Association of Botulinum Neurotoxin Serotype A Light Chain with Plasma Membrane-bound SNAP-25

Chen, Sheng; Barbieri, Joseph T.

doi:10.1074/jbc.m111.224493

Cited by 38 publications

(55 citation statements)

References 39 publications

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“…The results indicate that association of GFP-LCA with SNARE proteins is weak and/or transient and, hence, is unlikely to be responsible for membrane tethering of LCA as was previously suggested (Chen and Barbieri, 2011). The data provide no evidence for a specific interaction between LCA and adaptor proteins complexes.…”

Section: Identification Of Proteins That Interact With Lca Through Thcontrasting

confidence: 34%

“…Membrane cytoskeleton proteins rather than SNARE proteins are preferred LCA-interacting partners Recent studies have suggested that the interaction of LCA and SNAP-25 is responsible for stable association of LCA with the plasma membrane in neurons (Chen and Barbieri, 2011). However, our results from a global analysis of the LCA interactome in SiMa cells (Table 1; supplementary material Table S1) indicate that association of GFP-LCA with SNAP-25 is weak and transient and hence is unlikely to be responsible for the stabilization of LCA.…”

Section: Discussion New Properties Of Lca Dileucine Motifmentioning

confidence: 99%

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Recruitment of septin cytoskeletal proteins by Botulinum toxin A protease determines its remarkable stability

Vagin

Tokhtaeva

Garay

et al. 2014

Journal of Cell Science

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Proteolytic cleavage of synaptosomal-associated protein 25 by the light chain of botulinum neurotoxin type A (LCA) results in a blockade of neurotransmitter release that persists for several months in motor neurons. The L428A/L429A mutation in LCA is known to significantly shorten both the proteolytic and neuroparalytic effects of the neurotoxin in mice. To elucidate the cellular mechanism for LCA longevity, we studied the effects of L428A/L429A mutation on the interactome, localization and stability of LCA expressed in cultured neuronal cells. Mass spectrometry analysis of the LCA interactome showed that the mutation prevented the interaction of LCA with septins. The wild-type LCA was concentrated in plasma-membrane-associated clusters, colocalizing with septins-2 and septin-7, which accumulated in these clusters only in the presence of LCA. The L428A/L429A mutation decreased co-clustering of LCA and septins and accelerated proteasomal and non-proteasomal degradation of LCA. Similarly, the impairment of septin oligomerization by forchlorfenuron or silencing of septin-2 prevented LCA interaction and clustering with septins and increased LCA degradation. Therefore, the dileucine-mediated LCA-septin co-clustering is crucial for the long-lasting stabilization of LCA-related proteolytic and presumably neuroparalytic activity.

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Section: Identification Of Proteins That Interact With Lca Through Thcontrasting

confidence: 34%

Section: Discussion New Properties Of Lca Dileucine Motifmentioning

confidence: 99%

Recruitment of septin cytoskeletal proteins by Botulinum toxin A protease determines its remarkable stability

Vagin

Tokhtaeva

Garay

et al. 2014

Journal of Cell Science

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“…Neurons-BoNT/A promotes paralysis through the cleavage of 9 amino acid residues from the C terminus of SNAP25 (21,(13)(14)(15)(16)(17)(18)(19)(20)(21)(22)(23)(24)(25). We next investigated whether Dyngo-4a could prevent the cleavage of SNAP25 in hippocampal neurons.…”

Section: Dyngo-4a Prevents Bont/a-induced Snap25 Cleavage In Culturedmentioning

confidence: 99%

Dynamin Inhibition Blocks Botulinum Neurotoxin Type A Endocytosis in Neurons and Delays Botulism

Harper

Martin

Nguyen

et al. 2011

Journal of Biological Chemistry

145

149

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The botulinum neurotoxins (BoNTs) are di-chain bacterial proteins responsible for the paralytic disease botulism. Following binding to the plasma membrane of cholinergic motor nerve terminals, BoNTs are internalized into an endocytic compartment. Although several endocytic pathways have been characterized in neurons, the molecular mechanism underpinning the uptake of BoNTs at the presynaptic nerve terminal is still unclear. Here, a recombinant BoNT/A heavy chain binding domain (Hc) was used to unravel the internalization pathway by fluorescence and electron microscopy. BoNT/A-Hc initially enters cultured hippocampal neurons in an activity-dependent manner into synaptic vesicles and clathrin-coated vesicles before also entering endosomal structures and multivesicular bodies. We found that inhibiting dynamin with the novel potent Dynasore analog, Dyngo-4a TM , was sufficient to abolish BoNT/ A-Hc internalization and BoNT/A-induced SNAP25 cleavage in hippocampal neurons. Dyngo-4a also interfered with BoNT/ A-Hc internalization into motor nerve terminals. Furthermore, Dyngo-4a afforded protection against BoNT/A-induced paralysis at the rat hemidiaphragm. A significant delay of >30% in the onset of botulism was observed in mice injected with Dyngo-4a. Dynamin inhibition therefore provides a therapeutic avenue for the treatment of botulism and other diseases caused by pathogens sharing dynamin-dependent uptake mechanisms.

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“…Within the periphery of the motor neuron, the LC cleaves SNARE proteins, resulting in loss of stimulatory signaling between neurons and muscles, yielding flaccid paralysis. The LC of BoNT/A localizes to the plasma membrane to target synaptosomal-associated protein 25 (SNAP25) for cleavage within neurons (6).…”

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confidence: 99%

Entry of a Recombinant, Full-Length, Atoxic Tetanus Neurotoxin into Neuro-2a Cells

et al. 2014

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Tetanus neurotoxin (TeNT) and botulinum neurotoxin (BoNT) are clostridial neurotoxins (CNTs) that are the most toxic proteins for humans (1). TeNT and BoNT share ϳ35% identity and ϳ65% similarity and overall structure-function properties (2). BoNT intoxication results in flaccid paralysis through the inhibition of acetylcholine release by motor neurons, while TeNT intoxication yields a spastic paralysis due to inhibition of glycine release by inhibitory neurons (3). TeNT and BoNT are expressed as ϳ150-kDa single-chain proteins that are cleaved to form dichain proteins linked by a disulfide bond (2). The N-terminal 50-kDa light chain (LC) is a zinc-metalloprotease that cleaves neuronspecific soluble NSF attachment protein (SNAP) receptor (SNARE) proteins (4). TeNT and BoNT serotype B cleave the same residue within vesicle-associated membrane protein 2 (VAMP2), a SNARE protein of synaptic vesicles (SVs). The C-terminal 100-kDa heavy chain (HC) contains two structurally distinct domains with separate functions. The translocation domain (HCT) facilitates LC translocation from the SV lumen into the cell cytosol, and the receptor binding domain (HCR) binds dual host receptors.BoNT/A binds a ganglioside and synaptic vesicle protein 2 (SV2) and enters neurons upon SV recycling from the plasma membrane (5). Upon SV acidification within the periphery of motor neurons, the HCT undergoes a pH-dependent conformational change and inserts into the SV membrane, forming a channel that allows the LC to escape into the cytosol. Within the periphery of the motor neuron, the LC cleaves SNARE proteins, resulting in loss of stimulatory signaling between neurons and muscles, yielding flaccid paralysis. The LC of BoNT/A localizes to the plasma membrane to target synaptosomal-associated protein 25 (SNAP25) for cleavage within neurons (6).TeNT binds two gangliosides as functional receptors (7). TeNT can bind a glycophosphatidylinositol (GPI)-anchored protein (8) or SV2 (9), but the significance of these interactions has not been defined (10) or reproduced (11), respectively. TeNT enters motor neurons upon endocytosis (12) and traffics through motor neurons associated with Rab7-enriched endosomes that are of neutral pH (13,14). Retrograde trafficking proceeds from the axon to the soma, where TeNT transcytoses from the motor neuron into an inhibitory neuron of the central nervous system (CNS). Upon vesicle acidification, the LC is translocated into the cytosol and cleaves VAMP2. The block in signaling between the inhibitory neurons and motor neurons leads to the spastic paralysis characteristic of tetanus. The molecular mechanism responsible for the unique entry of BoNT and TeNT is not clearly understood.The modular structural domains of the CNTs have permitted the study of individual domains to assess protein structure-function in vitro; the LC is a functional SNARE protease (15), the HCT inserts into lipid bilayers and forms a channel (16), and the HCR binds host receptors (17). The tetanus HCR (HCR/T) can retrograde traffic in both cultu...

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Association of Botulinum Neurotoxin Serotype A Light Chain with Plasma Membrane-bound SNAP-25

Cited by 38 publications

References 39 publications

Recruitment of septin cytoskeletal proteins by Botulinum toxin A protease determines its remarkable stability

Recruitment of septin cytoskeletal proteins by Botulinum toxin A protease determines its remarkable stability

Dynamin Inhibition Blocks Botulinum Neurotoxin Type A Endocytosis in Neurons and Delays Botulism

Entry of a Recombinant, Full-Length, Atoxic Tetanus Neurotoxin into Neuro-2a Cells

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