Analysis of conditional heterozygous STXBP1 mutations in human neurons.
Heterozygous mutations in the syntaxin-binding protein 1 (STXBP1) gene, which encodes Munc18-1, a core component of the presynaptic membrane-fusion machinery, cause infantile early epileptic encephalopathy (Ohtahara syndrome), but it is unclear how a partial loss of Munc18-1 produces this severe clinical presentation. Here, we generated human ES cells designed to conditionally express heterozygous and homozygous STXBP1 loss-of-function mutations and studied isogenic WT and STXBP1-mutant human neurons derived from these conditionally mutant ES cells. We demonstrated that heterozygous STXBP1 mutations lower the levels of Munc18-1 protein and its binding partner, the t-SNARE-protein Syntaxin-1, by approximately 30% and decrease spontaneous and evoked neurotransmitter release by nearly 50%. Thus, our results confirm that using engineered human embryonic stem (ES) cells is a viable approach to studying disease-associated mutations in human neurons on a controlled genetic background, demonstrate that partial STXBP1 loss of function robustly impairs neurotransmitter release in human neurons, and suggest that heterozygous STXBP1 mutations cause early epileptic encephalopathy specifically through a presynaptic impairment.
Christopher
C
Patzke
Yan
Y
Han
Jason
J
Covy
Fei
F
Yi
Stephan
S
Maxeiner
Marius
M
Wernig
Thomas C
TC
Südhof
D053595:Embryonic Stem Cells / Q000378:metabolism / Q000473:pathology; D005071:Evoked Potentials / Q000235:genetics; D057809:HEK293 Cells; D006801:Humans; D007223:Infant; D007231:Infant, Newborn; D051938:Munc18 Proteins / Q000235:genetics / Q000378:metabolism; D009154:Mutation; D009474:Neurons / Q000378:metabolism / Q000473:pathology; D018377:Neurotransmitter Agents / Q000235:genetics / Q000378:metabolism; D013036:Spasms, Infantile / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D050827:Syntaxin 1 / Q000235:genetics / Q000378:metabolism
D051938:Munc18 Proteins; D018377:Neurotransmitter Agents; C497937:STXBP1 protein, human; D050827:Syntaxin 1