scholarly journals A Colicin M-Type Bacteriocin fromPseudomonas aeruginosaTargeting the HxuC Heme Receptor Requires a Novel Immunity Partner

2018 ◽  
Vol 84 (18) ◽  
Author(s):  
Maarten G. K. Ghequire ◽  
Başak Öztürk
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Membrane Protein ◽  
Receptor Binding ◽  
Binding Domain ◽  
Modular Organization ◽  
Receptor Binding Domain ◽  
Inner Membrane ◽  
Content Type ◽  
Transmembrane Topology ◽  
Inner Membrane Protein

ABSTRACTPyocins are bacteriocins secreted byPseudomonas aeruginosa, and they assist in the colonization of different niches. A major subset of these antibacterial proteins adopt a modular organization characteristic of polymorphic toxins. They include a receptor-binding domain, a segment enabling membrane passage, and a toxin module at the carboxy terminus, which eventually kills the target cells. To protect themselves from their own products, bacteriocin-producing strains express an immunity gene concomitantly with the bacteriocin. We show here that a pyocin equipped with a phylogenetically distinct ColM toxin domain, PaeM4, mediates antagonism against a large set ofP. aeruginosaisolates. Immunity to PaeM4 is provided by the inner membrane protein PmiC, which is equipped with a transmembrane topology not previously described for the ColM family. Given that strains lacking apmiCgene are killed by PaeM4, the presence of such an immunity partner likely is a key criterion for escaping cellular death mediated by PaeM4. The presence of a TonB box in PaeM4 and enhanced bacteriocin activity under iron-poor conditions strongly suggested the targeting of a TonB-dependent receptor. Evaluation of PaeM4 activities against TonB-dependent receptor knockout mutants inP. aeruginosaPAO1 revealed that the heme receptor HxuC (PA1302) serves as a PaeM4 target at the cellular surface. Because other ColM-type pyocins may target the ferrichrome receptor FiuA, our results illustrate the versatility in target recognition conferred by the polymorphic nature of ColM-type bacteriocins.IMPORTANCEThe antimicrobial armamentarium of a bacterium is a major asset for colonizing competitive environments. Bacteriocins comprise a subset of these compounds. Pyocins are an example of such antibacterial proteins produced byPseudomonas aeruginosa, killing otherP. aeruginosastrains. A large group of these molecules show a modular protein architecture that includes a receptor-binding domain for initial target cell attachment and a killer domain. In this study, we have shown that a novel modular pyocin (PaeM4) that kills target bacteria via interference with peptidoglycan assembly takes advantage of the HxuC heme receptor. Cells can protect themselves from killing by the presence of a dedicated immunity partner, an integral inner membrane protein that adopts a transmembrane topology distinct from that of proteins currently known to provide immunity against such toxin activity. Understanding the receptors with which pyocins interact and how immunity to pyocins is achieved is a pivotal step toward the rational design of bacteriocin cocktails for the treatment ofP. aeruginosainfections.

scholarly journals Tail-Anchored Inner Membrane Protein ElaB Increases Resistance to Stress While Reducing Persistence in Escherichia coli

2017 ◽  
Vol 199 (9) ◽  
Author(s):  
Yunxue Guo ◽  
Xiaoxiao Liu ◽  
Baiyuan Li ◽  
Jianyun Yao ◽  
Thomas K. Wood ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Escherichia Coli ◽  
Membrane Proteins ◽  
Membrane Protein ◽  
Stationary Phase ◽  
Transmembrane Domain ◽  
Inner Membrane ◽  
Content Type ◽  
E Coli ◽  
Inner Membrane Protein

ABSTRACT Host-associated bacteria, such as Escherichia coli, often encounter various host-related stresses, such as nutritional deprivation, oxidative stress, and temperature shifts. There is growing interest in searching for small endogenous proteins that mediate stress responses. Here, we characterized the small C-tail-anchored inner membrane protein ElaB in E. coli. ElaB belongs to a class of tail-anchored inner membrane proteins with a C-terminal transmembrane domain but lacking an N-terminal signal sequence for membrane targeting. Proteins from this family have been shown to play vital roles, such as in membrane trafficking and apoptosis, in eukaryotes; however, their role in prokaryotes is largely unexplored. Here, we found that the transcription of elaB is induced in the stationary phase in E. coli and stationary-phase sigma factor RpoS regulates elaB transcription by binding to the promoter of elaB. Moreover, ElaB protects cells against oxidative stress and heat shock stress. However, unlike membrane peptide toxins TisB and GhoT, ElaB does not lead to cell death, and the deletion of elaB greatly increases persister cell formation. Therefore, we demonstrate that disruption of C-tail-anchored inner membrane proteins can reduce stress resistance; it can also lead to deleterious effects, such as increased persistence, in E. coli. IMPORTANCE Escherichia coli synthesizes dozens of poorly understood small membrane proteins containing a predicted transmembrane domain. In this study, we characterized the function of the C-tail-anchored inner membrane protein ElaB in E. coli. ElaB increases resistance to oxidative stress and heat stress, while inactivation of ElaB leads to high persister cell formation. We also demonstrated that the transcription of elaB is under the direct regulation of stationary-phase sigma factor RpoS. Thus, our study reveals that small inner membrane proteins may have important cellular roles during the stress response.

scholarly journals Clostridium difficile Binary Toxin CDT Induces Clustering of the Lipolysis-Stimulated Lipoprotein Receptor into Lipid Rafts

mBio ◽  
2013 ◽  
Vol 4 (3) ◽  
Author(s):  
Panagiotis Papatheodorou ◽  
Daniel Hornuss ◽  
Thilo Nölke ◽  
Sarah Hemmasi ◽  
Jan Castonguay ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Clostridium Difficile ◽  
Lipid Rafts ◽  
Receptor Binding ◽  
Binding Domain ◽  
Binary Toxin ◽  
Lipoprotein Receptor ◽  
Receptor Binding Domain ◽  
Content Type ◽  
Binding Component

ABSTRACT Clostridium difficile is the leading cause of antibiotics-associated diarrhea and pseudomembranous colitis. Hypervirulent C. difficile strains produce the binary actin-ADP-ribosylating toxin CDT (C. difficile transferase), in addition to the Rho-glucosylating toxins A and B. We recently identified the lipolysis-stimulated lipoprotein receptor (LSR) as the host receptor that mediates uptake of CDT into target cells. Here we investigated in H1-HeLa cells, which ectopically express LSR, the influence of CDT on the plasma membrane distribution of the receptor. We found by fluorescence microscopy that the binding component of CDT (CDTb) induces clustering of LSR into subcompartments of the plasma membrane. Detergent extraction of cells treated with CDTb, followed by sucrose gradient fractionation, uncovered accumulation of LSR in detergent-resistant membranes (DRMs) that contained typical marker proteins of lipid rafts. Membrane cholesterol depletion with methyl-β-cyclodextrin inhibited the association of LSR with DRMs upon addition of CDTb. The receptor-binding domain of CDTb also triggered LSR clustering into DRMs. CDTb-triggered clustering of LSR into DRMs could be confirmed in Caco-2 cells. Our data suggest that CDT forces its receptor to cluster into lipid rafts and that oligomerization of the B component might enhance but is not essential for this process. IMPORTANCE C. difficile binary toxin CDT is a member of the iota-like, actin ADP-ribosylating toxin family. The mechanism that mediates endocytic uptake of these toxins still remains elusive. Previous studies highlighted the importance of lipid rafts for oligomerization of the binding component of these toxins and for cell entry. Recently, the host cell receptor for this toxin family, namely, the lipolysis-stimulated lipoprotein receptor (LSR), has been identified. Our study now demonstrates that the binding component of CDT (CDTb) induces clustering of LSR into lipid rafts. Importantly, LSR clustering is efficiently induced also by the receptor-binding domain of CDTb, suggesting that oligomerization of the B component of CDT is not the main trigger of this process. The current work extends our knowledge on the cooperative play between iota-like toxins and their receptor.

scholarly journals Growth Inhibition by External Potassium of Escherichia coli Lacking PtsN (EIIANtr) Is Caused by Potassium Limitation Mediated by YcgO

2016 ◽  
Vol 198 (13) ◽  
pp. 1868-1882 ◽  
Author(s):  
Ravish Sharma ◽  
Tomohiro Shimada ◽  
Vinod K. Mishra ◽  
Suchitra Upreti ◽  
Abhijit A. Sardesai
Keyword(s):  
Escherichia Coli ◽  
Membrane Protein ◽  
Phosphorylation Site ◽  
Phosphotransferase System ◽  
Inner Membrane ◽  
Content Type ◽  
E Coli ◽  
External Potassium ◽  
Inner Membrane Protein ◽  
External K

ABSTRACTThe absence of PtsN, the terminal phosphoacceptor of the phosphotransferase system comprising PtsP-PtsO-PtsN, inEscherichia coliconfers a potassium-sensitive (Ks) phenotype as the external K+concentration ([K+]e) is increased above 5 mM. A growth-inhibitory increase in intracellular K+content, resulting from hyperactivated Trk-mediated K+uptake, is thought to cause this Ks. We provide evidence that the Ksof the ΔptsNmutant is associated with K+limitation. Accordingly, the moderate Ksdisplayed by the ΔptsNmutant was exacerbated in the absence of the Trk and Kup K+uptake transporters and was associated with reduced cellular K+content. Conversely, overproduction of multiple K+uptake proteins suppressed the Ks. Expression of PtsN variants bearing the H73A, H73D, and H73E substitutions of the phosphorylation site histidine of PtsN complemented the Ks. Absence of the predicted inner membrane protein YcgO (also called CvrA) suppressed the Ks, which was correlated with elevated cellular K+content in the ΔptsNmutant, but the ΔptsNmutation did not alter YcgO levels. Heterologous overexpression ofycgOalso led to Ksthat was associated with reduced cellular K+content, exacerbated by the absence of Trk and Kup and alleviated by overproduction of Kup. Our findings are compatible with a model that postulates that Ksin the ΔptsNmutant occurs due to K+limitation resulting from activation of K+efflux mediated by YcgO, which may be additionally stimulated by [K+]e, implicating a role for PtsN (possibly its dephosphorylated form) as an inhibitor of YcgO activity.IMPORTANCEThis study examines the physiological link between the phosphotransferase system comprising PtsP-PtsO-PtsN and K+ion metabolism inE. coli. Studies on the physiological defect that renders anE. colimutant lacking PtsN to be growth inhibited by external K+indicate that growth impairment results from cellular K+limitation that is mediated by YcgO, a predicted inner membrane protein. Additional observations suggest that dephospho-PtsN may inhibit and external K+may stimulate K+limitation mediated by YcgO. It is speculated that YcgO-mediated K+limitation may be an output of a response to certain stresses, which by modulating the phosphotransfer capacity of the PtsP-PtsO-PtsN phosphorelay leads to growth cessation and stress tolerance.

The Pseudomonas aeruginosa type IV pilin receptor binding domain functions as an adhesin for both biotic and abiotic surfaces

2006 ◽  
Vol 60 (3) ◽  
pp. 813-813 ◽  
Author(s):  
Carmen L. Giltner ◽  
Erin J. van Schaik ◽  
Gerald F. Audette ◽  
Dan Kao ◽  
Robert S. Hodges ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Receptor Binding ◽  
Binding Domain ◽  
Receptor Binding Domain ◽  
Type Iv ◽  
Abiotic Surfaces ◽  
Type Iv Pilin

scholarly journals Pore-forming pyocin S5 utilizes the FptA ferripyochelin receptor to kill Pseudomonas aeruginosa

Microbiology ◽  
2014 ◽  
Vol 160 (2) ◽  
pp. 261-269 ◽  
Author(s):  
Ameer Elfarash ◽  
Jozef Dingemans ◽  
Lumeng Ye ◽  
Ahmed Amir Hassan ◽  
Michael Craggs ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Receptor Binding ◽  
Protein Product ◽  
Binding Domain ◽  
Burkholderia Cenocepacia ◽  
Receptor Binding Domain ◽  
Amino Acid Residues ◽  
Gene Encoding ◽  
Siderophore Receptors ◽  
Transposon Mutants

Pyocins are toxic proteins produced by some strains of Pseudomonas aeruginosa that are lethal for related strains of the same species. Some soluble pyocins (S2, S3 and S4) were previously shown to use the pyoverdine siderophore receptors to enter the cell. The P. aeruginosa PAO1 pore-forming pyocin S5 encoding gene (PAO985) was cloned into the expression vector pET15b, and the affinity-purified protein product tested for its killing activity against different P. aeruginosa strains. The results, however, did not show any correlation with a specific ferripyoverdine receptor. To further identify the S5 receptor, transposon mutants were generated. Pooled mutants were exposed to pyocin S5 and the resistant colonies growing in the killing zone were selected. The majority of S5-resistant mutants had an insertion in the fptA gene encoding the receptor for the siderophore pyochelin. Complementation of an fptA transposon mutant with the P. aeruginosa fptA gene in trans restored the sensitivity to S5. In order to define the receptor-binding domain of pyocin S5, two hybrid pyocins were constructed containing different regions from pyocin S5 fused to the C-terminal translocation and DNase killing domains of pyocin S2. Only the protein containing amino acid residues 151 to 300 from S5 showed toxicity, indicating that the pyocin S5 receptor-binding domain is not at the N-terminus of the protein as in other S-type pyocins. Pyocin S5 was, however, unable to kill Burkholderia cenocepacia strains producing a ferripyochelin FptA receptor, nor was the B. cenocepacia fptA gene able to restore the sensitivity of the resistant fptA mutant P. aeruginosa strain.

scholarly journals FoxB of Pseudomonas aeruginosa Functions in the Utilization of the Xenosiderophores Ferrichrome, Ferrioxamine B, and Schizokinen: Evidence for Transport Redundancy at the Inner Membrane

2006 ◽  
Vol 189 (1) ◽  
pp. 284-287 ◽  
Author(s):  
Páraic Ó Cuív ◽  
Damien Keogh ◽  
Paul Clarke ◽  
Michael O'Connell
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Membrane Protein ◽  
Sinorhizobium Meliloti ◽  
Inner Membrane ◽  
Ferrioxamine B ◽  
Inner Membrane Protein

ABSTRACT Expression of the inner membrane protein FoxB (PA2465) of Pseudomonas aeruginosa in mutants of Sinorhizobium meliloti that are defective in the utilization of ferrichrome, ferrioxamine B, and schizokinen resulted in the restoration of siderophore utilization. Mutagenesis of foxB in P. aeruginosa did not abolish siderophore utilization, suggesting that the function is redundant.

scholarly journals The Pseudomonas aeruginosa type IV pilin receptor binding domain functions as an adhesin for both biotic and abiotic surfaces

2006 ◽  
Vol 59 (4) ◽  
pp. 1083-1096 ◽  
Author(s):  
Carmen L. Giltner ◽  
Erin J. van Schaik ◽  
Gerald F. Audette ◽  
Dan Kao ◽  
Robert S. Hodges ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Receptor Binding ◽  
Binding Domain ◽  
Receptor Binding Domain ◽  
Type Iv ◽  
Abiotic Surfaces ◽  
Type Iv Pilin
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