Cysteine String Protein (CSP) Inhibition of N-type Calcium Channels Is Blocked by Mutant Huntingtin

Linda C Miller,Leigh Anne Swayne,Lina Chen,Zhong-Ping Feng,Jennifer L Wacker,Paul J Muchowski,Gerald W Zamponi,Janice E.A Braun

doi:10.1074/jbc.m306230200

Linda C Miller, Leigh Anne Swayne + Show 6 more

Open Access

https://doi.org/10.1074/jbc.m306230200

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Abstract

Cysteine string protein (CSP), a 34-kDa molecular chaperone, is expressed on synaptic vesicles in neurons and on secretory vesicles in endocrine, neuroendocrine, and exocrine cells. CSP can be found in a complex with two other chaperones, the heat shock cognate protein Hsc70, and small glutamine-rich tetratricopeptide repeat domain protein (SGT). CSP function is vital in synaptic transmission; however, the precise nature of its role remains controversial. We have previously reported interactions of CSP with both heterotrimeric GTP-binding proteins (G proteins) and N-type calcium channels. These associations give rise to a tonic G protein inhibition of the channels. Here we have examined the effects of huntingtin fragments (exon 1) with (huntingtin(exon1/exp)) and without (huntingtin(exon1/nonexp)) expanded polyglutamine (polyQ) tracts on the CSP chaperone system. In vitro huntingtin(exon1/exp) sequestered CSP and blocked the association of CSP with G proteins. In contrast, huntingtin(exon1/nonexp) did not interact with CSP and did not alter the CSP/G protein association. Similarly, co-expression of huntingtin(exon1/exp) with CSP and N-type calcium channels eliminated CSP's tonic G protein inhibition of the channels, while coexpression of huntingtin(exon1/nonexp) did not alter the robust inhibition promoted by CSP. These results indicate that CSP's modulation of G protein inhibition of calcium channel activity is blocked in the presence of a huntingtin fragment with expanded polyglutamine tracts.

Highlights

Cysteine string protein (CSP), a 34-kDa molecular chaperone, is expressed on synaptic vesicles in neurons and on secretory vesicles in endocrine, neuroendocrine, and exocrine cells
Exon 1 of Huntingtin with an Expanded Polyglutamine Tract Blocks the CSP Interaction with G Proteins—In order to investigate the possibility that mutant forms of huntingtin with expanded glutamine repeats alter the association of G protein with CSP, a GST fusion protein consisting of full-length CSP was coupled to glutathione-agarose beads and used in an in vitro binding assay
These results indicate that CSP modulation of G protein inhibition of calcium channel activity is blocked in the presence of huntingtin fragments with expanded polyglutamine tracts, suggesting that CSP becomes functionally inactivated in the presence of huntingtinexon1/exp

Summary

Introduction

Cysteine string protein (CSP), a 34-kDa molecular chaperone, is expressed on synaptic vesicles in neurons and on secretory vesicles in endocrine, neuroendocrine, and exocrine cells. These results indicate that CSP’s modulation of G protein inhibition of calcium channel activity is blocked in the presence of a huntingtin fragment with expanded polyglutamine tracts. Given that proteins with expanded polyglutamine repeats have been proposed to interfere with the chaperone balance of the cell, we have analyzed the effects of huntingtin on CSP modulation of N-type channels.

Results

Conclusion