A nontoxic cholera enterotoxin (CT) analog is chimeric with regard to both epitypes of CT-B subunits, CT-B-1 and CT-B-2

Boesman-Finkelstein, Mary; Peterson, Johnny W.; Thai, L. S.; Finkelstein, Richard A.

doi:10.1128/iai.64.1.346-348.1996

Cited by 5 publications

(1 citation statement)

References 17 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…CT-2* is a CT analog in which two codon substitutions altered the CT-A subunit (Arg73Lys and Glu1123Gln) and eliminated the toxin's ADP-ribosylation activity (19). The CT-2* protein was purified to homogeneity from V. cholerae CVD103[CT-2*] culture filtrates, as described and characterized previously (4). Briefly, the modified CT-2*-encoding gene was subcloned into V. cholerae CVD103 (ctxA ctxB ϩ ), and a recombinant strain that secreted inactive CT, referred to as CT-2*, was selected (4).…”

Section: Methodsmentioning

confidence: 99%

Cholera Toxin B Subunit Activates Arachidonic Acid Metabolism

et al. 1999

View full text Add to dashboard Cite

Cholera toxin (CT) increases intestinal secretion of water and electrolytes and modulates the mucosal immune response by stimulating cellular synthesis of arachidonic acid (AA) metabolites (e.g., prostaglandin E2), as well as the intracellular second messenger cyclic AMP (cAMP). While much is known about the mechanism of CT stimulation of adenylate cyclase, the toxin’s activation of phospholipase A2, which results in increased hydrolysis of AA from membrane phospholipids, is not well understood. To determine whether CT activation of AA metabolism requires CT’s known enzymatic activity (i.e., ADP-ribosylation of GSα), we used native CT and a mutant CT protein (CT-2*) lacking ADP-ribose transferase activity in combination with S49 wild-type (WT) and S49 cyc− murine Theta (Th)1.2-positive lymphoma cells deficient in GSα. The experimental results showed that native CT stimulated the release of [3H[AA from S49 cyc− cells at a level similar to that for S49 WT cells, indicating that GSα is not essential for this process. Further, levels of cAMP in the CT-treated cyc− cells remained the same as those in the untreated control cells. The ADP-ribosyltransferase-deficient CT-2* protein, which was incapable of increasing synthesis of cAMP, displayed about the same capacity as CT to evoke the release of [3H]AA metabolites from both S49 WT and cyc− cells. We concluded that stimulation of arachidonate metabolism in S49 murine lymphoma cells by native CT does not require enzymatically functional CT, capable of catalyzing the ADP-ribosylation reaction. These results demonstrated for the first time that stimulation of adenylate cyclase by CT and stimulation of AA metabolism by CT are not necessarily coregulated. In addition, the B subunits purified from native CT and CT-2* both simulated the release of [3H]AA from S49 cyc− cells and murine monocyte/macrophage cells (RAW 264.7), suggesting a receptor-mediated cell activation process of potential importance in enhancing immune responses to vaccine components.

show abstract

Section: Methodsmentioning

confidence: 99%

Cholera Toxin B Subunit Activates Arachidonic Acid Metabolism

et al. 1999

View full text Add to dashboard Cite

show abstract

Overexpression of a Mutant B Subunit in Toxigenic Vibrio cholerae Diminishes Production of Active Cholera Toxin In Vivo

1998

View full text Add to dashboard Cite

A mutant cholera toxin B subunit containing a G33E substitution was constructed and expressed in V. cholerae. The G33E amino acid substitution did not affect the amount of recombinant CTB secreted to the culture medium. The overexpression of the mutant B subunits in wild-type toxigenic cholera vibrios led to an 80% decrease in production of active cholera toxin in vitro and in vivo. Overexpression of BG33E subunits could be instrumental in the increase of the biosafety of live attenuated cholera candidate vaccine strains.

show abstract

A mutated cholera toxin without the ADP-ribosyltransferase activity induces cytokine production and inhibits apoptosis of splenocytes in mice possibly via toll-like receptor-4 signaling

Yang

Liu

et al. 2016

Molecular Immunology

View full text Add to dashboard Cite

Native cholera toxin (CT) and its mutated form (CT-2*) without ADP-ribosyltransferase activity differ in their immunomodulatory effects on host cells, and the mechanisms of these differences are poorly understood. In this study, we demonstrated that CT-2* induced higher levels of cytokine production and down-regulated ex-vivo apoptosis of splenocytes from C57BL/6 mice. After exposure of the splenocytes ex-vivo to CT or CT-2* (2μg/ml) for 48h, CT-2* stimulated expression of the toll-like receptor (TLR-4) gene was much higher and the cells produced increased levels of interleukin (IL)-12, interferon (IFN)-γ, and tumor necrosis factor (TNF)-α, compared to splenocytes of mice exposed to native CT. We confirmed these findings by observing that CT-2*, induced much lower levels of IL-12, IFN-γ, and TNF-α in a TLR-4 knockout macrophage cell line derived from C57BL/6 mice. In addition, while CT is known to stimulate apoptosis in splenocytes, we observed that CT-2* significantly down-regulated apoptosis (4.2%), compared to splenocytes exposed to CT (18.7%) or PBS (negative control, 8.5%). On the contrary, we noted both native CT and CT-2* to exhibit similar levels of apoptosis in TLR-4(-/-) cell line. Overall, the evidence supports the conclusion that CT-2* modulated cytokine production and apoptosis in splenocytes of mice possibly through the TLR-4 signaling pathway.

show abstract

A nontoxic cholera enterotoxin (CT) analog is chimeric with regard to both epitypes of CT-B subunits, CT-B-1 and CT-B-2

Cited by 5 publications

References 17 publications

Cholera Toxin B Subunit Activates Arachidonic Acid Metabolism

Cholera Toxin B Subunit Activates Arachidonic Acid Metabolism

Overexpression of a Mutant B Subunit in Toxigenic Vibrio cholerae Diminishes Production of Active Cholera Toxin In Vivo

A mutated cholera toxin without the ADP-ribosyltransferase activity induces cytokine production and inhibits apoptosis of splenocytes in mice possibly via toll-like receptor-4 signaling

Contact Info

Product

Resources

About