TheN-Ethylmaleimide-Sensitive Factor and Dysbindin Interact To Modulate Synaptic Plasticity

Abstract

Dysbindin is a schizophrenia susceptibility factor and subunit of the biogenesis of lysosome-related organelles complex 1 (BLOC-1) required for lysosome-related organelle biogenesis, and in neurons, synaptic vesicle assembly, neurotransmission, and plasticity. Protein networks, or interactomes, downstream of dysbindin/BLOC-1 remain partially explored despite their potential to illuminate neurodevelopmental disorder mechanisms. Here, we conducted a proteome-wide search for polypeptides whose cellular content is sensitive to dysbindin/BLOC-1 loss of function. We identified components of the vesicle fusion machinery as factors downregulated in dysbindin/ BLOC-1 deficiency in neuroectodermal cells and iPSC-derived human neurons, among them the N-ethylmaleimide-sensitive factor (NSF). Human dysbindin/BLOC-1 coprecipitates with NSF and vice versa, and both proteins colocalized in a Drosophila model synapse. To test the hypothesis that NSF and dysbindin/BLOC-1 participate in a pathway-regulating synaptic function, we examined the role for NSF in dysbindin/BLOC-1-dependent synaptic homeostatic plasticity in Drosophila. As previously described, we found that mutations in dysbindin precluded homeostatic synaptic plasticity elicited by acute blockage of postsynaptic receptors. This dysbindin mutant phenotype is fully rescued by presynaptic expression of either dysbindin or Drosophila NSF. However, neither reduction of NSF alone or in combination with dysbindin haploinsufficiency impaired homeostatic synaptic plasticity. Our results demonstrate that dysbindin/ BLOC-1 expression defects result in altered cellular content of proteins of the vesicle fusion apparatus and therefore influence synaptic plasticity.

Transmission, Development, and Plasticity of Synapses

“…Genetic interaction experiments support the idea that the BLOC-1 components Snapin and Blos-1 cooperate with Dysb to mediate homeostatic plasticity in Drosophila (Dickman et al 2012;Mullin et al 2015), and like Dysb, Snapin localizes to SVs (Dickman et al 2012). The mechanism of how Dysb and its interactors function in homeostatic compensation is unclear, but might involve regulation of SNARE-mediated fusion events, as BLOC-1 components have been shown to interact with SNAREs and NSF (Ghiani et al 2010;Gokhale et al 2015), and overexpression of NSF rescues the ability to respond to PhTx in dysb mutants (Gokhale et al 2015).…”

Recent Advances in Deciphering the Structure and Molecular Mechanism of the AAA+ ATPase N-Ethylmaleimide-Sensitive Factor (NSF)

“…In addition to its role as a SNARE-complex disassembly machine, NSF also interacts with other cellular proteins besides the SNARE complexes [18–21]. However, the possible function and mechanisms involving these interactions are unclear.…”

Rare Disease Mechanisms Identified by Genealogical Proteomics of Copper Homeostasis Mutant Pedigrees

“…Fibroblasts were labeled using the protocol described below and as cited (Gokhale et al, 2016; Gokhale et al, 2012; Gokhale et al, 2015; Ong et al, 2002; Ong and Mann, 2006; Ryder et al, 2013). Cells were grown in DMEM with either “light” unlabeled 12 C- and 14 N- arginine and lysine amino acids (R0K0).…”

“…Cell lysates were prepared, as described below. Mass spectrometry was performed using the services of MS Bioworks (Gokhale et al, 2016; Gokhale et al, 2015; Perez-Cornejo et al, 2012; Ryder et al, 2013). The clarified SILAC labeled supernatants were pooled 1:1:1 and 20μg of this mix was separated on a 4–12% Bis-Tris Novex mini-gel (Invitrogen) using the MOPS buffer system.…”