The discovery of major aggregated proteins in neurodegenerative diseases provides the first insight into disease pathogenesis. Recently, the dominant protein component within ubiquitin positive inclusions of frontotemporal lobar degeneration (FTLD-U) and sporadic amyotrophic lateral sclerosis (ALS) was found to be TAR DNA-binding protein 43 (TDP-43). The common pathology observed in these diseases suggests parallel approaches to treatment based on the understanding of TDP-43 aggregation events. In normal cells TDP-43 is a nuclear RNA binding protein involved transcriptional regulation. However, pathologic TDP-43 has been reported to redistribute from the nucleus to the cytoplasm where it is aggregated, phosphorylated, ubiquitinated and/or cleaved. Our preliminary results show that over-expressed human TDP-43 and the alternative splicing isoform (TDP-S6) are phosphorylated, ubiquitinated and cleaved in mammalian cell cultures. The expressed TDP-43 is localized diffusely and almost exclusively in the nucleus. In contrast, the TDP-S6 isoform is predominantly aggregated in the cytoplasm, recapitulating the disease-specific pathological hallmark. Therefore, our hypothesis is that TDP-43 aggregation and toxicity depends on the specific production of spliced and/or posttranslationally modified TDP-43 isoforms (i.e. cleavage, phosphorylation and ubiquitination). To test this hypothesis,we will investigate the expression of spliced TDP-43 isoforms in normal brain and FTLD-U tissue using RT-PCR and quantitative mass spectrometry (MS) approaches. We will also use MS to characterize phosphorylated and cleaved TDP-43 isoforms. Finally, we will assess TDP-43 mediated aggregation and neurotoxicity by expressing full-length TDP-43 and the S6 isoform in cell lines and primary neuronal cultures. This study will provide the first detailed characterization of TDP-43 isoforms either isolated from FTLD-U brain tissue or expressed in culture. Frontotemporal dementia (FTD) and amyotrophic lateral sclerosis (ALS) are severe neurodegenerative disorders that together affect approximately 30,000 people in the United States. The study of TDP-43 will provide deeper understanding into the pathogenesis of these diseases and potentially lead to novel therapeutic strategies.

Agency
National Institute of Health (NIH)
Institute
National Institute on Aging (NIA)
Type
Postdoctoral Individual National Research Service Award (F32)
Project #
1F32AG032848-01A1
Application #
7678208
Study Section
Special Emphasis Panel (ZRG1-F03A-F (20))
Program Officer
Miller, Marilyn
Project Start
2009-04-01
Project End
2010-08-31
Budget Start
2009-04-01
Budget End
2010-03-31
Support Year
1
Fiscal Year
2009
Total Cost
$52,898
Indirect Cost
Name
Emory University
Department
Neurology
Type
Schools of Medicine
DUNS #
066469933
City
Atlanta
State
GA
Country
United States
Zip Code
30322
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Seyfried, Nicholas T; Gozal, Yair M; Donovan, Laura E et al. (2012) Quantitative analysis of the detergent-insoluble brain proteome in frontotemporal lobar degeneration using SILAC internal standards. J Proteome Res 11:2721-38
Dammer, Eric B; Fallini, Claudia; Gozal, Yair M et al. (2012) Coaggregation of RNA-binding proteins in a model of TDP-43 proteinopathy with selective RGG motif methylation and a role for RRM1 ubiquitination. PLoS One 7:e38658
Herskowitz, Jeremy H; Seyfried, Nicholas T; Gearing, Marla et al. (2011) Rho kinase II phosphorylation of the lipoprotein receptor LR11/SORLA alters amyloid-beta production. J Biol Chem 286:6117-27
Herskowitz, Jeremy H; Seyfried, Nicholas T; Duong, Duc M et al. (2010) Phosphoproteomic analysis reveals site-specific changes in GFAP and NDRG2 phosphorylation in frontotemporal lobar degeneration. J Proteome Res 9:6368-79