Interaction of cholera toxin with cloned human goblet cells in monolayer culture

Abstract

Exposure of the intestinal mucosa to Vibrio cholerae enterotoxin (CT) results in mucus secretion from intestinal goblet cells. On the other hand, there is evidence that elevation of intracellular adenosine 3',5'-cyclic monophosphate levels is not sufficient to induce rapid mucin secretion. To determine whether CT has direct effects on human goblet cells and whether CT alone can elicit rapid exocytosis of apical mucin granules, purified CT was applied to monolayer cultures of well-differentiated HT-29-18 N2 cells, a goblet cell subclone derived from the human colon carcinoma line HT-29. CT bound with high affinity (dissociation constant = 10.5 +/- 1.9 nM) to receptors on these cells (approximately 60,000/apical membrane). Binding of radiolabeled CT was inhibited by excess CT or B subunit but not by A subunit. Preincubation of goblet cells with CT blocked the subsequent CT-specific ribosylation of a 45-kDa protein in membrane fractions of the cells and increased the activity of adenylate cyclase by 2- and 10-fold after 1 and 20 h, respectively. Light micrographs revealed that goblet cells incubated with CT, like control cells, contained abundant apical mucin granules. In contrast, goblet cells incubated with Ca2+ inophore A23187 and phorbol ester were rapidly depleted of mucin granules. Thus CT has direct physiological effects on HT-29 goblet cells, but these do not lead to rapid mucin secretion. These results raise the possibility that CT may accelerate mucin secretion in intact mucosa by an indirect mechanism perhaps mediated by mucosal nerves or other cell types.

Roles ofVibrio fischeriand Nonsymbiotic Bacteria in the Dynamics of Mucus Secretion during Symbiont Colonization of theEuprymna scolopesLight Organ

“…Previous studies of the colonization process have indicated that only gram-negative bacteria aggregate in the mucus (35) and that only V. fischeri sets up permanent residence in the crypt spaces (24). Taken together, these data indicate that the colonization of the E. scolopes light organ is a process of ever-increasing specificity.…”

“…In pathogenic associations, bacteria and bacterial products, such as cell surface molecules and toxins, have been implicated in the control of mucus secretion at both the cellular and molecular levels (12). For example, in the pathogenesis of Vibrio cholerae, cholera toxin triggers mucin release from cultured human goblet cells (16,24) and rat colonic tissue (7). In the lungs of patients suffering from cystic fibrosis, excess mucus secretion leads to creation of an environment that is conducive to persistent colonization by Pseudomonas aeruginosa (5,27).…”

Aberrant Expression of Human Mucin GeneMUC5B in Gastric Carcinoma and Cancer Cells

“…It contains an active TATA box, binds ATF-1 and Sp1 transcription factors, and is activated by cAMP-dependent protein kinase and PKC signaling pathways. In this report we showed that CTA, which has already been shown to activate mucin secretion in colon cancer cell (43), is capable of specifically inducing MUC5B promoter activity in KATO-III but not in AGS cells. Thus, we can postulate that MUC5B promoter activation via cAMP signaling in KATO-III cells involves activation of adenyl cyclase through activation of G s regulatory proteins.…”

Two Atypical Enteropathogenic Escherichia coli Strains Induce the Production of Secreted and Membrane-Bound Mucins To Benefit Their Own Growth at the Apical Surface of Human Mucin-Secreting Intestinal HT29-MTX Cells

“…Vibrio cholerae enterotoxin, by increasing intracellular adenosine 3Ј,5Ј-cyclic monophosphate, increases mucus secretion from intestinal goblet cells both in vitro and in vivo (14,32,40,41). The wild-type L. monocytogenes strain EGD increases mucus secretion in vivo (63), with the result that listeriolysin O toxin promotes the increased production of secreted and membrane-bound mucins and the upregulation of MUC genes in cultured mucin-secreting HT29-MTX cells (6,7,47).…”