Mutants of Type II Heat-Labile Enterotoxin LT-IIa with Altered Ganglioside-Binding Activities and Diminished Toxicity Are Potent Mucosal Adjuvants

Nawar, Hesham F.; Arce, Sergio; Russell, Michael W.; Connell, Terry D.

doi:10.1128/iai.01009-06

Cited by 22 publications

(64 citation statements)

References 58 publications

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“…The toxicity of purified enterotoxins was measured using a standard Y1 adrenal cell (ATCC #CCL-79) bioassay. Y1 cells are acutely sensitive to HLTs and respond by morphological changes in the cell, which are visible by phase-contrast microscopy (23,24). Briefly, 10 5 mouse Y1 adrenal cells were cultured to 75% confluence in 96-well tissue culture plates in F-12 medium supplemented with 30% horse serum and 10% fetal bovine serum at 37°C and in an atmosphere of 5% CO 2 .…”

Section: Methodsmentioning

confidence: 99%

“…Plates were incubated at 37°C in an atmosphere of 5% CO 2 and examined after 4 h to monitor for rounding of the cells, which is an indicator of toxicity. One unit of toxicity was defined as the smallest concentration of enterotoxin that induced rounding of 75 to 100% of the cultured mouse Y1 adrenal cells (23,24).…”

Section: Methodsmentioning

confidence: 99%

“…Binding of LT-IIc, LT-IIa, and LT-IIb to commercially available gangliosides (Matreya, State College, PA) was measured by ELISA (24).…”

Section: Methodsmentioning

confidence: 99%

“…Ganglioside-specific ELISA (23,24) using commercially available gangliosides demonstrated that LT-IIc bound to GM1, GM2, GM3, and GD1a (Fig. 7).…”

Section: Vol 78 2010 Lt-iic a New Type II Heat-labile Enterotoxin mentioning

confidence: 99%

“…To compare the cytotoxic activity of purified LT-IIc to those of CT, LT-IIa, and LT-IIb, a highly sensitive Y1 cell bioassay was employed (23,24). In comparison to other HLTs, LT-IIc exhibited the lowest cytotoxic activity.…”

Section: Vol 78 2010 Lt-iic a New Type II Heat-labile Enterotoxin mentioning

confidence: 99%

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LT-IIc, a New Member of the Type II Heat-Labile Enterotoxin Family Encoded by an Escherichia coli Strain Obtained from a Nonmammalian Host

Nawar

King-Lyons

et al. 2010

Infect Immun

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Infections caused by enterotoxigenic Escherichia coli (ETEC)are the leading cause of traveler's diarrhea and the major cause of diarrheal disease in underdeveloped nations, especially among children. ETEC, which is usually transmitted by food or water contaminated with animal or human feces, is estimated to be responsible annually for more than 650 million cases of enteric infections and nearly 800,000 deaths (29). Infection begins with ingestion of bacteria, followed by elaboration of enterotoxin and bacterial colonization of the gut, and presents as a profuse watery diarrhea which disseminates the bacteria back into the environment (10).ETEC strains are lactose-fermenting E. coli strains that produce a heat-labile enterotoxin (LT, hereafter referred to as LT-I), heat-stable enterotoxins (ST), or both and colonization factors which enable ETEC to colonize the small intestine (22). The pathogenesis of ETEC is dependent on the strains' capacity to produce LT-I and/or ST (10, 29). LT-I is closely related functionally, antigenically, and structurally to cholera toxin (CT), the heat-labile enterotoxin produced by Vibrio cholerae. Antiserum against CT neutralizes the toxicity of LT-I, and antiserum against LT-I neutralizes the toxicity of CT (15). Structurally, LT-I and CT are oligomeric proteins composed of an A polypeptide which is noncovalently coupled to a pentameric array of B polypeptides (15). The A polypeptide of LT-I and CT is enzymatically active and catalyzes an ADP-ribosylation of the G s ␣ regulatory protein in the intoxicated cell. Ribosylation of this regulatory protein constitutively activates adenylate cyclase, the enzyme which catalyzes production of cyclic AMP (cAMP) (3,20). Accumulation of cAMP induces the intoxicated cell to secrete electrolytes and chloride ions, thus generating the watery diarrhea, which is symptomatic of intoxication. Intracellular accumulation of cAMP modulates other cellular processes such as protein kinase activity, activation of calcium channels, etc. (15). Binding of LT-I and CT to ganglioside receptors is mediated by the B polypeptides. Gangliosides are members of a heterogeneous family of sialylated glycosphingolipids expressed on the surface of eukaryotic cells (9). Based on these characteristics, LT-I and CT have been designated as members of the large family of toxins known as the A 1 B 5 ADP-ribosylating heat-labile enterotoxins (HLTs).LT-IIa and LT-IIb, two new members of the A 1 B 5 family of HLTs produced by E. coli, were recently described (11,12,27). While it is clear that LT-IIa and LT-IIb are evolutionarily related to LT-I and CT, there are major differences between the two groups of enterotoxins. LT-IIa and LT-IIb are antigenically distinguishable from LT-I and CT and from each other (12). These antigenic differences are reflected in the low amino acid sequence similarity of the A polypeptides and the virtual absence of amino acid sequence homology of the B polypeptides between the two groups (LT-I and CT versus LT-IIa and LT-IIb) (35). To distinguish betwee...

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Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

“…Binding of LT-IIc, LT-IIa, and LT-IIb to commercially available gangliosides (Matreya, State College, PA) was measured by ELISA (24).…”

Section: Methodsmentioning

confidence: 99%

“…Ganglioside-specific ELISA (23,24) using commercially available gangliosides demonstrated that LT-IIc bound to GM1, GM2, GM3, and GD1a (Fig. 7).…”

Section: Vol 78 2010 Lt-iic a New Type II Heat-labile Enterotoxin mentioning

confidence: 99%

Section: Vol 78 2010 Lt-iic a New Type II Heat-labile Enterotoxin mentioning

confidence: 99%

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LT-IIc, a New Member of the Type II Heat-Labile Enterotoxin Family Encoded by an Escherichia coli Strain Obtained from a Nonmammalian Host

Nawar

King-Lyons

et al. 2010

Infect Immun

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2007

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Mutants of Type II Heat-Labile Enterotoxin LT-IIa with Altered Ganglioside-Binding Activities and Diminished Toxicity Are Potent Mucosal Adjuvants

Cited by 22 publications

References 58 publications

LT-IIc, a New Member of the Type II Heat-Labile Enterotoxin Family Encoded by an Escherichia coli Strain Obtained from a Nonmammalian Host

LT-IIc, a New Member of the Type II Heat-Labile Enterotoxin Family Encoded by an Escherichia coli Strain Obtained from a Nonmammalian Host

Mucosal SIgA Enhancement: Development of Safe and Effective Mucosal Adjuvants and Mucosal Antigen Delivery Vehicles

Bacterial Toxins as Immunomodulators

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