with high levels of σEactivity. We propose that the increased basal levels of σEsignaling primes the cell to respond to envelope stress before OMP assembly defects threaten cell viability. This finding addresses the importance of envelope stress responses in monitoring the OMP assembly process and underpins the critical balance between envelope defects and stress response activation.IMPORTANCEGram-negative bacteria, such asEscherichia coli, inhabit a natural environment that is prone to flux. In order to cope with shifting growth conditions and the changing availability of nutrients, cells must be capable of quickly responding to stress. Stress response pathways allow cells to rapidly shift gene expression profiles to ensure survival in this unpredictable environment. Here we describe a mutant that partially activates the σEstress response pathway. The elevated basal level of this stress response allows the cell to quickly respond to overwhelming stress to ensure cell survival.

scholarly journals Recognition of β-Strand Motifs by RseB Is Required for σEActivity in Escherichia coli

2011 ◽  
Vol 193 (22) ◽  
pp. 6179-6186 ◽  
Author(s):  
Adam Kulp ◽  
Meta J. Kuehn
Keyword(s):  
Stress Response ◽  
Stress Responses ◽  
Outer Membrane Proteins ◽  
Harsh Environments ◽  
Gram Negative Bacteria ◽  
Dependent Manner ◽  
Gram Negative ◽  
Stress Response Pathway ◽  
Envelope Stress ◽  
Different Types

Gram-negative bacteria react to misfolded proteins in the envelope through a myriad of different stress response pathways. This cohort of pathways allows the bacteria to specifically respond to different types of damage, and many of these have been discovered to have key roles in the virulence of bacterial pathogens. Misfolded outer membrane proteins (OMPs) are typically recognized by the σEpathway, a highly conserved envelope stress response pathway. We examined the features of misfolded OMPs with respect to their ability to generate envelope stress responses. We determined that the secondary structure, particularly the potential to form β strands, is critical to inducing the σEresponse in an RseB-dependent manner. The sequence of the potential β-strand motif modulates the strength of the σEresponse generated by the constructs. By understanding the details of how such stress response pathways are activated, we can gain a greater understanding of how bacteria survive in harsh environments.

scholarly journals Comparison of the d-Glutamate-Adding Enzymes from Selected Gram-Positive and Gram-Negative Bacteria

1999 ◽  
Vol 181 (17) ◽  
pp. 5395-5401 ◽  
Author(s):  
Ann W. Walsh ◽  
Paul J. Falk ◽  
Jane Thanassi ◽  
Linda Discotto ◽  
Michael J. Pucci ◽  
...  
Keyword(s):  
Cell Wall ◽  
Fusion Proteins ◽  
Early Stage ◽  
Substrate Inhibition ◽  
Biochemical Properties ◽  
Fine Tuning ◽  
Wall Structure ◽  
Gram Negative Bacteria ◽  
Gram Positive ◽  
Gram Negative

ABSTRACT The biochemical properties of the d-glutamate-adding enzymes (MurD) from Escherichia coli, Haemophilus influenzae, Enterococcus faecalis, andStaphylococcus aureus were investigated to detect any differences in the activity of this enzyme between gram-positive and gram-negative bacteria. The genes (murD) that encode these enzymes were cloned into pMAL-c2 fusion vector and overexpressed as maltose-binding protein–MurD fusion proteins. Each fusion protein was purified to homogeneity by affinity to amylose resin. Proteolytic treatments of the fusion proteins with factor Xa regenerated the individual MurD proteins. It was found that these fusion proteins retain d-glutamate-adding activity and haveKm and V max values similar to those of the regenerated MurDs, except for the H. influenzae enzyme. Substrate inhibition by UDP-N-acetylmuramyl-l-alanine, the acceptor substrate, was observed at concentrations greater than 15 and 30 μM for E. coli and H. influenzae MurD, respectively. Such substrate inhibition was not observed with the E. faecalisand S. aureus enzymes, up to a substrate concentration of 1 to 2 mM. In addition, the two MurDs of gram-negative origin were shown to require monocations such as NH4 + and/or K+, but not Na+, for optimal activity, while anions such as Cl− and SO4 2− had no effect on the enzyme activities. The activities of the two MurDs of gram-positive origin, on the other hand, were not affected by any of the ions tested. All four enzymes required Mg2+ for the ligase activity and exhibited optimal activities around pH 8. These differences observed between the gram-positive and gram-negative MurDs indicated that the two gram-negative bacteria may apply a more stringent regulation of cell wall biosynthesis at the early stage of peptidoglycan biosynthesis pathway than do the two gram-positive bacteria. Therefore, the MurD-catalyzed reaction may constitute a fine-tuning step necessary for the gram-negative bacteria to optimally maintain its relatively thin yet essential cell wall structure during all stages of growth.

scholarly journals Gram-positive bacteria cell wall-derived lipoteichoic acid induces inflammatory alveolar bone loss through prostaglandin E production in osteoblasts

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Tsukasa Tominari ◽  
Ayumi Sanada ◽  
Ryota Ichimaru ◽  
Chiho Matsumoto ◽  
Michiko Hirata ◽  
...  
Keyword(s):  
Cell Wall ◽  
Bone Loss ◽  
Alveolar Bone ◽  
Osteoclast Differentiation ◽  
Lipoteichoic Acid ◽  
Alveolar Bone Loss ◽  
Gram Negative Bacteria ◽  
Gram Positive ◽  
Gram Negative ◽  
Gram Positive Bacteria

AbstractPeriodontitis is an inflammatory disease associated with severe alveolar bone loss and is dominantly induced by lipopolysaccharide from Gram-negative bacteria; however, the role of Gram-positive bacteria in periodontal bone resorption remains unclear. In this study, we examined the effects of lipoteichoic acid (LTA), a major cell-wall factor of Gram-positive bacteria, on the progression of inflammatory alveolar bone loss in a model of periodontitis. In coculture of mouse primary osteoblasts and bone marrow cells, LTA induced osteoclast differentiation in a dose-dependent manner. LTA enhanced the production of PGE2 accompanying the upregulation of the mRNA expression of mPGES-1, COX-2 and RANKL in osteoblasts. The addition of indomethacin effectively blocked the LTA-induced osteoclast differentiation by suppressing the production of PGE2. Using ex vivo organ cultures of mouse alveolar bone, we found that LTA induced alveolar bone resorption and that this was suppressed by indomethacin. In an experimental model of periodontitis, LTA was locally injected into the mouse lower gingiva, and we clearly detected alveolar bone destruction using 3D-μCT. We herein demonstrate a new concept indicating that Gram-positive bacteria in addition to Gram-negative bacteria are associated with the progression of periodontal bone loss.

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