Location of the enterotoxin gene from Salmonella typhimurium and characterization of the gene products

Abstract

The enterotoxin gene (stn) in Salmonella typhimurium (Q1 strain) was confined to an 800 bp ClaI-EcoRI genomic DNA fragment (pCE3) that coded for two polypeptides (25 and 12 kDa) under the control of the T7 RNA polymerase/promoter system. The appearance of the 25 kDa protein corresponded to the enterotoxic activity, as determined by elongation of Chinese hamster ovary (CHO) cells, fluid accumulation in rabbit intestinal loops, and altered vascular permeability in rabbit skin. The stn gene products (STN) caused an elevation of intracellular cAMP in CHO cells. These values were at control levels in stn mutants devoid of enterotoxicity, and the 25-kDa protein concurrently disappeared. The biological activity of the heat-labile enterotoxin was blocked by GM1 ganglioside and neutralized by affinity-purified antibodies made against cholera toxin. The 12 kDa protein however was not correlated with an enterotoxic response.

Purification and Characterization of Novel Toxin Produced byVibrio choleraeO1

“…A similar observation was made for crude preparations of the CHO cell-elongating toxin produced by various strains of Salmonella enteritidis (1). Studies with enterotoxin produced by Salmonella typhimurium (4,5) established that the toxin does not require a strong homology to CT to elongate CHO cells and cause fluid accumulation in animal models. Although WO7 toxin resembled CT in some of its properties, the N-terminal amino acid sequences of both the subunits failed to show homology with any of the known cholera toxins, thereby suggesting that WO7 toxin is unique to the V. cholerae species.…”

Distribution of “classic” virulence factors amongSalmonellaspp.

“…At the amino acid level homologies were found between Stn and the CT‐A and the CT‐B subunit of the cholera toxin, LT‐1 (the heat labile toxin of E. coli ), exotoxin A of Pseudomonas aeruginosa and the diphtheria toxin of Corynebacterium diphtheriae [38]. Recombinant Stn displays biological properties, like the elongation of Chinese hamster ovary cells and the evocation of a fluid secretory response in ligated rabbit ileal loops, which are similar to those of CT [35,39]. Antibodies raised against CT are capable of neutralizing the biological activity of Stn [39].…”

Enteric bacterial toxins: mechanisms of action and linkage to intestinal secretion.

“…The stn gene has an atypical initiation codon and lacks an optimal upstream ribosomal binding sequence, resulting in inefficient production of the toxin by the organism (69). Expression of the cloned stn gene in E. coli has reproduced all of the biological properties ascribed to crude lysates of Salmonella organisms (59,68,422,446,447). Only minor histologic changes have been reported in intestinal epithelium treated with lysates of clones expressing the Salmonella enterotoxin (Stn) (422).…”

“…Despite the immunologic relatedness of Stn to CT (59,68,422,446,447), Stn lacks subunit structure and the biologically active protein is a single polypeptide (59,69,422). In addition, the predicted amino acid sequence is quite dissimilar from CT or E. coli LT-I, with only limited conservation in two regions including the potential active site of the toxin, which is predicted to be an ADP-ribosylating toxin (69,422).…”