The heat-labile enterotoxin of Escherichia coli binds to polylactosaminoglycan-containing receptors in CaCo-2 human intestinal epithelial cells.
The E. coli type I heat-labile enterotoxin (LT-I) shares considerable functional, structural, and immunological homology with cholera toxin (CT). Although the ganglioside GM1 is the sole receptor for CT, LT-I also appears to utilize additional, unique receptors on intestinal cells not recognized by CT. We characterized this second class of LT-I receptors using the human intestinal epithelial cell line, CaCo-2. CaCo-2 cells bound 8-fold more LT-I than CT, and some of these additional LT-I receptors appeared to be functional, as CT-B only partially inhibited LT-I activity at concentrations that completely inhibited CT activity. Membranes from unlabeled or [3H]galactose-labeled cells were incubated with toxin B subunits and extracted with Triton X-100, and the solubilized toxin B-receptor complexes were immunoabsorbed with anti-B bound to protein A-Sepharose. When organic extracts of the complexes were separated by thin-layer chromatography and overlayed with [125I]toxin, both toxins were found to bind only GM1. Separation of the complexes from [3H]galactose-labeled membranes by sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed a series of galactoproteins specifically recognized by LT-I but not by CT. Similar proteins were detected on Western blots probed with [125I]toxin. LT-I activity on intact cells and binding to membranes and the above galactoproteins were enhanced by neuraminidase treatment even in the presence of CT-B. beta-1,4-Galactosidase and endo-beta-1,4-galactosidase, but not beta-1,3-galactosidase, significantly reduced LT-I binding. LT-I binding to fetuin and transferrin exhibited a similar glycosidase sensitivity.(ABSTRACT TRUNCATED AT 250 WORDS)
Orlandi
P A
PA
Membrane Biochemistry Section, National Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, Maryland 20892.
Critchley
D R
DR
Fishman
P H
PH
D000606:Amino Sugars / Q000032:analysis*; D001427:Bacterial Toxins / Q000378:metabolism* / Q000494:pharmacology; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D002772:Cholera Toxin / Q000378:metabolism / Q000494:pharmacology; D000242:Cyclic AMP / Q000378:metabolism; D004768:Enterotoxins / Q000378:metabolism* / Q000494:pharmacology; D004848:Epithelium / Q000378:metabolism; D004926:Escherichia coli; D029968:Escherichia coli Proteins; D005677:G(M1) Ganglioside / Q000378:metabolism; D005696:Galactosidases / Q000494:pharmacology; D006023:Glycoproteins / Q000378:metabolism; D006162:Guanylate Cyclase / Q000032:analysis / Q000378:metabolism*; D006801:Humans; D007413:Intestinal Mucosa / Q000378:metabolism*; D011134:Polysaccharides / Q000032:analysis*; D000074261:Receptors, Enterotoxin; D050705:Receptors, Guanylate Cyclase-Coupled; D018000:Receptors, Peptide / Q000032:analysis / Q000378:metabolism*
D000606:Amino Sugars; D001427:Bacterial Toxins; D004768:Enterotoxins; D029968:Escherichia coli Proteins; D006023:Glycoproteins; D011134:Polysaccharides; D018000:Receptors, Peptide; C017336:galactoproteins; C039448:lactosaminoglycan; D005677:G(M1) Ganglioside; D002772:Cholera Toxin; C027538:heat-labile enterotoxin, E coli; D000242:Cyclic AMP; D005696:Galactosidases; D006162:Guanylate Cyclase; D000074261:Receptors, Enterotoxin; D050705:Receptors, Guanylate Cyclase-Coupled