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| Status |
Public on Jun 01, 2014 |
| Title |
Loss of nuclear TDP-43 in ALS causes altered expression of splicing machinery and widespread dysregulation of RNA splicing in motor neurons [fibroblasts] |
| Organism |
Homo sapiens |
| Experiment type |
Expression profiling by array
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| Summary |
Aims: Loss of nuclear TDP-43 characterises sporadic and most familial forms of amyotrophic lateral sclerosis (ALS). TDP-43 (encoded by TARDBP) has multiple roles in RNA processing. We aimed to determine whether 1) RNA splicing dysregulation is present in lower motor neurons in ALS and in a motor neuron-like cell model, and 2) TARDBP mutations (mtTARDBP) are associated with aberrant RNA splicing using patient-derived fibroblasts.
Methods: Affymetrix exon arrays were used to study mRNA expression and splicing in lower motor neurons obtained by laser capture microdissection of autopsy tissue from individuals with sporadic ALS and TDP-43 proteinopathy. Findings were confirmed by qRT-PCR and in NSC34 motor neuronal cells following shRNA-mediated TDP-43 depletion. Exon arrays and immunohistochemistry were used to study mRNA splicing and TDP-43 expression in fibroblasts from patients with mtTARDBP-associated, sporadic and mutant SOD1-associated ALS.
Results: We found altered expression of spliceosome components in motor neurons and widespread aberrations of mRNA splicing that specifically affected genes involved in ribonucleotide binding. This was confirmed in TDP-43 depleted NSC34 cells. Fibroblasts with mtTARDBP showed loss of nuclear TDP-43 protein and demonstrated similar changes in splicing and gene expression, that were not present in fibroblasts from patients with sporadic or SOD1-related ALS.
Conclusion: Loss of nuclear TDP-43 is associated with RNA processing abnormalities in ALS motor neurons, patient-derived cells with mtTARDBP, and following artificial TDP-43 depletion, suggesting that splicing dysregulation directly contributes to disease pathogenesis. Key functional pathways affected include those central to RNA metabolism.
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| Overall design |
RNA was extracted from fibroblasts grown from neurologically healthy controls (n=6) and 3 groups of patients with ALS: 1) sporadic cases (n=6); 2) cases due to mutations of SOD1 (n=4); 3) cases due to mutations of TARDBP (n=3). The three ALS groups were compared to the controls.
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| Contributor(s) |
Highley JR, Hewamadduma CA, Jansweijer JA, Kirby J, Heath PR, Kalaitzis A, Higginbottom A, Raman R, Ferraiuolo L, Allen SP, McDermott CJ, Lawrence N, Cooper-Knock J, Milo M, Wilson SA, Ince PG, Shaw PJ |
| Citation(s) |
24750229 |
| |
| Submission date |
Nov 21, 2011 |
| Last update date |
Aug 31, 2014 |
| Contact name |
Paul Roy Heath |
| E-mail(s) |
p.heath@sheffield.ac.uk
|
| Phone |
+44 114 2222254
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| Organization name |
University of Sheffield
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| Department |
Academic Neurology Unit
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| Lab |
Heath
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| Street address |
SITraN, 385a Glossop Road
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| City |
Sheffield |
| State/province |
S. Yorks |
| ZIP/Postal code |
S10 2HQ |
| Country |
United Kingdom |
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| Platforms (1) |
| GPL5188 |
[HuEx-1_0-st] Affymetrix Human Exon 1.0 ST Array [probe set (exon) version] |
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| Samples (19)
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| This SubSeries is part of SuperSeries: |
| GSE56504 |
Loss of nuclear TDP-43 in ALS causes altered expression of splicing machinery and widespread dysregulation of RNA splicing in motor neurons |
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| Relations |
| BioProject |
PRJNA148163 |
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