Characterization of the micro-environment of Salmonella typhimurium-containing vacuoles within MDCK epithelial cells

Mol Microbiol. 1992 Nov;6(22):3289-97. doi: 10.1111/j.1365-2958.1992.tb02197.x.

Abstract

Salmonella typhimurium has the capacity to enter into and multiply within epithelial cells. During the entire intracellular stage, bacteria are enclosed within a vacuole. To characterize the micro-environment of the bacteria-containing vacuoles, we have used a new method to measure the expression levels of several S. typhimurium genes in intracellular bacteria within Madin-Darby canine kidney (MDCK) epithelial cells. Our study was based on the determination of beta-galactosidase activity derived from lacZ transcriptional fusions using the highly sensitive substrate fluorescein-di-beta-D-galactoside (FDG). Expression of the iroA and mgtB genes (induced by Fe2+ and Mg2+ limitation respectively), and cadA (induced by pH 6.0 in the presence of lysine, with enhanced expression under anaerobiosis) were characterized at different post-infection times. High intracellular expression levels were detected for the iroA and mgtB genes, suggesting that the concentrations of free Fe2+ and Mg2+ in the vacuole may be low. cadA activity was detected only at early post-infection times (4 h), suggesting that the vacuole may have a mild-acidic pH, and oxygen and lysine present at this time. Globally, the results reported indicate that the use of a highly sensitive beta-galactosidase substrate can provide information about the micro-environment within which an intracellular pathogen, such as S. typhimurium, resides.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Aerobiosis
  • Animals
  • Bacterial Proteins / biosynthesis
  • Bacterial Proteins / genetics
  • Cell Line
  • Dogs
  • Epithelium / microbiology*
  • Epithelium / ultrastructure
  • Gene Expression Regulation, Bacterial*
  • Hydrogen-Ion Concentration
  • Iron / physiology
  • Kidney
  • Lysine / metabolism
  • Magnesium / physiology
  • Recombinant Fusion Proteins / biosynthesis
  • Salmonella typhimurium / genetics
  • Salmonella typhimurium / physiology*
  • Vacuoles / microbiology*

Substances

  • Bacterial Proteins
  • Recombinant Fusion Proteins
  • Iron
  • Magnesium
  • Lysine