In this proposal, we plan to further delineate the molecular and cellular basis of SBMA disease pathogenesis. In our previously funded research grant, we sought the molecular and mechanistic basis of SBMA disease pathogenesis by defining the role of polyglutamine-expanded androgen receptor in the transcriptional dysregulation originally uncovered in our highly representative mouse model for SBMA. Among the crucial advances that we have made toward achieving this goal is our demonstration that androgen receptor (AR) normal transcription function is central to understanding SBMA disease pathogenesis. The discovery of a role for AR normal function in SBMA motor neuron disease suggests that AR may function as a trophic factor in motor neurons. Recent work has focused upon delineating which AR domains are involved in mediating AR protein neurotoxicity, and we have identified a key domain that enhances AR protein toxicity. In a parallel line of investigation, we have developed a new BAC transgenic mouse model for SBMA that features a floxed first exon to permit cell-type specific excision of mutant AR gene expression. Based upon these advances, we will determine the molecular and cellular basis of SBMA by pursuing three lines of investigation: First, we will identify the cell types responsible for SBMA disease pathogenesis in our floxed first exon BAC transgenic model of SBMA (fxAR121) by crossing fxAR121 mice with transgenic mice expressing muscle-specific or motor neuron-specific Cre recombinase;confirming excision of AR121 in targeted cell types;and determining if elimination of expression of mutant AR from muscle cells or motor neurons affects onset or progression of SBMA. Second, we will test if loss of AR normal function contributes to SBMA by performing combined RNA-Seq and ChIP-Seq analysis of MN-1 cells stably transfected with AR24Q protein under conditions of hypoxia in the presence or absence of DHT;comparing the two gene lists to identify primary target genes in the AR regulome that mediate protection from hypoxic stress in motor neurons;and measuring the level of expression of AR neuroprotection genes in presymptomatic and early symptomatic spinal cords of SBMA and ALS transgenic mice. Third, we will test if post-translation modification of lysine 385 in AR protein plays a key role in neurotoxicity by confirming that the K385 mutation results in proteotoxicity in a fly model of SBMA;determining if transgenic expression of normal Q length AR with the K385R mutation is sufficient to produce neurodegeneration in mice;testing if transgenic mice expressing polyQ-expanded AR with the K385R mutation display altered disease progression;and determining if SUMOylation alters turnover of AR protein, or affects native protein complex interactions of AR. These studies should provide insights into mechanisms of motor neuron disease, and could reveal targets for therapy development for SBMA and related motor neuron diseases.

Public Health Relevance

Studies of X-linked spinal &bulbar muscular atrophy (SBMA) and related motor neuron diseases have highlighted the importance of understanding the cellular and molecular basis of disease pathogenesis. In this project, we will examine the cellular basis of SBMA through the use of a powerful floxed first exon BAC transgenic mouse model, we will define how AR-regulated gene expression is neuroprotective in motor neurons, and we will determine if a post-translational modification of AR is fundamental to AR proteotoxicity using a combination of approaches in cell culture, Drosophila, and mouse models. These studies should provide insights into mechanisms of motor neuron disease, and could reveal targets for therapy development for SBMA and related motor neuron diseases.

Agency
National Institute of Health (NIH)
Institute
National Institute of Neurological Disorders and Stroke (NINDS)
Type
Research Project (R01)
Project #
5R01NS041648-12
Application #
8136010
Study Section
Cellular and Molecular Biology of Neurodegeneration Study Section (CMND)
Program Officer
Gubitz, Amelie
Project Start
2001-06-01
Project End
2015-08-31
Budget Start
2011-09-01
Budget End
2012-08-31
Support Year
12
Fiscal Year
2011
Total Cost
$331,531
Indirect Cost
Name
University of California San Diego
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
804355790
City
La Jolla
State
CA
Country
United States
Zip Code
92093
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