Plasma membrane localization signals in the light chain of botulinum neurotoxin.
Botulinum neurotoxin (BoNT) is a potent biological substance used to treat neuromuscular and pain disorders. Both BoNT type A and BoNT type E display high-affinity uptake into motor neurons and inhibit exocytosis through cleavage of the synaptosome-associated protein of 25 kDa (SNAP25). The therapeutic effects of BoNT/A last from 3 to 12 months, whereas the effects of BoNT/E last less than 4 weeks. Using confocal microscopy and site-specific mutagenesis, we have determined that the protease domain of BoNT/A light chain (BoNT/A-LC) localizes in a punctate manner to the plasma membrane, colocalizing with the cleaved product, SNAP25(197). In contrast, the short-duration BoNT/E serotype is cytoplasmic. Mutations in the BoNT/A-LC have revealed sequences at the N terminus necessary for plasma membrane localization, and an active dileucine motif in the C terminus that is likely involved in trafficking and interaction with adaptor proteins. These data support sequence-specific signals as determinants of intracellular localization and as a basis for the different durations of action in these two BoNT serotypes.
Fernández-Salas
Ester
E
Neurotoxin Research Program, Department of Biological Sciences, Allergan Inc., 2525 Dupont Drive, Irvine, CA 92612-1599, USA. fernandez_ester@allergan.com
Steward
Lance E
LE
Ho
Helen
H
Garay
Patton E
PE
Sun
Sarah W
SW
Gilmore
Marcella A
MA
Ordas
Joseph V
JV
Wang
Joanne
J
Francis
Joseph
J
Aoki
K Roger
KR
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D015153:Blotting, Western; D001905:Botulinum Toxins / Q000737:chemistry* / Q000378:metabolism; D019274:Botulinum Toxins, Type A / Q000737:chemistry* / Q000378:metabolism; D002352:Carrier Proteins / Q000378:metabolism; D002462:Cell Membrane / Q000502:physiology*; D002868:Chromosomal Proteins, Non-Histone; D007930:Leucine; D046911:Macromolecular Substances; D009419:Nerve Tissue Proteins / Q000378:metabolism; D016716:PC12 Cells; D051381:Rats; D011994:Recombinant Proteins / Q000378:metabolism; D014162:Transfection
D002352:Carrier Proteins; D002868:Chromosomal Proteins, Non-Histone; D046911:Macromolecular Substances; D009419:Nerve Tissue Proteins; D011994:Recombinant Proteins; C459407:Zwint protein, rat; D001905:Botulinum Toxins; D019274:Botulinum Toxins, Type A; D007930:Leucine; C096326:botulinum toxin type E