Diminishing evidence for torsinA-positive neuronal inclusions in DYT1 dystonia
- Drew Pratt†1,
- Karin Mente†2Email authorView ORCID ID profile,
- Shervin Rahimpour3,
- Nancy A. Edwards4,
- Sule Tinaz2, 5,
- Brian D. Berman6,
- Mark Hallett2 and
- Abhik Ray-Chaudhury4
© The Author(s). 2016
Received: 22 July 2016
Accepted: 8 August 2016
Published: 17 August 2016
KeywordsDYT1 Dystonia TorsinA Inclusions
DYT1 dystonia, an early onset generalized dystonia, also known as Oppenheim’s dystonia, is an inherited isolated dystonia characterized by progressive generalized muscle spasms and sustained postures leading to significant disability . The disease is inherited in an autosomal dominant manner with incomplete penetrance (30–40 %) and typically presents in childhood . Patients harbor a 3-bp (GAG) deletion in the coding region of the TOR1A gene on chromosome 9q34 that encodes the protein torsinA . This deletion corresponds to loss of a single glutamate at amino acid residue 302 or 303 (torsinA ΔE) . The function of wildtype torsinA has been speculative, but relatively recent studies have demonstrated its involvement in protein trafficking, quality control, secretion, and degradation (for review, see Dauer 2014 ). The pathogenic mechanism leading to disease as a result of this deletion is thought to likely involve disruption of sensorimotor circuit development and function . Recent evidence also suggests striatal cholinergic dysfunction, or dysregulation, as a potential mechanism underlying the pathophysiology of DYT1 dystonia .
DYT1 dystonia neuropathology studies
Subcellular TorsinA immunoreactivity
Walker et al. 2002 
1 DYT1 Dystonia subject
Anti-torsinA (rabbit polyclonal: AA residues 323–332)
Nucleus, cytoplasm, and dendrites in both DYT1 dystonia and control groups
4 control subjects
Rostasy et al. 2003 
5 DYT1 Dystonia subjects
Anti-torsinA (rabbit polyclonal TAB1: AA residues 299–312; mouse monoclonal D-MG10: AA residues 208–249)
Cytoplasm, dendrites, and axons in both DYT1 dystonia and control groups
20 control subjects
McNaught et al. 2004 
4 DYT1 Dystonia subjects
Yes: midbrain, pons
Anti-torsinA (rabbit polyclonal; AA residues 323–332), Anti-ubiquitin, Anti-UPC, Anti-ChAT
Perinuclear and intranuclear inclusions in DYT1 dystonia group only
4 control subjects
Paudel et al. 2014 
7 DYT1 Dystonia subjects
Our results add to the growing body of evidence that there are no consistent torsinA immunoreactive protein inclusions associated with, or specific to, DYT1 dystonia in humans. In cell culture studies of neural cells transfected with mutant torsinA, cells demonstrated large perikaryal inclusions ultrastructurally composed of spheroid whorled membranes . The location of the collections appeared to be mutation status-related, with overexpression of mutant torsinA forming inclusions adjacent to the nuclear membrane and overexpressed wildtype protein aggregating within the cytoplasm. This latter feature is in keeping with our findings but was nonspecific and may possibly represent a response of wildtype torsinA to cellular stress (e.g., agonal state). The perinuclear accentuation of torsinA immunoreactivity we observed has previously been described in cultured cells exposed to oxidative stress . Lastly, it has been suggested that endogenous torsinA levels may be lower in vivo as compared to those in transfected cells due to overexpression in cell culture models, which may affect subcellular localization of torsinA .
Potential reasons for recurrent failure to demonstrate inclusions in human DYT1 tissue have been noted [5, 10, 12] and deserve mention here. Foremost, DYT1 dystonia is rare and tissue availability for sufficient sample size analysis is very limited; this is a limitation in the context of an incompletely penetrant disease, as disease-specific changes may be related to clinically apparent disease. Notably, while our evaluation was already limited to six DYT1 patients, only five included brainstem tissue (including the PPN, CN, and PAG) for evaluation. Second, studies involving human and animal tissues have failed to recapitulate the immunoreactive protein aggregates observed in culture cells overexpressing torsinA (see Dauer  for review), supporting the suspected lack of neurodegeneration in the disease. The human tissue studies that failed to identify torsinA inclusions used various anti-torsinA antibodies directed against different epitopes, which further supports that lack of inclusion body formation in DYT1 dystonia. Furthermore, there is strong evidence linking altered cholinergic striatal function, and its inputs from other structures such as the brainstem, cerebellum and thalamus, to the underlying pathophysiology in DYT1 dystonia (for review see Eskow Jaunarajs et al. ). Indeed, postmortem analyses of the putamen of DYT1 subjects have shown decreased levels of cholinergic markers .
In conclusion, there is little evidence supporting the presence of specific cellular morphologic changes in DYT1 dystonia at the light microscopic level in human tissue. The significance and specificity of the changes we observed are best addressed with larger postmortem studies combining histopathology and disease-specific cell models, such as patient-specific induced pluripotent stem cells (iPSCs). Use of iPSC-derived neurons would allow the subcellular localization of torsinA, and any inclusions, to be explored in living cells without altering the level of torsinA expression.
We are grateful for the contribution of the patients (and their families) who donated their tissues for research studies. The authors would like to thank Michael Feldman for performing the Western blot.
DP wrote the manuscript, contributed to the study design, performed the imaging analysis, and assisted in the histologic examination. KM co-wrote the manuscript, contributed to the study design, and assisted in performing the histologic examination. NE and SR performed the immunohistochemical and histochemical staining. ST, BDB, and MH conceptualized the study, gathered the materials, and edited the manuscript. ARC performed the histologic examination and conceptualized the study. All authors read and approved the final manuscript.
The authors declare that they have no competing interests.
This research was supported by the Intramural Research Program of the NIH, National Cancer Institute, Center for Cancer Research and the National Institute of Neurological Disorders and Stroke.
The NIH Office of Human Subject Research Protection has determined that this study is exempt from Institutional Review Board review.
The content of this publication does not necessarily reflect the views or policies of the Department of Health and Human Services, nor does mention of trade names, commercial products, or organizations imply endorsement by the US Government.
Open AccessThis article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.
- Greene P, Kang UJ, Fahn S. Spread of symptoms in idiopathic torsion dystonia. Mov Disord. 1995;10:143–52.View ArticlePubMedGoogle Scholar
- Risch NJ, Bressman SB, deLeon D, Brin MF, Burke RE, Greene PE, Shale H, Claus EB, Cupples LA, Fahn S. Segregation analysis of idiopathic torsion dystonia in Ashkenazi Jews suggests autosomal dominant inheritance. Am J Hum Genet. 1990;46:533–8.PubMedPubMed CentralGoogle Scholar
- Ozelius L, Kramer PL, Moskowitz CB, Kwiatkowski DJ, Brin MF, Bressman SB, Schuback DE, Falk CT, Risch N, de Leon D, et al. Human gene for torsion dystonia located on chromosome 9q32-q34. Neuron. 1989;2:1427–34.View ArticlePubMedGoogle Scholar
- Ozelius LJ, Hewett JW, Page CE, Bressman SB, Kramer PL, Shalish C, de Leon D, Brin MF, Raymond D, Corey DP, Fahn S, Risch NJ, Buckler AJ, Gusella JF, Breakefield XO. The early-onset torsion dystonia gene (DYT1) encodes an ATP-binding protein. Nat Genet. 1997;17:40–8.View ArticlePubMedGoogle Scholar
- Dauer W. Inherited isolated dystonia: clinical genetics and gene function. Neurotherapeutics. 2014;11:807–16.View ArticlePubMedPubMed CentralGoogle Scholar
- Liang CC, Tanabe LM, Jou S, Chi F, Dauer WT. TorsinA hypofunction causes abnormal twisting movements and sensorimotor circuit neurodegeneration. J Clin Invest. 2014;124:3080–92.View ArticlePubMedPubMed CentralGoogle Scholar
- Pappas SS, Darr K, Holley SM, Cepeda C, Mabrouk OS, Wong JM, LeWitt TM, Paudel R, Houlden H, Kennedy RT, Levine MS, Dauer WT. Forebrain deletion of the dystonia protein torsinA causes dystonic-like movements and loss of striatal cholinergic neurons. Elife. 2015;4:e08352.View ArticlePubMedPubMed CentralGoogle Scholar
- Oleas J, Yokoi F, DeAndrade MP, Pisani A, Li Y. Engineering animal models of dystonia. Mov Disord. 2013;28:990–1000. doi:https://doi.org/10.1002/mds.25583.View ArticlePubMedPubMed CentralGoogle Scholar
- McNaught KS, Kapustin A, Jackson T, Jengelley TA, Jnobaptiste R, Shashidharan P, Perl DP, Pasik P, Olanow CW. Brainstem pathology in DYT1 primary torsion dystonia. Ann Neurol. 2004;56(4):540–7.View ArticlePubMedGoogle Scholar
- Paudel R, Kiely A, Li A, Lashley T, Bandopadhyay R, Hardy J, Jinnah HA, Bhatia K, Houlden H, Holton JL. Neuropathological features of genetically confirmed DYT1 dystonia: investigating disease-specific inclusions. Acta Neuropathol Commun. 2014;2:159.View ArticlePubMedPubMed CentralGoogle Scholar
- Walker RH, Brin MF, Sandu D, Good PF, Shashidharan P. TorsinA immunoreactivity in brains of patients with DYT1 and non-DYT1 dystonia. Neurology. 2002;58:120–4.View ArticlePubMedGoogle Scholar
- Rostasy K, Augood SJ, Hewett JW, Leung JC, Sasaki H, Ozelius LJ, Ramesh V, Standaert DG, Breakefield XO, Hedreen JC. TorsinA protein and neuropathology in early onset generalized dystonia with GAG deletion. Neurobiol Dis. 2003;12:11–24.View ArticlePubMedGoogle Scholar
- Hewett J, Gonzalez-Agosti C, Slater D, Ziefer P, Li S, Bergeron D, Jacoby DJ, Ozelius LJ, Ramesh V, Breakefield XO. Mutant torsinA, responsible for early-onset torsion dystonia, forms membrane inclusions in cultured neural cells. Hum Mol Genet. 2000;9:1403–13.View ArticlePubMedGoogle Scholar
- Hewett J, Ziefer P, Bergeron D, Naismith T, Boston H, Slater D, Wilbur J, Schuback D, Kamm C, Smith N, Camp S, Ozelius LJ, Ramesh V, Hanson PI, Breakefield XO. TorsinA in PC12 cells: localization in the endoplasmic reticulum and response to stress. J Neurosci Res. 2003;72:158–68.View ArticlePubMedGoogle Scholar
- Harata NC. Current gaps in the understanding of the subcellular distribution of exogenous and endogenous protein TorsinA. Tremor Other Hyperkinet Mov (N Y). 2014;4:260.Google Scholar
- Eskow Jaunarajs KL, Bonsi P, Chesselet MF, Standaert DG, Pisani A. Striatal cholinergic dysfunction as a unifying theme in the pathophysiology of dystonia. Prog Neurobiol. 2015;127–128:91–107.View ArticlePubMedGoogle Scholar