Escherichia coli heat-stable enterotoxin b (STb) in vivo internalization within rat intestinal epithelial cells

Abstract

-Heat-stable enterotoxin b (STb) is a low molecular weight toxin known to bind sulfatide, its receptor. The fate of STb bound to rat intestinal epithelium cells was followed using an anti-toxin gold labeled assay and transmission electron microscopy. The data suggest that STb toxin and the fusion protein maltose binding protein (MBP)-STb were internalized whereas its mutant I41E-M42R with reduced hydrophobicity did not show internalization. There was a significant difference in the mean of gold particles per field between rat intestine incubated with STb or the fusion protein MBP-STb and the negative control consisting of intestine incubated with PBS alone. No subcellular compartment seems to be particularly aimed by the toxin as gold particles were randomly distributed within the cell. Escherichia coli / entérotoxine STb / internalisation 223 Vet. Res. 33 (2002) 223-228

Escherichia coli STb toxin induces apoptosis in intestinal epithelial cell lines

“…Internalization of wild-type and the mutant I41E-M42R characterized by decreased hydrophobicity were compared and revealed diminished uptake of the mutant indicating these amino acids are essential to proper internalization of STb (Labrie et al, 2002). Indeed, site-specific mutagenesis of the residues Phe 37 , Ile 41 and Met 42 demonstrated they are necessary to the binding of STb to its receptor as these mutations resulted in reduced binding (Labrie et al, 2001a), thus supporting the decrease of the uptake of these mutants observed by Labrie et al (2002).…”

“…Indeed, site-specific mutagenesis of the residues Phe 37 , Ile 41 and Met 42 demonstrated they are necessary to the binding of STb to its receptor as these mutations resulted in reduced binding (Labrie et al, 2001a), thus supporting the decrease of the uptake of these mutants observed by Labrie et al (2002). The toxicity of these mutants was evaluated in the rat loop assay revealing decreased toxicity compared to the wildtype.…”

“…The fate of STb following uptake was studied by Labrie et al (2002) in vivo in rat intestinal epithelial cells using an anti-STb gold labeled assay and transmission electron microscopy revealing that STb does not seem to target a particular organelle as gold particles were observed dispersed throughout cells.…”

“…HRT-18 cells are human colon cancer cells used in apoptosis studies . IEC-18 cells are rat ileum epithelial cells and were chosen since in vivo studies on STb are conducted in a rat model (Labrie et al, 2002). The results obtained during our study are presented in the next section.…”

“…Indeed, Labrie et al (2002) demonstrated STb mutants in the C-terminal region, the domain responsible for oligomer formation, exhibited reduced enterotoxicity. Thus, the implication of oligomer formation in the modulation of cell death by STb could be evaluated in a future study and would permit a greater understanding of the structurefunction relationship required for STb toxicity.…”