Mutant SNAP25B causes myasthenia, cortical hyperexcitability, ataxia, and intellectual disability.
OBJECTIVE
To identify and characterize the molecular basis of a syndrome associated with myasthenia, cortical hyperexcitability, cerebellar ataxia, and intellectual disability.


METHODS
We performed in vitro microelectrode studies of neuromuscular transmission, performed exome and Sanger sequencing, and analyzed functional consequences of the identified mutation in expression studies.


RESULTS
Neuromuscular transmission at patient endplates was compromised by reduced evoked quantal release. Exome sequencing identified a dominant de novo variant, p.Ile67Asn, in SNAP25B, a SNARE protein essential for exocytosis of synaptic vesicles from nerve terminals and of dense-core vesicles from endocrine cells. Ca(2+)-triggered exocytosis is initiated when synaptobrevin attached to synaptic vesicles (v-SNARE) assembles with SNAP25B and syntaxin anchored in the presynaptic membrane (t-SNAREs) into an α-helical coiled-coil held together by hydrophobic interactions. Pathogenicity of the Ile67Asn mutation was confirmed by 2 measures. First, the Ca(2+) triggered fusion of liposomes incorporating v-SNARE with liposomes containing t-SNAREs was hindered when t-SNAREs harbored the mutant SNAP25B moiety. Second, depolarization of bovine chromaffin cells transfected with mutant SNAP25B or with mutant plus wild-type SNAP25B markedly reduced depolarization-evoked exocytosis compared with wild-type transfected cells.


CONCLUSIONS
Ile67Asn variant in SNAP25B is pathogenic because it inhibits synaptic vesicle exocytosis. We attribute the deleterious effects of the mutation to disruption of the hydrophobic α-helical coiled-coil structure of the SNARE complex by replacement of a highly hydrophobic isoleucine by a strongly hydrophilic asparagine.
From the Department of Neurology and Neuromuscular Research Laboratory, Mayo Clinic, Rochester, MN. shen.xinming@mayo.edu age@mayo.edu.
Xin-Ming
XM
Shen
From the Department of Neurology and Neuromuscular Research Laboratory, Mayo Clinic, Rochester, MN.
Duygu
D
Selcen
From the Department of Neurology and Neuromuscular Research Laboratory, Mayo Clinic, Rochester, MN.
Joan
J
Brengman
From the Department of Neurology and Neuromuscular Research Laboratory, Mayo Clinic, Rochester, MN. shen.xinming@mayo.edu age@mayo.edu.
Andrew G
AG
Engel
D000293:Adolescent; D000818:Animals; D001259:Ataxia / Q000235:genetics* / Q000503:physiopathology; D001927:Brain Diseases / Q000235:genetics* / Q000503:physiopathology; D019556:COS Cells; D002417:Cattle; D002522:Chlorocebus aethiops; D019439:Chromaffin Cells / Q000502:physiology; D004252:DNA Mutational Analysis; D005089:Exocytosis / Q000502:physiology; D005260:Female; D006801:Humans; D008607:Intellectual Disability / Q000235:genetics* / Q000503:physiopathology; D008839:Microelectrodes; D009045:Motor Endplate / Q000503:physiopathology; D009154:Mutation; D050600:SNARE Proteins / Q000378:metabolism; D013572:Synaptic Vesicles / Q000502:physiology; D050825:Synaptosomal-Associated Protein 25 / Q000235:genetics* / Q000378:metabolism; D013577:Syndrome; D014162:Transfection
C494208:SNAP25 protein, human; D050600:SNARE Proteins; D050825:Synaptosomal-Associated Protein 25