Huntington's disease is a neurodegenerative disorder caused by an expanding CAG repeat coding for polyglutamine in the huntingtin gene. In the previous grant period we established and studied a transgenic mouse model of HD using an N-terminal fragment of huntingtin. We have now developed a new inducible transgenic model of HD expressing full-length huntingtin with an N-terminal myc tag to facilitate biochemical and cell biological studies. Preliminary data indicate that the mice have a robust progressive behavioral phenotype and characteristic pathology, as well as the presence of an N-terminal fragment of huntingtin. We believe proteolytic cleavage may be a critical event in HD pathogenesis. We study this HD model in parallel with a mouse model of the closely related polyglutamine disorder DRPLA, in which proteolytic processing appears to be critical for pathogenesis.
In specific aim 1 we will breed sufficient numbers of these HD inducible transgenic mice to characterize the time course of the behavioral phenotype and to provide tissue for pathological and biochemical studies.
In specific aim 2 we will define the pathology of the mice, including the regional distribution of intranuclear inclusions and other aggregates of the huntingtin protein.
In specific aim 3 we will study the time course and regional and cellular distribution of the huntingtin fragment. We will determine its length and use methods of protein purification and mass spectrometry to determine the cleavage site. We will develop a cell model with stable inducible expression of the same mutant huntingtin construct for comparative in vitro studies.
In specific aim 4 we will generate new transgenic mice with alterations in the huntingtin protein sequence to eliminate proteolytic cleavage. We predict that these mice will have a substantially decreased HD phenotype. Taken together these studies will establish cleavage as a key event in HD pathogenesis, and as a novel and accessible target for therapeutics. The new mouse model will be useful for studying pathogenic features of HD, and for testing experimental therapeutic interventions.

Agency
National Institute of Health (NIH)
Institute
National Institute of Neurological Disorders and Stroke (NINDS)
Type
Research Project (R01)
Project #
5R01NS038144-08
Application #
7087786
Study Section
Special Emphasis Panel (ZRG1-BDCN-4 (01))
Program Officer
Sutherland, Margaret L
Project Start
1998-12-01
Project End
2007-06-30
Budget Start
2006-07-01
Budget End
2007-06-30
Support Year
8
Fiscal Year
2006
Total Cost
$369,209
Indirect Cost
Name
Johns Hopkins University
Department
Psychiatry
Type
Schools of Medicine
DUNS #
001910777
City
Baltimore
State
MD
Country
United States
Zip Code
21218
Ratovitski, Tamara; Gucek, Marjan; Jiang, Haibing et al. (2009) Mutant huntingtin N-terminal fragments of specific size mediate aggregation and toxicity in neuronal cells. J Biol Chem 284:10855-67
Masuda, Naoki; Peng, Qi; Li, Qing et al. (2008) Tiagabine is neuroprotective in the N171-82Q and R6/2 mouse models of Huntington's disease. Neurobiol Dis 30:293-302
Ratovitski, Tamara; Nakamura, Masayuki; D'Ambola, James et al. (2007) N-terminal proteolysis of full-length mutant huntingtin in an inducible PC12 cell model of Huntington's disease. Cell Cycle 6:2970-81
Schilling, Gabriele; Klevytska, Alexandra; Tebbenkamp, Andrew T N et al. (2007) Characterization of huntingtin pathologic fragments in human Huntington disease, transgenic mice, and cell models. J Neuropathol Exp Neurol 66:313-20
Schilling, Birgit; Gafni, Juliette; Torcassi, Cameron et al. (2006) Huntingtin phosphorylation sites mapped by mass spectrometry. Modulation of cleavage and toxicity. J Biol Chem 281:23686-97
Schilling, Gabriele; Savonenko, Alena V; Klevytska, Alexandra et al. (2004) Nuclear-targeting of mutant huntingtin fragments produces Huntington's disease-like phenotypes in transgenic mice. Hum Mol Genet 13:1599-610
Goti, Daniel; Katzen, Scott M; Mez, Jesse et al. (2004) A mutant ataxin-3 putative-cleavage fragment in brains of Machado-Joseph disease patients and transgenic mice is cytotoxic above a critical concentration. J Neurosci 24:10266-79
Ross, Christopher A; Poirier, Michelle A (2004) Protein aggregation and neurodegenerative disease. Nat Med 10 Suppl:S10-7
Nucifora Jr, Frederick C; Ellerby, Lisa M; Wellington, Cheryl L et al. (2003) Nuclear localization of a non-caspase truncation product of atrophin-1, with an expanded polyglutamine repeat, increases cellular toxicity. J Biol Chem 278:13047-55
Wheeler, Vanessa C; Gutekunst, Claire-Anne; Vrbanac, Vladimir et al. (2002) Early phenotypes that presage late-onset neurodegenerative disease allow testing of modifiers in Hdh CAG knock-in mice. Hum Mol Genet 11:633-40

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