Gangliosides are the binding substances in neural cells for tetanus and botulinum toxins in mice

Kitamura, Masaru; Takamiya, Kogo; Aizawa, Shinichi; Furukawa, Koichi

doi:10.1016/s1388-1981(99)00140-7

Cited by 106 publications

(71 citation statements)

References 15 publications

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“…Chen et al (10) demonstrated that treatment of gangliosideloaded PC12 cells, a pheochromocytoma cell line, with proteinase K does not change the cell association of HCR/T and that loading HeLa cells with exogenous ganglioside is sufficient for HCR/T cell entry. A role for gangliosides in TeNT entry is also supported by earlier determinations that mice lacking complex gangliosides by disruption of GM2/GD2 synthase are ϳ600-fold more resistant to TeNT than wild-type mice (23).…”

Section: Discussionmentioning

confidence: 65%

Tetanus Toxin and Botulinum Toxin A Utilize Unique Mechanisms To Enter Neurons of the Central Nervous System

et al. 2012

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ABSTRACTBotulinum neurotoxins (BoNTs) and tetanus neurotoxin (TeNT) are the most toxic proteins for humans. While BoNTs cause flaccid paralysis, TeNT causes spastic paralysis. Characterized BoNT serotypes enter neurons upon binding dual receptors, a ganglioside and a neuron-specific protein, either synaptic vesicle protein 2 (SV2) or synaptotagmin, while TeNT enters upon binding gangliosides as dual receptors. Recently, TeNT was reported to enter central nervous system (CNS) neurons upon synaptic vesicle cycling that was mediated by the direct binding to SV2, implying that TeNT and BoNT utilize common mechanisms to enter CNS neurons. This prompted an assessment of TeNT entry into CNS neurons, using the prototypic BoNT serotype A as a reference for SV2-mediated entry into synaptic vesicles, analyzing the heavy-chain receptor binding domain (HCR) of each toxin. Synaptic vesicle cycling stimulated the entry of HCR/A into neurons, while HCR/T entered neurons with similar levels of efficiency in depolarized and nondepolarized neurons. ImageJ analysis identified two populations of cell-associated HCR/T in synaptic vesicle cycling neurons, a major population which segregated from HCR/A and a minor population which colocalized with HCR/A. HCR/T did not inhibit HCR/A entry into neurons in competition experiments and did not bind SV2, the protein receptor for BoNT/A. Intoxication experiments showed that TeNT efficiently cleaved VAMP2 in depolarized neurons and neurons blocked for synaptic vesicle cycling. These experiments demonstrate that TeNT enters neurons by two pathways, one independent of stimulated synaptic vesicle cycling and one by synaptic vesicles independent of SV2, showing that TeNT and BoNT/A enter neurons by unique mechanisms.

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Section: Discussionmentioning

confidence: 65%

Tetanus Toxin and Botulinum Toxin A Utilize Unique Mechanisms To Enter Neurons of the Central Nervous System

et al. 2012

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“…As for gangliosides, it is well known that GT1b/GD1b are receptors for botulinum toxins and/or tetanus toxins (23), and this has been confirmed using complex ganglioside-lacking mice (24), which were relatively resistant to the injected toxins. However, the fact that the mice finally died suggested that coreceptors for individual toxins were present (23).…”

Section: Discussionmentioning

confidence: 73%

Targeted Disruption of Gb3/CD77 Synthase Gene Resulted in the Complete Deletion of Globo-series Glycosphingolipids and Loss of Sensitivity to Verotoxins

Okuda

Tokuda

Numata

et al. 2006

Journal of Biological Chemistry

174

166

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To examine whether globotriaosylceramide (Gb3/CD77) is a receptor for verotoxins (VTs) in vivo, sensitivity of Gb3/CD77 synthase null mutant mice to VT-2 and VT-1 was analyzed. Although wild-type mice died after administration of 0.02 g of VT-2 or 1.0 g of VT-1, the mutant mice showed no reaction to doses as much as 100 times that administered to wild types. Expression analysis of Gb3/CD77 in mouse tissues with antibody revealed that low, but definite, levels of Gb3/CD77 were expressed in the microvascular endothelial cells of the brain cortex and pia mater and in renal tubular capillaries. Corresponding to the Gb3/CD77 expression, tissue damage with edema, congestion, and cytopathic changes was observed, indicating that Gb3/CD77 (and its derivatives) exclusively function as a receptor for VTs in vivo. The lethal kinetics were similar regardless of lipopolysaccharide elimination in VT preparation, suggesting that basal Gb3/CD77 levels are sufficient for lethal effects of VTs.

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“…Furthermore, mice lacking all complex gangliosides through disruption of the ␤1,4-N-acetylgalactosaminyltransferase gene (GM2/GD2 synthase) displayed an ϳ600-fold decrease in sensitivity to TeNT relative to wild type littermates. In contrast, the related botulinum neurotoxins types A and B (BoNT/A and BoNT/B), which bind a single molecule of ganglioside in conjunction with a protein co-receptor (31-33), displayed only 40-and 24-fold reductions in toxicity, respectively, relative to wild type littermates (34). Together, these observations support a model in which TeNT toxicity requires simultaneous interaction with two molecules of ganglioside.…”

Section: Discussionmentioning

confidence: 80%

Gangliosides as High Affinity Receptors for Tetanus Neurotoxin

Chen

Kim

et al. 2009

Journal of Biological Chemistry

118

134

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Tetanus neurotoxin (TeNT) is an exotoxin produced byClostridium tetani that causes paralytic death to hundreds of thousands of humans annually. TeNT cleaves vesicle-associated membrane protein-2, which inhibits neurotransmitter release in the central nervous system to elicit spastic paralysis, but the molecular basis for TeNT entry into neurons remains unclear. TeNT is a ϳ150-kDa protein that has AB structure-function properties; the A domain is a zinc metalloprotease, and the B domain encodes a translocation domain and C-terminal receptor-binding domain (HCR/T). Earlier studies showed that HCR/T bound gangliosides via two carbohydrate-binding sites, termed the lactose-binding site (the "W" pocket) and the sialic acid-binding site (the "R" pocket). Here we report that TeNT high affinity binding to neurons is mediated solely by gangliosides. Glycan array and solid phase binding analyses identified gangliosides that bound exclusively to either the W pocket or the R pocket of TeNT; GM1a bound to the W pocket, and GD3 bound to the R pocket. Using these gangliosides and mutated forms of HCR/T that lacked one or both carbohydrate-binding pocket, gangliosides binding to both of the W and R pockets were shown to be necessary for high affinity binding to neuronal and non-neuronal cells. The crystal structure of a ternary complex of HCR/T with sugar components of two gangliosides bound to the W and R supported the binding of gangliosides to both carbohydrate pockets. These data show that gangliosides are functional dual receptors for TeNT.Tetanus is an acute, often fatal disease of humans that was first described by Hippocrates over 24 centuries ago (1). Tetanus is characterized by generalized increased rigidity and convulsive spasms of skeletal muscles. Tetanus is caused by exposure to tetanus neurotoxin (TeNT) 3 produced by the sporeforming bacterium Clostridium tetani. TeNT is delivered from the bloodstream to the peripheral nervous system, from where TeNT traffics to the central nervous system to cleave vesicleassociated membrane protein-2 (VAMP2), which inhibits neurotransmitter release and elicits spastic paralysis (2). Although prevented by vaccination, tetanus is responsible for hundreds of thousands of deaths per year in countries where vaccination is not common (3).TeNT is produced as a ϳ150-kDa protein that is cleaved to a di-chain protein, comprising an N-terminal light chain (ϳ50 kDa) and a C-terminal heavy chain domain (ϳ100 kDa) linked through a single disulfide bond (4). TeNT light chain is a zinc metalloprotease that cleaves the neuronal SNARE protein VAMP2 (2). The TeNT heavy chain contains two functional domains: a translocation domain and a C-terminal receptorbinding domain (HCR/T, ϳ50 kDa).The first step in TeNT action involves binding to a receptor(s) on the presynaptic membrane of ␣-motor neurons. Although the molecular basis for TeNT entry remains undetermined, an unambiguous role for gangliosides has been demonstrated (5-9). Current models implicate a dual receptor mechanism for the binding of t...

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Gangliosides are the binding substances in neural cells for tetanus and botulinum toxins in mice

Cited by 106 publications

References 15 publications

Tetanus Toxin and Botulinum Toxin A Utilize Unique Mechanisms To Enter Neurons of the Central Nervous System

Tetanus Toxin and Botulinum Toxin A Utilize Unique Mechanisms To Enter Neurons of the Central Nervous System

Targeted Disruption of Gb3/CD77 Synthase Gene Resulted in the Complete Deletion of Globo-series Glycosphingolipids and Loss of Sensitivity to Verotoxins

Gangliosides as High Affinity Receptors for Tetanus Neurotoxin

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