scholarly journals Chemical or genetic alteration of proton motive force results in loss of virulence of Burkholderia glumae , the cause of rice bacterial panicle blight.

2021 ◽  
Author(s):  
Asif Iqbal ◽  
Pradip R. Panta ◽  
John Ontoy ◽  
Jobelle Bruno ◽  
Jong Hyun Ham ◽  
...  
Keyword(s):  
Sodium Bicarbonate ◽  
Wild Type Strain ◽  
Proton Motive Force ◽  
Membrane Transporters ◽  
Motive Force ◽  
Wild Type ◽  
Bacterial Genomes ◽  
Burkholderia Glumae ◽  
Sheath Rot ◽  
Panicle Blight

Rice is an important source of food for more than half the world’s population. Bacterial panicle blight (BPB) is a disease of rice characterized by grain discoloration or sheath rot caused mainly by Burkholderia glumae . B. glumae synthesizes toxoflavin, an essential virulence factor, that is required for symptoms of the disease. The products of the tox operons, ToxABCDE and ToxFGHI, are responsible for the synthesis and the proton motive force (PMF)-dependent secretion of toxoflavin, respectively. The DedA family is a highly conserved membrane protein family found in most bacterial genomes that likely function as membrane transporters. Our previous work has demonstrated that absence of certain DedA family members results in pleiotropic effects, impacting multiple pathways that are energized by PMF. We have demonstrated that a member of the DedA family from Burkholderia thailandensis , named DbcA, is required for the extreme polymyxin resistance observed in this organism. B. glumae encodes a homolog of DbcA with 73% amino acid identity to Burkholderia thailandensis DbcA. Here, we created and characterized a B. glumae Δ dbcA strain. In addition to polymyxin sensitivity, B. glumae Δ dbcA is compromised for virulence in several BPB infection models and secretes only low amounts of toxoflavin (∼15% of wild type levels). Changes in membrane potential in B. glumae Δ dbcA were reproduced in the wild type strain by the addition of sub-inhibitory concentrations of sodium bicarbonate, previously demonstrated to cause disruption of PMF. Sodium bicarbonate inhibited B. glumae virulence in rice suggesting a possible non-toxic chemical intervention for bacterial panicle blight. IMPORTANCE Bacterial panicle blight (BPB) is a disease of rice characterized by grain discoloration or sheath rot caused mainly by Burkholderia glumae . The DedA family is a highly conserved membrane protein family found in most bacterial genomes that likely function as membrane transporters. Here, we constructed a B. glumae mutant with a deletion in a DedA family member named dbcA and report a loss of virulence in models of BPB. Physiological analysis of the mutant shows that the proton motive force is disrupted, leading to reduction of secretion of the essential virulence factor toxoflavin. The mutant phenotypes are reproduced in the virulent wild type strain without an effect on growth using sodium bicarbonate, a nontoxic buffer that has been reported to disrupt the PMF. The results presented here suggest that bicarbonate may be an effective antivirulence agent capable of controlling BPB without imposing an undue burden on the environment.

scholarly journals 4-Chlorobenzoate Uptake in Comamonas sp. Strain DJ-12 Is Mediated by a Tripartite ATP-Independent Periplasmic Transporter

2006 ◽  
Vol 188 (24) ◽  
pp. 8407-8412 ◽  
Author(s):  
Jong-Chan Chae ◽  
Gerben J. Zylstra
Keyword(s):  
Growth Rate ◽  
Gene Cluster ◽  
Type Strain ◽  
Wild Type Strain ◽  
Proton Motive Force ◽  
Significant Inhibition ◽  
Motive Force ◽  
Wild Type ◽  
Knockout Mutant

ABSTRACT The fcb gene cluster involved in the hydrolytic dehalogenation of 4-chlorobenzoate is organized in the order fcbB-fcbA-fcbT1-fcbT2-fcbT3-fcbC in Comamonas sp. strain DJ-12. The genes are operonic and inducible with 4-chloro-, 4-iodo-, and 4-bromobenzoate. The fcbT1, fcbT2, and fcbT3 genes encode a transporter in the secondary TRAP (tripartite ATP-independent periplasmic) family. An fcbT1T2T3 knockout mutant shows a much slower growth rate on 4-chlorobenzoate compared to the wild type. 4-Chlorobenzoate is transported into the wild-type strain five times faster than into the fcbT1T2T3 knockout mutant. Transport of 4-chlorobenzoate shows significant inhibition by 4-bromo-, 4-iodo-, and 4-fluorobenzoate and mild inhibition by 3-chlorobenzoate, 2-chlorobenzoate, 4-hydroxybenzoate, 3-hydroxybenzoate, and benzoate. Uptake of 4-chlorobenzoate is significantly inhibited by ionophores which collapse the proton motive force.

scholarly journals Yersinia enterocolitica Type III Secretion Depends on the Proton Motive Force but Not on the Flagellar Motor Components MotA and MotB

2004 ◽  
Vol 72 (7) ◽  
pp. 4004-4009 ◽  
Author(s):  
Gottfried Wilharm ◽  
Verena Lehmann ◽  
Kristina Krauss ◽  
Beatrix Lehnert ◽  
Susanna Richter ◽  
...  
Keyword(s):  
Yersinia Enterocolitica ◽  
Type Iii Secretion ◽  
Proton Motive Force ◽  
Motive Force ◽  
Host Cells ◽  
Infection Model ◽  
Flagellar Motor ◽  
Wild Type ◽  
Secretion Systems ◽  
Type Iii

ABSTRACT The flagellum is believed to be the common ancestor of all type III secretion systems (TTSSs). In Yersinia enterocolitica, expression of the flagellar TTSS and the Ysc (Yop secretion) TTSS are inversely regulated. We therefore hypothesized that the Ysc TTSS may adopt flagellar motor components in order to use the pathogenicity-related translocon in a drill-like manner. As a prerequisite for this hypothesis, we first tested a requirement for the proton motive force by both systems using the protonophore carbonyl cyanide m-chlorophenylhydrazone (CCCP). Motility as well as type III-dependent secretion of Yop proteins was inhibited by CCCP. We deleted motAB, which resulted in an immotile phenotype. This mutant, however, secreted amounts of Yops to the supernatant comparable to those of the wild type. Translocation of Yops into host cells was also not affected by the motAB deletion. Virulence of the mutant was comparable to that of the wild type in the mouse oral infection model. Thus, the hypothesis that the Ysc TTSS might adopt flagellar motor components was not confirmed. The finding that, in addition to consumption of ATP, Ysc TTSS requires the proton motive force is discussed.

scholarly journals Limitations to photosynthesis by proton motive force-induced photosystem II photodamage

eLife ◽  
2016 ◽  
Vol 5 ◽  
Author(s):  
Geoffry A Davis ◽  
Atsuko Kanazawa ◽  
Mark Aurel Schöttler ◽  
Kaori Kohzuma ◽  
John E Froehlich ◽  
...  
Keyword(s):  
Photosystem Ii ◽  
Proton Motive Force ◽  
Motive Force ◽  
Limiting Factor ◽  
Wild Type ◽  
Recombination Rates ◽  
Atp Production ◽  
Photosynthetic Regulation ◽  
Light Capture

The thylakoid proton motive force (pmf) generated during photosynthesis is the essential driving force for ATP production; it is also a central regulator of light capture and electron transfer. We investigated the effects of elevated pmf on photosynthesis in a library of Arabidopsis thaliana mutants with altered rates of thylakoid lumen proton efflux, leading to a range of steady-state pmf extents. We observed the expected pmf-dependent alterations in photosynthetic regulation, but also strong effects on the rate of photosystem II (PSII) photodamage. Detailed analyses indicate this effect is related to an elevated electric field (Δψ) component of the pmf, rather than lumen acidification, which in vivo increased PSII charge recombination rates, producing singlet oxygen and subsequent photodamage. The effects are seen even in wild type plants, especially under fluctuating illumination, suggesting that Δψ-induced photodamage represents a previously unrecognized limiting factor for plant productivity under dynamic environmental conditions seen in the field.

scholarly journals The FlgN chaperone activates the Na+-driven engine of the Salmonella flagellar protein export apparatus

2021 ◽  
Vol 4 (1) ◽  
Author(s):  
Tohru Minamino ◽  
Miki Kinoshita ◽  
Yusuke V. Morimoto ◽  
Keiichi Namba
Keyword(s):  
Conformational Change ◽  
Environmental Conditions ◽  
Proton Motive Force ◽  
Protein Export ◽  
Motive Force ◽  
Structural Proteins ◽  
Wild Type ◽  
Flagellar Protein

AbstractThe bacterial flagellar protein export machinery consists of a transmembrane export gate complex and a cytoplasmic ATPase complex. The gate complex has two intrinsic and distinct H+-driven and Na+-driven engines to drive the export of flagellar structural proteins. Salmonella wild-type cells preferentially use the H+-driven engine under a variety of environmental conditions. To address how the Na+-driven engine is activated, we analyzed the fliJ(Δ13–24) fliH(Δ96–97) mutant and found that the interaction of the FlgN chaperone with FlhA activates the Na+-driven engine when the ATPase complex becomes non-functional. A similar activation can be observed with either of two single-residue substitutions in FlhA. Thus, it is likely that the FlgN-FlhA interaction generates a conformational change in FlhA that allows it to function as a Na+ channel. We propose that this type of activation would be useful for flagellar construction under conditions in which the proton motive force is severely restricted.

scholarly journals The TonB Dimeric Crystal Structures Do Not Exist In Vivo

mBio ◽  
2010 ◽  
Vol 1 (5) ◽  
Author(s):  
Kathleen Postle ◽  
Kyle A. Kastead ◽  
Michael G. Gresock ◽  
Joydeep Ghosh ◽  
Cheryl D. Swayne
Keyword(s):  
Outer Membrane ◽  
Crystal Structures ◽  
Cytoplasmic Membrane ◽  
Proton Motive Force ◽  
Membrane Transporters ◽  
Motive Force ◽  
Carboxy Terminus ◽  
Tonb System ◽  
Outer Membrane Transporters

ABSTRACTThe TonB system energizes transport of nutrients across the outer membrane ofEscherichia coliusing cytoplasmic membrane proton motive force (PMF) for energy. Integral cytoplasmic membrane proteins ExbB and ExbD appear to harvest PMF and transduce it to TonB. The carboxy terminus of TonB then physically interacts with outer membrane transporters to allow translocation of ligands into the periplasmic space. The structure of the TonB carboxy terminus (residues ~150 to 239) has been solved several times with similar results. Our previous results hinted thatin vitrostructures might not mimic the dimeric conformations that characterize TonBin vivo. To test structural predictions and to identify irreplaceable residues, the entire carboxy terminus of TonB was scanned with Cys substitutions. TonB I232C and N233C, predicted to efficiently form disulfide-linked dimers in the crystal structures, did not do so. In contrast, Cys substitutions positioned at large distances from one another in the crystal structures efficiently formed dimers. Cys scanning identified seven functionally important residues. However, no single residue was irreplaceable. The phenotypes conferred by changes of the seven residues depended on both the specific assay used and the residue substituted. All seven residues were synergistic with one another. The buried nature of the residues in the structures was also inconsistent with these properties. Taken together, these results indicate that the solved dimeric crystal structures of TonB do not exist. The most likely explanation for the aberrant structures is that they were obtained in the absence of the TonB transmembrane domain, ExbB, ExbD, and/or the PMF.IMPORTANCEThe TonB system of Gram-negative bacteria is an attractive target for development of novel antibiotics because of its importance in iron acquisition and virulence. Logically, therefore, the structure of TonB must be accurately understood. TonB functions as a dimerin vivo, and two different but similar crystal structures of the dimeric carboxy-terminal ~90 amino acids gave rise to mechanistic models. Here we demonstrate that the crystal structures, and therefore the models based on them, are not biologically relevant. The idiosyncratic phenotypes conferred by substitutions at the only seven functionally important residues in the carboxy terminus suggest that similar to interaction of cytochromes P450 with numerous substrates, these residues allow TonB to differentially interact with different outer membrane transporters. Taken together, data suggest that TonB is maintained poised between order and disorder by ExbB, ExbD, and the proton motive force (PMF) before energy transduction to the outer membrane transporters.

scholarly journals Membrane Transporters of the Major Facilitator Superfamily Are Essential for Long-Term Maintenance of Phenotypic Tolerance to Multiple Antibiotics in E. coli

2021 ◽  
Author(s):  
Yingkun Wan ◽  
Miaomiao Wang ◽  
Edward Wai Chi Chan ◽  
Sheng Chen
Keyword(s):  
Proton Motive Force ◽  
Membrane Transporters ◽  
Motive Force ◽  
Major Facilitator Superfamily ◽  
Starvation Stress ◽  
E Coli ◽  
Major Facilitator ◽  
Term Maintenance ◽  
Multiple Antibiotics

We recently showed that the antibiotic-tolerant subpopulation of bacteria or persisters actively maintain the transmembrane proton motive force (PMF) to survive starvation stress for a prolonged period. This work further shows that the reason why antibiotic persisters need to maintain PMF is that PMF is required to support a range of efflux or transportation functions.

scholarly journals Deletion ofmtrCin Haemophilus ducreyi Increases Sensitivity to Human Antimicrobial Peptides and Activates the CpxRA Regulon

2011 ◽  
Vol 79 (6) ◽  
pp. 2324-2334 ◽  
Author(s):  
Sherri D. Rinker ◽  
Michael P. Trombley ◽  
Xiaoping Gu ◽  
Kate R. Fortney ◽  
Margaret E. Bauer
Keyword(s):  
Antimicrobial Peptides ◽  
Antimicrobial Peptide ◽  
Resistance Mechanism ◽  
Proton Motive Force ◽  
Human Infection ◽  
Motive Force ◽  
Haemophilus Ducreyi ◽  
Wild Type ◽  
Content Type ◽  
Antimicrobial Peptide Resistance

ABSTRACTHaemophilus ducreyiresists killing by antimicrobial peptides encountered during human infection, including cathelicidin LL-37, α-defensins, and β-defensins. In this study, we examined the role of the proton motive force-dependent multiple transferable resistance (MTR) transporter in antimicrobial peptide resistance inH. ducreyi. We found a proton motive force-dependent effect onH. ducreyi's resistance to LL-37 and β-defensin HBD-3, but not α-defensin HNP-2. Deletion of the membrane fusion protein MtrC renderedH. ducreyimore sensitive to LL-37 and human β-defensins but had relatively little effect on α-defensin resistance. ThemtrCmutant 35000HPmtrCexhibited phenotypic changes in outer membrane protein profiles, colony morphology, and serum sensitivity, which were restored to wild type bytrans-complementation withmtrC. Similar phenotypes were reported in acpxAmutant; activation of the two-component CpxRA regulator was confirmed by showing transcriptional effects on CpxRA-regulated genes in 35000HPmtrC. AcpxRmutant had wild-type levels of antimicrobial peptide resistance; acpxAmutation had little effect on defensin resistance but led to increased sensitivity to LL-37. 35000HPmtrCwas more sensitive than thecpxAmutant to LL-37, indicating that MTR contributed to LL-37 resistance independent of the CpxRA regulon. The CpxRA regulon did not affect proton motive force-dependent antimicrobial peptide resistance; however, 35000HPmtrChad lost proton motive force-dependent peptide resistance, suggesting that the MTR transporter promotes proton motive force-dependent resistance to LL-37 and human β-defensins. This is the first report of a β-defensin resistance mechanism inH. ducreyiand shows that LL-37 resistance inH. ducreyiis multifactorial.

scholarly journals Bacillus subtilis mutants harbouring a single copy of the rRNA operon exhibit severe defects in growth and sporulation

Microbiology ◽  
2010 ◽  
Vol 156 (10) ◽  
pp. 2944-2952 ◽  
Author(s):  
Hideaki Nanamiya ◽  
Makiko Sato ◽  
Kenta Masuda ◽  
Mikiko Sato ◽  
Tetsuya Wada ◽  
...  
Keyword(s):  
Bacillus Subtilis ◽  
Functional Significance ◽  
Wild Type Strain ◽  
Bacterial Species ◽  
Single Copy ◽  
Rrna Operon ◽  
Rrna Genes ◽  
Wild Type ◽  
Bacterial Genomes ◽  
Mutant Strains

The number of copies of rRNA genes in bacterial genomes differs greatly among bacterial species. It is difficult to determine the functional significance of the heterogeneity of each rRNA operon fully due to the existence of multiple rRNA operons and because the sequence heterogeneity among the rRNA genes is extremely low. To overcome this problem, we sequentially deleted the ten rrn operons of Bacillus subtilis and constructed seven mutant strains that each harboured a single rrn operon (either rrnA, B, D, E, I, J or O) in their genome. The growth rates and sporulation frequencies of these mutants were reduced drastically compared with those of the wild-type strain, and this was probably due to decreased levels of ribosomes in the mutants. Interestingly, the ability to sporulate varied significantly among the mutant strains. These mutants have proved to be invaluable in our initial attempts to reveal the functional significance of the heterogeneity of each rRNA operon.

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