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Visualization of vacuolar acidification-induced transcription of genes of pathogens inside macrophages

Natalia Martin-Orozco1, Nicolas Touret, Michael L Zaharik

  • 1Cell Biology Program, Hospital for Sick Children and Department of Biochemistry, University of Toronto, Toronto, Ontario M5G 1X8, Canada.

Molecular Biology of the Cell
|October 28, 2005
PubMed

Insights

Macrophages control pathogens by regulating the phagosomal environment. This study reveals pH and IFN-gamma signaling as key activators of Salmonella PhoP transcription, independent of magnesium levels.

Area of Science:

  • Immunology
  • Microbiology
  • Cell Biology

Background:

  • Macrophages are crucial in host defense against intracellular pathogens like Salmonella.
  • The phagosome's ionic environment plays a role in pathogen control.
  • The PhoPQ regulon in Salmonella is essential for virulence and adaptation within host cells.

Purpose of the Study:

  • To investigate the influence of the phagosomal ionic milieu on macrophage-mediated pathogen control.
  • To elucidate the specific environmental factors within the Salmonella-containing vacuole (SCV) that regulate PhoP transcription.

Main Methods:

  • Utilized digital imaging and flow cytometry to monitor Salmonella enterica Typhimurium phoP promoter activity in live macrophages.
  • Employed nanosensor particles for direct measurement of magnesium ion concentration ([Mg2+]) within the SCV.
  • Investigated the impact of altering SCV pH gradients and inhibiting vacuolar proton pumps on PhoP induction.
  • Assessed the effect of interferon-gamma (IFN-γ) pretreatment on PhoP activation.

Main Results:

  • Early induction of PhoP was unaffected by manipulating Mg2+ levels within the SCV; [Mg2+] was rapidly stabilized around 1 mM.
  • Nramp1 efflux pump activity did not significantly contribute to PhoP regulon activation.
  • Dissipating the SCV pH gradient significantly attenuated PhoP induction.
  • A pH-independent component of PhoP induction was revealed upon vacuolar proton pump inhibition.
  • IFN-γ pretreatment eliminated the pH-independent component but enhanced the overall PhoP response.

Conclusions:

  • PhoP transcription is regulated by at least three distinct activators: a pH-sensitive component in resting macrophages, an IFN-γ-stimulated pH-sensitive component, and a pH-independent component.
  • The phagosomal pH gradient is a critical factor for PhoP activation, while Mg2+ concentration plays a limited role in early induction.
  • IFN-γ modulates PhoP induction through distinct pathways, highlighting complex immune signaling.

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