title: Early Golgi Abnormalities and Neurodegeneration upon Loss of Presynaptic Proteins Munc18-1, Syntaxin-1, or SNAP-25.
abstract: The loss of presynaptic proteins Munc18-1, syntaxin-1, or SNAP-25 is known to produce cell death, but the underlying features have not been compared experimentally. Here, we investigated these features in cultured mouse CNS and DRG neurons. Side-by-side comparisons confirmed massive cell death, before synaptogenesis, within 1-4 DIV upon loss of t-SNAREs (syntaxin-1, SNAP-25) or Munc18-1, but not v-SNAREs (synaptobrevins/VAMP1/2/3 using tetanus neurotoxin (TeNT), also in TI-VAMP/VAMP7 knock-out (KO) neurons). A condensedcis-Golgi was the first abnormality observed upon Munc18-1 or SNAP-25 loss within 3 DIV. This phenotype was distinct from the Golgi fragmentation observed in apoptosis. Cell death was too rapid after syntaxin-1 loss to study Golgi abnormalities. Syntaxin-1 and Munc18-1 depend on each other for normal cellular levels. We observed that endogenous syntaxin-1 accumulates at the Golgi of Munc18-1 KO neurons. However, expression of a non-neuronal Munc18 isoform that does not bind syntaxin-1, Munc18-3, in Munc18-1 KO neurons prevented cell death and restored normalcis-Golgi morphology, but not synaptic transmission or syntaxin-1 targeting. Finally, we observed that DRG neurons are the only Munc18-1 KO neurons that do not degeneratein vivoorin vitroIn these neurons,cis-Golgi abnormalities were less severe, with no changes in Golgi shape. Together, these data demonstrate that cell death upon Munc18-1, syntaxin-1, or SNAP-25 loss occurs via a degenerative pathway unrelated to the known synapse function of these proteins and involving earlycis-Golgi abnormalities, distinct from apoptosis.SIGNIFICANCE STATEMENTThis study provides new insights in a neurodegeneration pathway triggered by the absence of specific proteins involved in synaptic transmission (syntaxin-1, Munc18-1, SNAP-25), whereas other proteins involved in the same molecular process (synaptobrevins, Munc13-1/2) do not cause degeneration. Massive cell death occurs in cultured neurons upon depleting syntaxin-1, Munc18-1, and/or SNAP-25, well before synapse formation. This study characterizes several relevant cellular phenotypes, especially earlycis-Golgi abnormalities, distinct from abnormalities observed during apoptosis, and rules out several other phenotypes as causal (defects in syntaxin-1 targeting and synaptic transmission). As proteins, such as syntaxin-1, Munc18-1, or SNAP-25, modulate α-synuclein neuropathy and/or are dysregulated in Alzheimer's disease, understanding this type of neurodegeneration may provide new links between synaptic defects and neurodegeneration in humans.
authors.affiliation: Department of Functional Genomics and.
authors.forename: Tatiana C
authors.initials: TC
authors.lastname: Santos
authors.affiliation: Department of Functional Genomics and.
authors.forename: Keimpe
authors.identifier: 0000-0002-8784-9490
authors.initials: K
authors.lastname: Wierda
authors.affiliation: Department of Clinical Genetics, Center for Neurogenomics and Cognitive Research, VU University Amsterdam and VU Medical Center, 1081 HV Amsterdam, The Netherlands.
authors.forename: Jurjen H
authors.identifier: 0000-0002-4295-6938
authors.initials: JH
authors.lastname: Broeke
authors.affiliation: Department of Functional Genomics and.
authors.forename: Ruud F
authors.initials: RF
authors.lastname: Toonen
authors.affiliation: Department of Functional Genomics and matthijs@cncr.vu.nl.|Department of Clinical Genetics, Center for Neurogenomics and Cognitive Research, VU University Amsterdam and VU Medical Center, 1081 HV Amsterdam, The Netherlands.
authors.forename: Matthijs
authors.initials: M
authors.lastname: Verhage
mesh_terms: D000818:Animals; D017209:Apoptosis; D016923:Cell Death / Q000235:genetics; D002478:Cells, Cultured; D005089:Exocytosis / Q000235:genetics; D006056:Golgi Apparatus / Q000235:genetics* / Q000378:metabolism* / Q000473:pathology; D051379:Mice; D018345:Mice, Knockout; D051938:Munc18 Proteins / Q000235:genetics*; D019636:Neurodegenerative Diseases / Q000235:genetics* / Q000378:metabolism* / Q000473:pathology; D013569:Synapses / Q000473:pathology; D050825:Synaptosomal-Associated Protein 25 / Q000235:genetics*; D050827:Syntaxin 1 / Q000235:genetics*
chemical_list: D051938:Munc18 Proteins; C494209:Snap25 protein, mouse; D050825:Synaptosomal-Associated Protein 25; D050827:Syntaxin 1
keywords: exocytosis; neurodegeneration; presynaptic proteins; synaptic transmission