title: Role of a potential endoplasmic reticulum retention sequence (RDEL) and the Golgi complex in the cytotonic activity of Escherichia coli heat-labile enterotoxin.
abstract: Recent experimental evidence indicates that Escherichia coli heat-labile enterotoxin and the closely related cholera toxin gain access to intracellular target substrates through a brefeldin A-sensitive pathway that may involve retrograde transport through the Golgi-endoplasmic reticulum network. The A subunits of both toxins possess a carboxy-terminal tetrapeptide sequence (KDEL in cholera toxin and RDEL in the heat-labile enterotoxins) that is known to mediate the retention of eukaryotic proteins in the endoplasmic reticulum. To investigate the potential role of the RDEL sequence in the toxic activity of the heat-labile enterotoxin we constructed mutant analogues of the toxin containing single substitutions (RDGL and RDEV) or a reversed sequence (LEDR). The single substitutions had little effect on Chinese hamster ovary cell elongation or the ability to stimulate cAMP accumulation in Caco-2 cells. Reversal of the sequence reduced the ability of the toxin to increase cAMP levels in Caco-2 cells by approximately 60% and decreased the ability to elicit elongation of Chinese hamster ovary cells. The effects of the heat-labile enterotoxin were not diminished in a mutant Chinese hamster ovary cell line (V.24.1) that belongs to the End4 complementation group and possesses a temperature-sensitive block in secretion that correlates directly with the disappearance of the Golgi stacks. Collectively, these findings suggest that the brefeldin A-sensitive process involved in intoxication by the heat-labile enterotoxin does not involve RDEL-dependent retrograde transport of the A subunit through the Golgi-endoplasmic reticulum complex. The results are more consistent with a model of internalization involving translocation of the A subunit from an endosomal or a trans-Golgi network compartment.
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D001427:Bacterial Toxins / Q000235:genetics* / Q000633:toxicity; D001483:Base Sequence; D020126:Brefeldin A; D016466:CHO Cells; D002460:Cell Line; D006224:Cricetinae; D000242:Cyclic AMP / Q000378:metabolism; D003517:Cyclopentanes / Q000494:pharmacology; D004269:DNA, Bacterial / Q000235:genetics; D004721:Endoplasmic Reticulum / Q000187:drug effects; D004768:Enterotoxins / Q000235:genetics* / Q000633:toxicity; D004926:Escherichia coli / Q000187:drug effects / Q000235:genetics* / Q000472:pathogenicity; D029968:Escherichia coli Proteins; D006056:Golgi Apparatus / Q000187:drug effects; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D011487:Protein Conformation
chemical_list: D000900:Anti-Bacterial Agents; D001427:Bacterial Toxins; D003517:Cyclopentanes; D004269:DNA, Bacterial; D004768:Enterotoxins; D029968:Escherichia coli Proteins; D020126:Brefeldin A; C027538:heat-labile enterotoxin, E coli; D000242:Cyclic AMP