International Journal of Microbiology and Biotechnology

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Staphylococcal Pore-forming Leukotoxins: Opening of Ca2+-activated K+ Channels and Specificity of Membrane Pores in Human Neutrophils

Received: 14 July 2020    Accepted: 27 July 2020    Published: 20 August 2020
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Abstract

Pore-forming toxins are key virulence determinants produced by human bacterial pathogens Staphylococcus aureus, inducing two independent cellular events in neutrophils. Upon a specific binding to membrane receptors, both Panton and Valentin Leukocidin and γ-hemolysin induced an increase of Na+ and K+ fluxes, likely associated to the activation of preexisting ionic channels or to the membrane pores formation. This was investigated by using, spectrofluorometry techniques and, specific molecular probes in human neutrophils. Interestingly, we found that, in the absence of extracellular Ca2+, leukotoxins did form membrane pores, which were large enough to allow a massive entry of ethidium into neutrophils. Simultaneously, sustained Na+ influx and K+ efflux were observed. Another set of experiments carried out in the presence of extracellular Ca2+ did show that, the percentage of pores formed by leukotoxins was significantly, reduced due to the Ca2+ effect to eventually protect cells from lysis. The simultaneous recording of Na+ and K+ movements showed a significant increase of the K+ efflux although, the Na+ influx was reduced. By using potassium channels blockers, we found that, the potassium efflux enhanced by the presence of extracellular Ca2+, was markedly, inhibited in apamin-, charybdotoxin-, tetrodotoxin-, and quinine-pretreatment neutrophils. We also found that, the increase of the K+ efflux was reduced by either, thapsigargin or TMB8, potent blockers of the internal Ca2+ stores depletion. Consequently, we proposed that, the activation of another potassium pathway by leukotoxins, known as Ca2+-activated K+ channels following the Ca2+ stores depletion. Furthermore, potassium channels blockers did not affect ethidium, Na+ and K+ movements, in the absence of extracellular Ca2+. Moreover, in this condition, no monovalent ions movement was recorded, when the pores formation was altered by tetra-ethyl-ammonium. In the present study, we further highlighted the specificity of membrane pores to Na+ and K+ ions when, the pores formation was completely blocked by divalent ions blockers (Ca2+ and Zn2+). Under these conditions, no monovalent ions movement, was recorded although, a significant influx of Ca2+ and Zn2+ was observed after the leukotoxins application. In conclusion, our data provided an evidence that, staphylococcal leukotoxins induced in human neutrophils: 1) the opening of Ca2+-activated K+ channels, only in the presence of 1 mM extracellular Ca2+; 2) the formation of membrane pores, which exhibited a high specificity to monovalent cations and, 3) an influx of sodium, through a tetrodotoxin not-sensitive pathway ruling out the hypothesis that, Na+ channels could be activated by leukotoxins.

DOI 10.11648/j.ijmb.20200503.19
Published in International Journal of Microbiology and Biotechnology (Volume 5, Issue 3, September 2020)
Page(s) 135-151
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This is an Open Access article, distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution and reproduction in any medium or format, provided the original work is properly cited.

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Copyright © The Author(s), 2024. Published by Science Publishing Group

Keywords

Staphylococcal Pore-forming Leukotoxins: Opening of Ca2+-activated K+ Channels and Specificity of Membrane Pores in Human Neutrophils

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  • Department of Biotechnology, Natural and Life Sciences Faculty, Ahmed Ben Bella Oran1-University, Oran, Algeria; Bacteriology Institute of Medical Faculty, Louis Pasteur University, Strasbourg, France

  • Bacteriology Institute of Medical Faculty, Louis Pasteur University, Strasbourg, France

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    Leila Staali, Didier Andre Colin. (2020). Staphylococcal Pore-forming Leukotoxins: Opening of Ca2+-activated K+ Channels and Specificity of Membrane Pores in Human Neutrophils. International Journal of Microbiology and Biotechnology, 5(3), 135-151. https://doi.org/10.11648/j.ijmb.20200503.19

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    Leila Staali; Didier Andre Colin. Staphylococcal Pore-forming Leukotoxins: Opening of Ca2+-activated K+ Channels and Specificity of Membrane Pores in Human Neutrophils. Int. J. Microbiol. Biotechnol. 2020, 5(3), 135-151. doi: 10.11648/j.ijmb.20200503.19

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    Leila Staali, Didier Andre Colin. Staphylococcal Pore-forming Leukotoxins: Opening of Ca2+-activated K+ Channels and Specificity of Membrane Pores in Human Neutrophils. Int J Microbiol Biotechnol. 2020;5(3):135-151. doi: 10.11648/j.ijmb.20200503.19

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  • @article{10.11648/j.ijmb.20200503.19,
      author = {Leila Staali and Didier Andre Colin},
      title = {Staphylococcal Pore-forming Leukotoxins: Opening of Ca2+-activated K+ Channels and Specificity of Membrane Pores in Human Neutrophils},
      journal = {International Journal of Microbiology and Biotechnology},
      volume = {5},
      number = {3},
      pages = {135-151},
      doi = {10.11648/j.ijmb.20200503.19},
      url = {https://doi.org/10.11648/j.ijmb.20200503.19},
      eprint = {https://download.sciencepg.com/pdf/10.11648.j.ijmb.20200503.19},
      abstract = {Pore-forming toxins are key virulence determinants produced by human bacterial pathogens Staphylococcus aureus, inducing two independent cellular events in neutrophils. Upon a specific binding to membrane receptors, both Panton and Valentin Leukocidin and γ-hemolysin induced an increase of Na+ and K+ fluxes, likely associated to the activation of preexisting ionic channels or to the membrane pores formation. This was investigated by using, spectrofluorometry techniques and, specific molecular probes in human neutrophils. Interestingly, we found that, in the absence of extracellular Ca2+, leukotoxins did form membrane pores, which were large enough to allow a massive entry of ethidium into neutrophils. Simultaneously, sustained Na+ influx and K+ efflux were observed. Another set of experiments carried out in the presence of extracellular Ca2+ did show that, the percentage of pores formed by leukotoxins was significantly, reduced due to the Ca2+ effect to eventually protect cells from lysis. The simultaneous recording of Na+ and K+ movements showed a significant increase of the K+ efflux although, the Na+ influx was reduced. By using potassium channels blockers, we found that, the potassium efflux enhanced by the presence of extracellular Ca2+, was markedly, inhibited in apamin-, charybdotoxin-, tetrodotoxin-, and quinine-pretreatment neutrophils. We also found that, the increase of the K+ efflux was reduced by either, thapsigargin or TMB8, potent blockers of the internal Ca2+ stores depletion. Consequently, we proposed that, the activation of another potassium pathway by leukotoxins, known as Ca2+-activated K+ channels following the Ca2+ stores depletion. Furthermore, potassium channels blockers did not affect ethidium, Na+ and K+ movements, in the absence of extracellular Ca2+. Moreover, in this condition, no monovalent ions movement was recorded, when the pores formation was altered by tetra-ethyl-ammonium. In the present study, we further highlighted the specificity of membrane pores to Na+ and K+ ions when, the pores formation was completely blocked by divalent ions blockers (Ca2+ and Zn2+). Under these conditions, no monovalent ions movement, was recorded although, a significant influx of Ca2+ and Zn2+ was observed after the leukotoxins application. In conclusion, our data provided an evidence that, staphylococcal leukotoxins induced in human neutrophils: 1) the opening of Ca2+-activated K+ channels, only in the presence of 1 mM extracellular Ca2+; 2) the formation of membrane pores, which exhibited a high specificity to monovalent cations and, 3) an influx of sodium, through a tetrodotoxin not-sensitive pathway ruling out the hypothesis that, Na+ channels could be activated by leukotoxins.},
     year = {2020}
    }
    

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  • TY  - JOUR
    T1  - Staphylococcal Pore-forming Leukotoxins: Opening of Ca2+-activated K+ Channels and Specificity of Membrane Pores in Human Neutrophils
    AU  - Leila Staali
    AU  - Didier Andre Colin
    Y1  - 2020/08/20
    PY  - 2020
    N1  - https://doi.org/10.11648/j.ijmb.20200503.19
    DO  - 10.11648/j.ijmb.20200503.19
    T2  - International Journal of Microbiology and Biotechnology
    JF  - International Journal of Microbiology and Biotechnology
    JO  - International Journal of Microbiology and Biotechnology
    SP  - 135
    EP  - 151
    PB  - Science Publishing Group
    SN  - 2578-9686
    UR  - https://doi.org/10.11648/j.ijmb.20200503.19
    AB  - Pore-forming toxins are key virulence determinants produced by human bacterial pathogens Staphylococcus aureus, inducing two independent cellular events in neutrophils. Upon a specific binding to membrane receptors, both Panton and Valentin Leukocidin and γ-hemolysin induced an increase of Na+ and K+ fluxes, likely associated to the activation of preexisting ionic channels or to the membrane pores formation. This was investigated by using, spectrofluorometry techniques and, specific molecular probes in human neutrophils. Interestingly, we found that, in the absence of extracellular Ca2+, leukotoxins did form membrane pores, which were large enough to allow a massive entry of ethidium into neutrophils. Simultaneously, sustained Na+ influx and K+ efflux were observed. Another set of experiments carried out in the presence of extracellular Ca2+ did show that, the percentage of pores formed by leukotoxins was significantly, reduced due to the Ca2+ effect to eventually protect cells from lysis. The simultaneous recording of Na+ and K+ movements showed a significant increase of the K+ efflux although, the Na+ influx was reduced. By using potassium channels blockers, we found that, the potassium efflux enhanced by the presence of extracellular Ca2+, was markedly, inhibited in apamin-, charybdotoxin-, tetrodotoxin-, and quinine-pretreatment neutrophils. We also found that, the increase of the K+ efflux was reduced by either, thapsigargin or TMB8, potent blockers of the internal Ca2+ stores depletion. Consequently, we proposed that, the activation of another potassium pathway by leukotoxins, known as Ca2+-activated K+ channels following the Ca2+ stores depletion. Furthermore, potassium channels blockers did not affect ethidium, Na+ and K+ movements, in the absence of extracellular Ca2+. Moreover, in this condition, no monovalent ions movement was recorded, when the pores formation was altered by tetra-ethyl-ammonium. In the present study, we further highlighted the specificity of membrane pores to Na+ and K+ ions when, the pores formation was completely blocked by divalent ions blockers (Ca2+ and Zn2+). Under these conditions, no monovalent ions movement, was recorded although, a significant influx of Ca2+ and Zn2+ was observed after the leukotoxins application. In conclusion, our data provided an evidence that, staphylococcal leukotoxins induced in human neutrophils: 1) the opening of Ca2+-activated K+ channels, only in the presence of 1 mM extracellular Ca2+; 2) the formation of membrane pores, which exhibited a high specificity to monovalent cations and, 3) an influx of sodium, through a tetrodotoxin not-sensitive pathway ruling out the hypothesis that, Na+ channels could be activated by leukotoxins.
    VL  - 5
    IS  - 3
    ER  - 

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