Cautionary note on the use of the B subunit of cholera toxin as a ganglioside GM1 probe: Detection of cholera toxin A subunit in B subunit preparations by a sensitive adenylate cyclase assay

1990 ◽  
Vol 42 (3) ◽  
pp. 143-152 ◽  
Author(s):  
Sarah Spiegel
Keyword(s):  
Adenylate Cyclase ◽  
Cholera Toxin ◽  
Ganglioside Gm1 ◽  
Cautionary Note ◽  
Toxin A ◽  
Probe Detection ◽  
B Subunit ◽  
A Subunit

Interaction of ganglioside GM1 with the B subunit of cholera toxin modulates intracellular free calcium in sensory neurons

1992 ◽  
Vol 33 (3) ◽  
pp. 466-475 ◽  
Author(s):  
D. Milani ◽  
M.-C. Minozzi ◽  
L. Petrelli ◽  
D. Guidolin ◽  
S. D. Skaper ◽  
...  
Keyword(s):  
Cholera Toxin ◽  
Sensory Neurons ◽  
Intracellular Free Calcium ◽  
Free Calcium ◽  
Ganglioside Gm1 ◽  
B Subunit

scholarly journals Expression and mutagenesis of recombinant cholera toxin A subunit

1994 ◽  
Vol 17 (5) ◽  
pp. 339-346 ◽  
Author(s):  
Kirsten L. Vadheim ◽  
Yogendra Singh ◽  
Jerry M. Keith
Keyword(s):  
Cholera Toxin ◽  
Toxin A ◽  
A Subunit

scholarly journals Differential Biological and Adjuvant Activities of Cholera Toxin and Escherichia coli Heat-Labile Enterotoxin Hybrids

2001 ◽  
Vol 69 (3) ◽  
pp. 1528-1535 ◽  
Author(s):  
Christal C. Bowman ◽  
John D. Clements
Keyword(s):  
Escherichia Coli ◽  
Cyclic Amp ◽  
Cholera Toxin ◽  
The Other ◽  
Toxin Gene ◽  
Wild Type ◽  
B Subunit ◽  
Heat Labile ◽  
A Subunit ◽  
Ifn Γ

ABSTRACT Two bacterial products that have been demonstrated to function as mucosal adjuvants are cholera toxin (CT), produced by various strains of Vibrio cholerae, and the heat-labile enterotoxin (LT) produced by some enterotoxigenic strains of Escherichia coli. Although LT and CT have many features in common, they are clearly distinct molecules with biochemical and immunologic differences which make them unique. The goal of this study was to determine the basis for these biological differences by constructing and characterizing chimeric CT-LT molecules. Toxin gene fragments were subcloned to create two constructs, each expressing the enzymatically active A subunit of one toxin and the receptor binding B subunit of the other toxin. These hybrid toxins were purified, and the composition and assembly of CT A subunit (CT-A)-LT B subunit (LT-B) and LT A subunit (LT-A)-CT B subunit (CT-B) were confirmed. Hybrids were evaluated for enzymatic activity, as measured by the accumulation of cyclic AMP in Caco-2 cells, and the enterotoxicity of each toxin was assessed in a patent-mouse assay. The results demonstrated that LT-A–CT-B induces the accumulation of lower levels of cyclic AMP and has less enterotoxicity than either wild-type toxin or the other hybrid. Nonetheless, this hybrid retains adjuvant activity equivalent to or greater than that of either wild-type toxin or the other hybrid when used in conjunction with tetanus toxoid for intranasal immunization of BALB/c mice. Importantly, the ability of LT to induce a type 1 cytokine response was found to be a function of LT-A. Specifically, LT-A–CT-B was able to augment the levels of antigen-specific gamma interferon (IFN-γ) and interleukin 5 to levels comparable to those achieved with native LT, while CT-A–LT-B and native CT both produced lower levels of antigen-specific IFN-γ. Thus, these toxin hybrids possess unique biological characteristics and provide information about the basis for differences in the biological activities observed for CT and LT.

Intracellular Signal Triggered by Cholera Toxin inSaccharomyces boulardii and Saccharomyces cerevisiae

1998 ◽  
Vol 64 (2) ◽  
pp. 564-568 ◽  
Author(s):  
Rogelio L. Brandão ◽  
Ieso M. Castro ◽  
Eduardo A. Bambirra ◽  
Sheila C. Amaral ◽  
Luciano G. Fietto ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cholera Toxin ◽  
Mammalian Cells ◽  
Specific Binding ◽  
Saccharomyces Boulardii ◽  
Yeast Cells ◽  
B Subunit ◽  
Nonfermentable Carbon Source ◽  
A Subunit ◽  
Yeast Surface

ABSTRACT As is the case for Saccharomyces boulardii, Saccharomyces cerevisiae W303 protects Fisher rats against cholera toxin (CT). The addition of glucose or dinitrophenol to cells of S. boulardii grown on a nonfermentable carbon source activated trehalase in a manner similar to that observed for S. cerevisiae. The addition of CT to the same cells also resulted in trehalase activation. Experiments performed separately on the A and B subunits of CT showed that both are necessary for activation. Similarly, the addition of CT but not of its separate subunits led to a cyclic AMP (cAMP) signal in both S. boulardii and S. cerevisiae. These data suggest that trehalase stimulation by CT probably occurred through the cAMP-mediated protein phosphorylation cascade. The requirement of CT subunit B for both the cAMP signal and trehalase activation indicates the presence of a specific receptor on the yeasts able to bind to the toxin, a situation similar to that observed for mammalian cells. This hypothesis was reinforced by experiments with 125I-labeled CT showing specific binding of the toxin to yeast cells. The adhesion of CT to a receptor on the yeast surface through the B subunit and internalization of the A subunit (necessary for the cAMP signal and trehalase activation) could be one more mechanism explaining protection against the toxin observed for rats treated with yeasts.

scholarly journals Interactions of cholera toxin with isolated hepatocytes. Effects of low pH, chloroquine and monensin on toxin internalization, processing and action

1988 ◽  
Vol 253 (3) ◽  
pp. 735-743 ◽  
Author(s):  
M Janicot ◽  
J P Clot ◽  
B Desbuquois
Keyword(s):  
Plasma Membrane ◽  
Adenylate Cyclase ◽  
Cholera Toxin ◽  
Acidic Medium ◽  
Isolated Hepatocytes ◽  
Surface Generation ◽  
Lag Phase ◽  
Single Class ◽  
Endosomal Membrane ◽  
A Subunit

The major steps in cholera-toxin action, i.e. binding, internalization, generation of A1 peptide and activation of adenylate cyclase, were examined in isolated hepatocytes. The binding of toxin involves a single class of high-affinity sites (KD congruent to 0.1 nM; Bmax. congruent to 10(7) sites/cell). At 37 degrees C, cell-associated toxin is progressively internalized, as judged by the loss of its accessibility to antibodies against whole toxin, A and B subunits (about 50, 75 and 30% of initially bound toxin after 40 min respectively). Two distinct pathways are involved in this process: endocytosis of the whole toxin, and selective penetration of the A subunit into the plasma membrane. Exposure of hepatocytes to an acidic medium (pH 5) results in a rapid and marked disappearance of the A subunit from the cell surface. Generation of A1 peptide and activation of adenylate cyclase by the toxin occur after a lag phase (10 min at 37 degrees C), and increase with time in a parallel manner up to 2-3% A1 peptide generated; they are unaffected by exposure of cells to an acidic medium. Chloroquine and monensin, which elevate the pH in acidic organelles, inhibit by 2-4-fold both the generation of A1 peptide and the activation of adenylate cyclase. Unexpectedly, these drugs also inhibit the internalization of the toxin. These results suggest that an acidic pH facilitates the penetration of A subunit into the plasma membrane and presumably the endosomal membrane as well, and that endocytosis of cholera toxin is required for generation of A1 peptide and activation of adenylate cyclase.

scholarly journals Gangliosides That Associate with Lipid Rafts Mediate Transport of Cholera and Related Toxins from the Plasma Membrane to Endoplasmic Reticulm

2003 ◽  
Vol 14 (12) ◽  
pp. 4783-4793 ◽  
Author(s):  
Yukako Fujinaga ◽  
Anne A. Wolf ◽  
Chiara Rodighiero ◽  
Heidi Wheeler ◽  
Billy Tsai ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Plasma Membrane ◽  
Cholera Toxin ◽  
Lipid Rafts ◽  
Intestinal Cells ◽  
B Subunit ◽  
High Affinity ◽  
A Subunit ◽  
Human Intestinal Cells

Cholera toxin (CT) travels from the plasma membrane of intestinal cells to the endoplasmic reticulum (ER) where a portion of the A-subunit, the A1 chain, crosses the membrane into the cytosol to cause disease. A related toxin, LTIIb, binds to intestinal cells but does not cause toxicity. Here, we show that the B-subunit of CT serves as a carrier for the A-subunit to the ER where disassembly occurs. The B-subunit binds to gangliosides in lipid rafts and travels with the ganglioside to the ER. In many cells, LTIIb follows a similar pathway, but in human intestinal cells it binds to a ganglioside that fails to associate with lipid rafts and it is sorted away from the retrograde pathway to the ER. Our results explain why LTIIb does not cause disease in humans and suggest that gangliosides with high affinity for lipid rafts may provide a general vehicle for the transport of toxins to the ER.

Anthrax toxin-mediated delivery of cholera toxin-A subunit into the cytosol of mammalian cells

2000 ◽  
Vol 32 (1) ◽  
pp. 69 ◽  
Author(s):  
Manju Sharma ◽  
Hemant Khanna ◽  
Naveen Arora ◽  
Yogendra Singh
Keyword(s):  
Cholera Toxin ◽  
Mammalian Cells ◽  
Anthrax Toxin ◽  
Toxin A ◽  
A Subunit

scholarly journals Sleep-inducing effect of substance P-cholera toxin A subunit in mice

2017 ◽  
Vol 659 ◽  
pp. 44-47 ◽  
Author(s):  
Mark R. Zielinski ◽  
Dmitry Gerashchenko
Keyword(s):  
Substance P ◽  
Cholera Toxin ◽  
Toxin A ◽  
A Subunit
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