The Cpx stress response system of Escherichia coli senses plasma membrane proteins and controls HtpX, a membrane protease with a cytosolic active site

2002 ◽  
Vol 7 (7) ◽  
pp. 653-662 ◽  
Author(s):  
Nobuyuki Shimohata ◽  
Shinobu Chiba ◽  
Naoya Saikawa ◽  
Koreaki Ito ◽  
Yoshinori Akiyama
Keyword(s):  
Escherichia Coli ◽  
Plasma Membrane ◽  
Stress Response ◽  
Membrane Proteins ◽  
Active Site ◽  
Plasma Membrane Proteins ◽  
Response System ◽  
Stress Response System

scholarly journals The Cpx Stress Response System Potentiates the Fitness and Virulence of Uropathogenic Escherichia coli

2013 ◽  
Vol 81 (5) ◽  
pp. 1450-1459 ◽  
Author(s):  
Irina Debnath ◽  
J. Paul Norton ◽  
Amelia E. Barber ◽  
Elizabeth M. Ott ◽  
Bijaya K. Dhakal ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Urinary Tract ◽  
Stress Response ◽  
Aminoglycoside Antibiotic ◽  
Deletion Mutants ◽  
Content Type ◽  
Response System ◽  
Stress Response System ◽  
Tract Infections ◽  
Auxiliary Factor

ABSTRACTStrains of uropathogenicEscherichia coli(UPEC) are the primary cause of urinary tract infections, representing one of the most widespread and successful groups of pathogens on the planet. To colonize and persist within the urinary tract, UPEC must be able to sense and respond appropriately to environmental stresses, many of which can compromise the bacterial envelope. The Cpx two-component envelope stress response system is comprised of the inner membrane histidine kinase CpxA, the cytosolic response regulator CpxR, and the periplasmic auxiliary factor CpxP. Here, by using deletion mutants along with mouse and zebrafish infection models, we show that the Cpx system is critical to the fitness and virulence of two reference UPEC strains, the cystitis isolate UTI89 and the urosepsis isolate CFT073. Specifically, deletion of thecpxRAoperon impaired the ability of UTI89 to colonize the murine bladder and greatly reduced the virulence of CFT073 during both systemic and localized infections within zebrafish embryos. These defects coincided with diminished host cell invasion by UTI89 and increased sensitivity of both strains to complement-mediated killing and the aminoglycoside antibiotic amikacin. Results obtained with thecpxPdeletion mutants were more complicated, indicating variable strain-dependent and niche-specific requirements for this well-conserved auxiliary factor.

scholarly journals Activation of the Cpx-envelope stress response system promotes tolerance to antibacterials delivered by arginine-rich peptides and aminoglycosides in Escherichia coli

2020 ◽  
Author(s):  
Jakob Frimodt-Møller ◽  
Andreas Koulouktsis ◽  
Godefroid Charbon ◽  
Marit Otterlei ◽  
Peter E. Nielsen ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Stress Response ◽  
Cell Penetrating Peptides ◽  
Resistant Bacteria ◽  
Antimicrobial Compounds ◽  
Inner Membrane ◽  
Response System ◽  
Envelope Stress ◽  
Cell Penetrating ◽  
Stress Response System

AbstractCell penetrating peptides (CPP) are increasingly used for cellular drug delivery in both pro- and eukaryotic cells, and oligoarginines have attracted special attention. However; their mechanism of action, particularly for prokaryotes is still unknown. Arginine-rich CPPs (R-CPP) efficiently delivers the antimicrobial peptide nucleic acid (PNA) into bacteria. Here, we show that resistance to an R-CPP PNA conjugate in Escherichia coli requires multiple genetic modifications and is specific to R-CPP and not to the PNA-part. An integral part of the resistance was the constitutively activated Cpx-envelope stress response system (cpx*), which decreased the cytoplasmic membrane potential and thereby indicates an indirectly energy dependent uptake mechanism. Interestingly, cpx* mutants also showed increased tolerance to aminoglycosides and R-CPP conjugated to a peptide targeting the DNA sliding clamp; i.e., similar uptake in E. coli for these antimicrobial compounds. We speculate that the cpx* phenotype could create an evolutionary opportunity to adapt and evolve in the presence of either compounds.Author summaryThe emergence of multidrug resistant bacteria is raising the need for new classes of antibiotics. Peptide nucleic acids (PNAs) may fill this requirement by their ability to block translation of essential mRNAs and hence inhibit growth. PNA needs conjugation to a delivery peptide (cell penetrating peptide; CPP) to enter the bacteria. Arginine-rich CPPs (CPPR) are receiving a lot of attention for use as delivery vessels. Here, we show, for the first time, CPPR-PNA resistance in Escherichia coli directed towards the delivery peptide. Consequently, resistance also applies to other antimicrobial compounds delivered by the same carrier. An integral part of CPPR resistance is due to a constitutive active Cpx-response system, which leads to a decreased electric potential (ΔΨ) across the inner membrane. The decreased ΔΨ is a result of down-regulation of two aerobic respiratory operons, namely NADH:ubiquinone oxidoreductase complex I and cytochrome bo3 ubiquinol oxidase. The decreased ΔΨ also led to increased tolerance to aminoglycosides. This shows that a (large) negative ΔΨ is important for providing sufficient free energy for membrane translocation of both CPPR and that the inner membrane is the main barrier for entry of both arginine-rich delivery peptides and aminoglycosides.

scholarly journals Induction Kinetics of a Conditional pH Stress Response System in Escherichia coli

2009 ◽  
Vol 393 (2) ◽  
pp. 272-286 ◽  
Author(s):  
Georg Fritz ◽  
Christiane Koller ◽  
Korinna Burdack ◽  
Larissa Tetsch ◽  
Ina Haneburger ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Stress Response ◽  
Response System ◽  
Induction Kinetics ◽  
Ph Stress ◽  
Stress Response System ◽  
Kinetics Of

scholarly journals The Escherichia coli CpxA-CpxR Envelope Stress Response System Regulates Expression of the Porins OmpF and OmpC

2005 ◽  
Vol 187 (16) ◽  
pp. 5723-5731 ◽  
Author(s):  
Eric Batchelor ◽  
Don Walthers ◽  
Linda J. Kenney ◽  
Mark Goulian
Keyword(s):  
Escherichia Coli ◽  
Stress Response ◽  
Response Regulator ◽  
Gene Encoding ◽  
Fluorescent Reporter ◽  
Response System ◽  
Dnase I Footprinting ◽  
Envelope Stress ◽  
Two Component ◽  
Stress Response System

ABSTRACT We performed transposon mutagenesis of a two-color fluorescent reporter strain to identify new regulators of the porin genes ompF and ompC in Escherichia coli. Screening of colonies by fluorescence microscopy revealed numerous mutants that exhibited interesting patterns of porin expression. One mutant harbored an insertion in the gene encoding the histidine kinase CpxA, the sensor for a two-component signaling system that responds to envelope stress. The cpxA mutant exhibited increased transcription of ompC and a very strong decrease in transcription of ompF under conditions in which acetyl phosphate levels were high. Subsequent genetic analysis revealed that this phenotype is dependent on phosphorylation of the response regulator CpxR and that activation of CpxA in wild-type cells results in similar regulation of porin expression. Using DNase I footprinting, we demonstrated that CpxR binds upstream of both the ompF and ompC promoters. It thus appears that two distinct two-component systems, CpxA-CpxR and EnvZ-OmpR, converge at the porin promoters. Within the context of envelope stress, outer membrane beta-barrel proteins have generally been associated with the sigma E pathway. However, at least for the classical porins OmpF and OmpC, our results show that the Cpx envelope stress response system plays a role in regulating their expression.

scholarly journals Toward a Semisynthetic Stress Response System To Engineer Microbial Solvent Tolerance

mBio ◽  
2012 ◽  
Vol 3 (5) ◽  
Author(s):  
Kyle A. Zingaro ◽  
Eleftherios Terry Papoutsakis
Keyword(s):  
Escherichia Coli ◽  
Stress Response ◽  
Heat Shock ◽  
Heat Shock Proteins ◽  
Solvent Tolerance ◽  
Content Type ◽  
Response System ◽  
Wide Range ◽  
Stress Response System ◽  
Shock Proteins

ABSTRACT Strain tolerance to toxic metabolites is an important trait for many biotechnological applications, such as the production of solvents as biofuels or commodity chemicals. Engineering a complex cellular phenotype, such as solvent tolerance, requires the coordinated and tuned expression of several genes. Using combinations of heat shock proteins (HSPs), we engineered a semisynthetic stress response system in Escherichia coli capable of tolerating high levels of toxic solvents. Simultaneous overexpression of the HSPs GrpE and GroESL resulted in a 2-fold increase in viable cells (CFU) after exposure to 5% (vol/vol) ethanol for 24 h. Co-overexpression of GroESL and ClpB on coexisting plasmids resulted in 1,130%, 78%, and 25% increases in CFU after 24 h in 5% ethanol, 1% n-butanol, and 1% i-butanol, respectively. Co-overexpression of GrpE, GroESL, and ClpB on a single plasmid produced 200%, 390%, and 78% increases in CFU after 24 h in 7% ethanol, 1% n-butanol, or 25% 1,2,4-butanetriol, respectively. Overexpression of other autologous HSPs (DnaK, DnaJ, IbpA, and IbpB) alone or in combinations failed to improve tolerance. Expression levels of HSP genes, tuned through inducible promoters and the plasmid copy number, affected the effectiveness of the engineered stress response system. Taken together, these data demonstrate that tuned co-overexpression of GroES, GroEL, ClpB, and GrpE can be engaged to engineer a semisynthetic stress response system capable of greatly increasing the tolerance of E. coli to solvents and provides a starting platform for engineering customized tolerance to a wide variety of toxic chemicals. IMPORTANCE Microbial production of useful chemicals is often limited by the toxicity of desired products, feedstock impurities, and undesired side products. Improving tolerance is an essential step in the development of practical platform organisms for production of a wide range of chemicals. By overexpressing autologous heat shock proteins in Escherichia coli, we have developed a modular semisynthetic stress response system capable of improving tolerance to ethanol, n-butanol, and potentially other toxic solvents. Using this system, we demonstrate that a practical stress response system requires both tuning of individual gene components and a reliable framework for gene expression. This system can be used to seek out new interacting partners to improve the tolerance phenotype and can be used in the development of more robust solvent production strains.

Detergent fractionation with subsequent subtractive suppression hybridization as a tool for identifying genes coding for plasma membrane proteins

2009 ◽  
Vol 18 (6) ◽  
pp. 527-535 ◽  
Author(s):  
Andreas Lange ◽  
Claudia Kistler ◽  
Tanja B. Jutzi ◽  
Alexandr V. Bazhin ◽  
Claus Detlev Klemke ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Membrane Proteins ◽  
Plasma Membrane Proteins ◽  
Subtractive Suppression Hybridization

scholarly journals pH-dependent Cargo Sorting from the Golgi

2011 ◽  
Vol 286 (12) ◽  
pp. 10058-10065 ◽  
Author(s):  
Chunjuan Huang ◽  
Amy Chang
Keyword(s):  
Plasma Membrane ◽  
Membrane Proteins ◽  
Atpase Activity ◽  
Secretory Pathway ◽  
Ph Dependence ◽  
Glucose Deprivation ◽  
Ph Homeostasis ◽  
Plasma Membrane Proteins ◽  
Cytosolic Ph ◽  
Cargo Sorting

The vacuolar proton-translocating ATPase (V-ATPase) plays a major role in organelle acidification and works together with other ion transporters to maintain pH homeostasis in eukaryotic cells. We analyzed a requirement for V-ATPase activity in protein trafficking in the yeast secretory pathway. Deficiency of V-ATPase activity caused by subunit deletion or glucose deprivation results in missorting of newly synthesized plasma membrane proteins Pma1 and Can1 directly from the Golgi to the vacuole. Vacuolar mislocalization of Pma1 is dependent on Gga adaptors although no Pma1 ubiquitination was detected. Proper cell surface targeting of Pma1 was rescued in V-ATPase-deficient cells by increasing the pH of the medium, suggesting that missorting is the result of aberrant cytosolic pH. In addition to mislocalization of the plasma membrane proteins, Golgi membrane proteins Kex2 and Vrg4 are also missorted to the vacuole upon loss of V-ATPase activity. Because the missorted cargos have distinct trafficking routes, we suggest a pH dependence for multiple cargo sorting events at the Golgi.

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