Childhood onset dystonia is an autosomal dominant movement disorder that has been linked to a three base pair (GAG) deletion in the DYT1 gene localized to chromosome 9. The DYT1 gene encodes a novel ATP binding protein of 332 amino acid residues called torsinA (Ozelius et al, 1997). We have recently demonstrated that torsinA indeed binds to ATP and is expressed in brain and other peripheral tissues (Shashidharan et al, 2000a). In the brain, torsinA expression is restricted to neurons as shown by in situ hybridization (Augood et al, 1999) and immunohistochemical techniques (Shashidharan et al, 2000a; Walker et al., 2001a). Neither the cellular function of torsinA nor its role in dystonia is known. In order to better understand the cellular function of torsinA and its role in childhood-onset dystonia, we have: a) developed transgenic mice expressing human mutant (deltaE)-torsinA, b) generated PC12 clonal cell lines expressing normal and (deltaE)-torsinA, c) developed and characterized polyclonal and monoclonal antibodies to human torsinA d) cloned and characterized normal and deltaE-torsinA cDNAs, e) generated fusion proteins with GST and GFP, f) obtained postmortem brain tissue from a patient with childhood-onset dystonia, g) established a program to obtain postmortem brains by rapid autopsy from patients with dystonia. Equipped with these we have begun investigations into the cellular function of torsinA and its role in childhood onset dystonia. Our long-term objective is to better understand the pathogenesis and cellular dysfunction in DYT1 dystonia and develop rational methods for therapeutic intervention. In this grant proposal we will carry out the following specific aims:
AIM 1 : To characterize DYT1 transgenic mice expressing human torsinA by biochemical, molecular and immunohistochemical techniques and behavioral studies. Our preliminary studies revealed that 35 percent of transgenic mice expressing deltaE-torsinA develop abnormal hyperkinetic motor function. The symptoms develop between 3-10 weeks of life in mice. Interestingly the age of onset and penetrance level exhibited by transgenic mice is comparable to that of the human disorder. The age of onset in humans is between ages 5-12 and the penetrance level is 30-40 percent.
AIM 2 : To investigate the biochemical properties of torsinA. Our preliminary studies utilizing co-immunoprecipitation techniques revealed that torsinA may interact with other proteins. Whether these interacting proteins are necessary for the biological function of torsinA remains to be further elucidated.
AIM 3 : To isolate and identify protein partners of torsinA. TorsinA belongs to AAA+ family, a class of proteins associated with complex assembly, operation and disassembly of cellular proteins. The carboxy terminal region of torsinA, that undergoes a point mutation in DYT1 dystonia, is thought to be involved in either formation of an oligomeric ring structure or in an interaction with other proteins. Our initial studies described in Specific aim 2 will provide us with insights into the presence of interacting proteins with torsinA, which will be further investigated by yeast two-hybrid system.

Agency
National Institute of Health (NIH)
Institute
National Institute of Neurological Disorders and Stroke (NINDS)
Type
Research Project (R01)
Project #
5R01NS043038-02
Application #
6625759
Study Section
Special Emphasis Panel (ZRG1-BDCN-5 (01))
Program Officer
Sheehy, Paul A
Project Start
2002-04-15
Project End
2007-01-31
Budget Start
2003-02-01
Budget End
2004-01-31
Support Year
2
Fiscal Year
2003
Total Cost
$281,794
Indirect Cost
Name
Mount Sinai School of Medicine
Department
Neurology
Type
Schools of Medicine
DUNS #
078861598
City
New York
State
NY
Country
United States
Zip Code
10029
Shashidharan, Pullanipally; Plaitakis, Andreas (2014) The discovery of human of GLUD2 glutamate dehydrogenase and its implications for cell function in health and disease. Neurochem Res 39:460-70
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Shashidharan, P; Paris, Nicolae; Sandu, Daniela et al. (2004) Overexpression of torsinA in PC12 cells protects against toxicity. J Neurochem 88:1019-25
Torres, Gonzalo E; Sweeney, Ava L; Beaulieu, Jean-Martin et al. (2004) Effect of torsinA on membrane proteins reveals a loss of function and a dominant-negative phenotype of the dystonia-associated DeltaE-torsinA mutant. Proc Natl Acad Sci U S A 101:15650-5

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