The Dystonia-associated Protein TorsinA Modulates Synaptic Vesicle Recycling

Abstract

The loss of a glutamic acid residue in the AAA-ATPase (ATPases associated with diverse cellular activities) torsinA is responsible for most cases of early onset autosomal dominant primary dystonia. In this study, we found that snapin, which binds SNAP-25 (synaptosome-associated protein of 25,000 Da) and enhances the association of the SNARE complex with synaptotagmin, is an interacting partner for both wild type and mutant torsinA. Snapin co-localized with endogenous torsinA on dense core granules in PC12 cells and was recruited to perinuclear inclusions containing mutant ⌬E-torsinA in neuroblastoma SH-SY5Y cells. In view of these observations, synaptic vesicle recycling was analyzed using the lipophilic dye FM1-43 and an antibody directed against an intravesicular epitope of synaptotagmin I. We found that overexpression of wild type torsinA negatively affects synaptic vesicle endocytosis. Conversely, overexpression of ⌬E-torsinA in neuroblastoma cells increases FM1-43 uptake. Knockdown of snapin and/or torsinA using small interfering RNAs had a similar inhibitory effect on the exo-endocytic process. In addition, down-regulation of torsinA causes the persistence of synaptotagmin I on the plasma membrane, which closely resembles the effect observed by the overexpression of the ⌬E-torsinA mutant. Altogether, these findings suggest that torsinA plays a role together with snapin in regulated exocytosis and that ⌬E-torsinA exerts its pathological effects through a loss of function mechanism. This may affect neuronal uptake of neurotransmitters, such as dopamine, playing a role in the development of dystonic movements.The majority of cases of early onset, primary torsion dystonia are caused by a dominantly inherited mutation in the DYT1 (TOR1A) gene on chromosome 9q34 (1). DYT1 dystonia manifests in childhood, typically with dystonia in a limb that spreads to the trunk and other limbs, usually sparing cranio-cervical muscles (2, 3). There is no evidence for neurodegeneration in DYT1 dystonia, implying that abnormal movements are caused by a functional neuronal defect (4). All cases of typical DYT1 dystonia are caused by an in-frame GAG deletion (⌬GAG302/ 303; ⌬E) in DYT1 gene, resulting in the loss of a glutamic acid in the C-terminal region of the encoded protein, torsinA (1).TorsinA is a member of the AAA ATPase superfamily of chaperone-like proteins (5). In mammalian neuronal cells, torsinA is found throughout the cytoplasm, neurite processes, and growth cones (6, 7). TorsinA has also been found in the lumen of the endoplasmic reticulum (ER) 3 and in the space between the inner and the outer membrane of the nuclear envelope (NE) (8 -11). In contrast, in cells overexpressing the mutant (⌬E-torsinA), the protein is redistributed from ER to NE and accumulates in large perinuclear membranous inclusions, which appeared to arise from the nuclear envelope (7,(12)(13)(14). TorsinA-positive inclusions have been found in the midbrain of DYT1 patients, suggesting that they are relevant to the pathogenesis of DYT1 dystonia (15...

Recent Advances in the Molecular Pathogenesis of Dystonia-Plus Syndromes and Heredodegenerative Dystonias

“…This is demonstrated by DRD, DAT gene related dystonia-parkinsonism and LND. In DYT1 dystonia there is also evidence to support a dopaminergic hypothesis with abnormal synaptic vesicle function [83,84]. Other forms of primary dystonia, including DYT6 , have been associated with reduced striatal D2 receptor availability, and it has been suggested this may lead to dystonic movements by affecting the indirect pathway of cortical-basal ganglia circuits.…”

Printor, a Novel TorsinA-interacting Protein Implicated in Dystonia Pathogenesis

“…Although torsinA is generally thought to be a ER lumenal protein, a small population of torsinA has been observed in the cytosol (10,70) and a significant pool of torsinA has been shown to have the AAA ϩ domain facing the cytoplasm (20). Moreover, torsinA has been reported to bind several cytosolic proteins (20,27,71). Thus, it is possible that a small pool of printor may interact with torsinA in the cytosol and regulate cytosolic protein folding.…”

“…Several binding partners of torsinA have been reported, including LAP1 and its homologous protein LULL1 (21), nesprins (22), snapin (27), and dopamine transporter (33). These torsinA binding partners have been proposed to be substrates of torsinA (20 -22, 27, 33), and some of these proteins have been shown to preferentially bind to the ATP-bound form of torsinA (21,22,33).…”

“…In support of this topology, torsinA is found in the cytoplasm, neuronal processes, and synaptic terminals (2,3,15,(23)(24)(25)(26) and has been shown to bind cytosolic proteins snapin (27) and kinesin light chain 1 (20). TorsinA has been proposed to play a role in several cellular processes, including dopaminergic neurotransmission (28 -31), NE organization and dynamics (17,22,32), and protein trafficking (27,33). However, the precise biological function of torsinA and its regulation remain unknown.…”

TorsinA and the Pathophysiology of DYT1 Dystonia

“…TorsinA might, however, play more than one role in cells. In particular, several lines of evidence suggest that torsinA might be part of the molecular machinery required to maintain nuclear envelope architecture and position, and to regulate neurite extension, protein secretion, and synaptic vesicle transport (FerrariToninelli et al, 2004;Hewett et al, 2006;Granata et al, 2008;Hewett et al, 2008).…”