Molecular defects of the dystonia-causing torsinA mutation

Pham, Phuong Ngoc; Frei, Karen; Woo, William; Truong, Daniel

doi:10.1097/wnr.0b013e3280101220

Cited by 25 publications

(19 citation statements)

References 24 publications

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“…Secondly, as Zhu and others have pointed out, previous studies that have investigated the ATPase activity of WT torsinA and torsinA (ΔE) have been contradictory and did not have an ATP hydrolysis mutant as a negative control. These studies have also shown much lower rates of activity than that of other AAA+ proteins as well as conflicting results between the ATPase activity of WT torsinA and torsinA (ΔE) (Kustedjo et al 2000;Konakova and Pulst 2005;Pham et al 2006).…”

Section: Discussionmentioning

confidence: 99%

The early-onset torsion dystonia-associated protein, torsinA, displays molecular chaperone activity in vitro

Burdette

Churchill

Caldwell

et al. 2010

Cell Stress and Chaperones

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TorsinA is a member of the AAA+ ATPase family of proteins and, notably, is the only known ATPase localized to the ER lumen. It has been suggested to act as a molecular chaperone, while a mutant form associated with early-onset torsion dystonia, a dominantly inherited movement disorder, appears to result in a net loss of function in vivo. Thus far, no studies have examined the chaperone activity of torsinA in vitro. Here we expressed and purified both wild-type (WT) and mutant torsinA fusion proteins in bacteria and examined their ability to function as molecular chaperones by monitoring suppression of luciferase and citrate synthase (CS) aggregation. We also assessed their ability to hold proteins in an intermediate state for refolding. As measured by light scattering and SDS-PAGE, both WT and mutant torsinA effectively, and similarly, suppressed protein aggregation compared to controls. This function was not further enhanced by the presence of ATP. Further, we found that while neither form of torsinA could protect CS from heat-induced inactivation, they were both able to reactivate luciferase when ATP and rabbit reticulocyte lysate were added. This suggests that torsinA holds luciferase in an intermediate state, which can then be refolded in the presence of other chaperones. These data provide conclusive evidence that torsinA acts as a molecular chaperone in vitro and suggests that early-onset torsion dystonia is likely not a consequence of a loss in torsinA chaperone activity but might be an outcome of insufficient torsinA localization at the ER to manage protein folding or trafficking.

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Section: Discussionmentioning

confidence: 99%

The early-onset torsion dystonia-associated protein, torsinA, displays molecular chaperone activity in vitro

Burdette

Churchill

Caldwell

et al. 2010

Cell Stress and Chaperones

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“…Mutant torsinA might decrease chaperone function simply by being inactive, resulting in haploinsufficiency, but this does not seem to be the case because heterozygote knockout cells had near wild-type levels of Gluc secretion. Rather, ⌬E-torsinA appears to inhibit torsinA activity, possibly by forming inactive multimers with torsinA (41,42). Mutant ⌬E-torsinA protein might also sensitize DYT1 cells to the UPR, which can restrict protein processing in the ER (31,43).…”

Section: Resultsmentioning

confidence: 99%

Mutant torsinA interferes with protein processing through the secretory pathway in DYT1 dystonia cells

Hewett

Tannous

Niland

et al. 2007

Proc. Natl. Acad. Sci. U.S.A.

132

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TorsinA is an AAA ؉ protein located predominantly in the lumen of the endoplasmic reticulum (ER) and nuclear envelope responsible for early onset torsion dystonia (DYT1). Most cases of this dominantly inherited movement disorder are caused by deletion of a glutamic acid in the carboxyl terminal region of torsinA. We used a sensitive reporter, Gaussia luciferase (Gluc) to evaluate the role of torsinA in processing proteins through the ER. In primary fibroblasts from controls and DYT1 patients most Gluc activity (95%) was released into the media and processed through the secretory pathway, as confirmed by inhibition with brefeldinA and nocodazole. Fusion of Gluc to a fluorescent protein revealed coalignment and fractionation with ER proteins and association of Gluc with torsinA. Notably, fibroblasts from DYT1 patients were found to secrete markedly less Gluc activity as compared with control fibroblasts. This decrease in processing of Gluc in DYT1 cells appear to arise, at least in part, from a loss of torsinA activity, because mouse embryonic fibroblasts lacking torsinA also had reduced secretion as compared with control cells. These studies demonstrate the exquisite sensitivity of this reporter system for quantitation of processing through the secretory pathway and support a role for torsinA as an ER chaperone protein.early onset dystonia ͉ endoplasmic reticulum ͉ luciferase ͉ protein translation

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“…The molecular etiology of EOTD is believed to result from a dominant negative loss of torsinA activity mediated by the mutant protein (Pham et al, 2006); thus, molecules that function to elevate the activity of the remaining WT torsinA within patients may represent the best candidates for therapy (i.e. quinolone and blactam antibiotics).…”

Section: Structure-activity Relationships Between Torsina Effector Momentioning

confidence: 99%

“…Mutant torsinA (DE) exhibits reduced ATPase activity as compared with WT torsinA, and the mutant form inhibits WT ATPase activity in mixed protein preparations in vitro (Pham et al, 2006). It has been further demonstrated, in cell culture, that the absence of the glutamic acid (DE) residue in torsinA can result in the formation of aberrant membranous inclusions and increased subcellular distribution of this protein at the nuclear envelope (NE) Chemical enhancement of torsinA function in cell and animal models of torsion dystonia (Gonzalez-Alegre and Paulson, 2004;Goodchild and Dauer, 2004;Naismith et al, 2004).…”

Section: Introductionmentioning

confidence: 99%

Chemical enhancement of torsinA function in cell and animal models of torsion dystonia

Cao

Hewett

Yokoi³

et al. 2010

Disease Models &Amp; Mechanisms

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SUMMARYMovement disorders represent a significant societal burden for which therapeutic options are limited and focused on treating disease symptomality. Early-onset torsion dystonia (EOTD) is one such disorder characterized by sustained and involuntary muscle contractions that frequently cause repetitive movements or abnormal postures. Transmitted in an autosomal dominant manner with reduced penetrance, EOTD is caused in most cases by the deletion of a glutamic acid (DE) in the DYT1 (also known as TOR1A) gene product, torsinA. Although some patients respond well to anticholingerics, therapy is primarily limited to either neurosurgery or chemodenervation. As mutant torsinA (DE) expression results in decreased torsinA function, therapeutic strategies directed toward enhancement of wild-type (WT) torsinA activity in patients who are heterozygous for mutant DYT1 may restore normal cellular functionality. Here, we report results from the first-ever screen for candidate small molecule therapeutics for EOTD, using multiple activity-based readouts for torsinA function in Caenorhabditis elegans, subsequent validation in human DYT1 patient fibroblasts, and behavioral rescue in a mouse model of DYT1 dystonia. We exploited the nematode to rapidly discern chemical effectors of torsinA and identified two classes of antibiotics, quinolones and aminopenicillins, which enhance WT torsinA activity in two separate in vivo assays. Representative molecules were assayed in EOTD patient fibroblasts for improvements in torsinA-dependent secretory function, which was improved significantly by ampicillin. Furthermore, a behavioral defect associated with an EOTD mouse knock-in model was also rescued following administration of ampicillin. These combined data indicate that specific small molecules that enhance torsinA activity represent a promising new approach toward therapeutic development for EOTD, and potentially for other diseases involving the processing of mutant proteins.

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Molecular defects of the dystonia-causing torsinA mutation

Cited by 25 publications

References 24 publications

The early-onset torsion dystonia-associated protein, torsinA, displays molecular chaperone activity in vitro

The early-onset torsion dystonia-associated protein, torsinA, displays molecular chaperone activity in vitro

Mutant torsinA interferes with protein processing through the secretory pathway in DYT1 dystonia cells

Chemical enhancement of torsinA function in cell and animal models of torsion dystonia

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