Identification of CSPα Clients Reveals a Role in Dynamin 1 Regulation

Abstract

SUMMARY Cysteine string protein α (CSPα), a presynaptic co-chaperone for Hsc70, is required for synapse maintenance. Deletion of CSPα leads to neuronal dysfunction, synapse loss, and neurodegeneration. We utilized unbiased, systematic proteomics to identify putative CSPα protein clients. We found 22 such proteins whose levels are selectively decreased in CSPα knockout synapses. Of these putative CSPα protein clients, two directly bind to the CSPα chaperone complex and are bona fide clients. They are the t-SNARE SNAP-25 and the GTPase dynamin 1, which are necessary for synaptic vesicle fusion and fission, respectively. Using hippocampal cultures, we show CSPα regulates the stability of client proteins and synaptic vesicle number. Our analysis of CSPα-dynamin 1 interactions reveals unexpectedly that CSPα regulates the polymerization of dynamin 1. CSPα therefore participates in synaptic vesicle endocytosis and may facilitate exo- and endocytic coupling. These findings advance the understanding of how synapses are functionally and structurally maintained.

Phosphomimetic Mutation of Cysteine String Protein-α Increases the Rate of Regulated Exocytosis by Modulating Fusion Pore Dynamics in PC12 Cells

“…In addition to conventional SNARE proteins and Syt isoforms, CSPα has been shown to interact with dynamin I, a protein essential for vesicle fission, suggesting that CSPα may regulate synaptic vesicle endocytosis and facilitate exo- and endocytotic coupling [29]. Consistent with this finding, our results suggest that phosphorylation of CSPα may regulate fusion pore dynamics.…”

Monomeric Synucleins Generate Membrane Curvature

“…(v) ␣-Synuclein is a component of clathrincoated vesicles (66). (vi) Overexpression of ␣-synuclein rescues the synapse loss seen in knock-out mice for the co-chaperone CSP␣ (28) which participates in both exo-and endocytosis at the presynaptic terminal (31). Collectively, the above findings support the hypothesis that synucleins are participatants in the exo-and endocytic steps of the synaptic vesicle cycle.…”

“…Mass Spectrometry-A quantitative analysis of the synaptic proteome of wild type and ␣␤␥-synuclein KO mice was performed using DIGE according to previously published protocols (30,31). Equal amounts of protein from wild type and ␣␤␥-synuclein KO samples were differentially labeled in vitro with Cy3 and Cy5 N-hydroxysuccinimidyl ester dyes and separated on two-dimensional gels.…”

Latrophilins Function as Heterophilic Cell-adhesion Molecules by Binding to Teneurins

“…Therefore, we performed quantitative RT-PCR assays to detect the mRNA levels of 20 genes encoding neuronal proteins comprising cell-adhesion molecules, SNARE proteins, and others. Cell-adhesion molecule targets were selected on the basis that they do not constitute the group of described CSP␣ clients, therefore excluding a direct contribution of CSP␣-chaperoning function (43).…”