The BiP Molecular Chaperone Plays Multiple Roles during the Biogenesis of TorsinA, an AAA+ ATPase Associated with the Neurological Disease Early-onset Torsion Dystonia

Lucía F Zacchi,Hui-Chuan Wu,Samantha L Bell,Linda Millen,Adrienne W Paton,James C Paton,Philip J Thomas,Michal Zolkiewski,Jeffrey L Brodsky

doi:10.1074/jbc.m113.529123

Abstract

Early-onset torsion dystonia (EOTD) is a neurological disorder characterized by involuntary and sustained muscle contractions that can lead to paralysis and abnormal posture. EOTD is associated with the deletion of a glutamate (ΔE) in torsinA, an endoplasmic reticulum (ER) resident AAA(+) ATPase. To date, the effect of ΔE on torsinA and the reason that this mutation results in EOTD are unclear. Moreover, there are no specific therapeutic options to treat EOTD. To define the underlying biochemical defects associated with torsinAΔE and to uncover factors that might be targeted to offset defects associated with torsinAΔE, we developed a yeast torsinA expression system and tested the roles of ER chaperones in mediating the folding and stability of torsinA and torsinAΔE. We discovered that the ER lumenal Hsp70, BiP, an associated Hsp40, Scj1, and a nucleotide exchange factor, Lhs1, stabilize torsinA and torsinAΔE. BiP also maintained torsinA and torsinAΔE solubility. Mutations predicted to compromise specific torsinA functional motifs showed a synthetic interaction with the ΔE mutation and destabilized torsinAΔE, suggesting that the ΔE mutation predisposes torsinA to defects in the presence of secondary insults. In this case, BiP was required for torsinAΔE degradation, consistent with data that specific chaperones exhibit either pro-degradative or pro-folding activities. Finally, using two independent approaches, we established that BiP stabilizes torsinA and torsinAΔE in mammalian cells. Together, these data define BiP as the first identified torsinA chaperone, and treatments that modulate BiP might improve symptoms associated with EOTD.

Highlights

The ⌬E mutation in the AAAϩ ATPase torsinA is associated with the neurological disease torsion dystonia
TorsinA and torsinA⌬E were expressed at similar levels and migrated by SDS-PAGE as an ϳ33-kDa species, which corresponds to the monomeric forms of the proteins (Fig. 1C)
At pH 10.5, Pdi1 was almost completely released into the supernatant, whereas a considerable fraction of torsinA and torsinA⌬E remained associated with the pellet fraction, similar to Kar2/BiP (Fig. 2A, compare lanes 5 and 6 and 13 and 14)

Summary

Introduction

The ⌬E mutation in the AAAϩ ATPase torsinA is associated with the neurological disease torsion dystonia. Mutations predicted to compromise specific torsinA functional motifs showed a synthetic interaction with the ⌬E mutation and destabilized torsinA⌬E, suggesting that the ⌬E mutation predisposes torsinA to defects in the presence of secondary insults. In this case, BiP was required for torsinA⌬E degradation, consistent with data that specific chaperones exhibit either pro-degradative or pro-folding activities. Using two independent approaches, we established that BiP stabilizes torsinA and torsinA⌬E in mammalian cells Together, these data define BiP as the first identified torsinA chaperone, and treatments that modulate BiP might improve symptoms associated with EOTD

Methods

Results

Conclusion