Glycosylated SV2A and SV2B mediate the entry of botulinum neurotoxin E into neurons.
Botulinum neurotoxin E (BoNT/E) can cause paralysis in humans and animals by blocking neurotransmitter release from presynaptic nerve terminals. How this toxin targets and enters neurons is not known. Here we identified two isoforms of the synaptic vesicle protein SV2, SV2A and SV2B, as the protein receptors for BoNT/E. BoNT/E failed to enter neurons cultured from SV2A/B knockout mice; entry was restored by expressing SV2A or SV2B, but not SV2C. Mice lacking SV2B displayed reduced sensitivity to BoNT/E. The fourth luminal domain of SV2A or SV2B alone, expressed in chimeric receptors by replacing the extracellular domain of the low-density lipoprotein receptor, can restore the binding and entry of BoNT/E into neurons lacking SV2A/B. Furthermore, we found disruption of a N-glycosylation site (N573Q) within the fourth luminal domain of SV2A rendered the mutant unable to mediate the entry of BoNT/E and also reduced the entry of BoNT/A. Finally, we demonstrate that BoNT/E failed to bind and enter ganglioside-deficient neurons; entry was rescued by loading exogenous gangliosides into neuronal membranes. Together, the data reported here demonstrate that glycosylated SV2A and SV2B act in conjunction with gangliosides to mediate the entry of BoNT/E into neurons.
Dong
Min
M
Howard Hughes Medical Institute and Department of Physiology, University of Wisconsin, Madison, WI 53706, USA.
Liu
Huisheng
H
Tepp
William H
WH
Johnson
Eric A
EA
Janz
Roger
R
Chapman
Edwin R
ER
D000595:Amino Acid Sequence; D000818:Animals; D001905:Botulinum Toxins / Q000378:metabolism*; D002478:Cells, Cultured; D005732:Gangliosides / Q000378:metabolism; D006031:Glycosylation; D006624:Hippocampus / Q000166:cytology; D006801:Humans; D008562:Membrane Glycoproteins / Q000235:genetics / Q000378:metabolism*; D008564:Membrane Potentials / Q000502:physiology; D051379:Mice; D018345:Mice, Knockout; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000378:metabolism*; D009474:Neurons / Q000166:cytology / Q000378:metabolism*; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism*; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D013572:Synaptic Vesicles / Q000378:metabolism; D053586:Virus Internalization
D005732:Gangliosides; D008562:Membrane Glycoproteins; D009419:Nerve Tissue Proteins; D020033:Protein Isoforms; D011993:Recombinant Fusion Proteins; C489614:Sv2a protein, mouse; C489615:Sv2b protein, mouse; D001905:Botulinum Toxins; C096326:botulinum toxin type E