Loss of nuclear TDP-43 in amyotrophic lateral sclerosis (ALS) causes altered expression of splicing machinery and widespread dysregulation of RNA splicing in motor neurones.

Abstract

AIMS:

Loss of nuclear TDP-43 characterizes 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 neurones in ALS and in a motor neurone-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 neurones obtained by laser capture microdissection of autopsy tissue from individuals with sporadic ALS and TDP-43 proteinopathy. Findings were confirmed by quantitative reverse transcription-polymerase chain reaction (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 neurones 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, which 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 neurones, 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.

• Authors: J Robin Highley, Janine Kirby, Joeri A Jansweijer, Philip S Webb, Channa A Hewamadduma, Paul R Heath, Adrian Higginbottom, Rohini Raman, Laura Ferraiuolo, Johnathan Cooper-Knock, Christopher J McDermott, Stephen B Wharton, Pamela J Shaw, Paul G Ince
• Journal: Neuropathology and applied neurobiology (Neuropathol Appl Neurobiol) Vol. 40 Issue 6 Pg. 670-85 (Oct 2014) ISSN: 1365-2990 [Electronic] England
• PMID: 24750229 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)
• Copyright: © 2014 British Neuropathological Society.

Chemical References

• DNA-Binding Proteins
• Nuclear Proteins

Topics

• Aged
• Amyotrophic Lateral Sclerosis (genetics)
• Animals
• DNA-Binding Proteins (genetics)
• Female
• Gene Expression
• Humans
• Male
• Mice
• Middle Aged
• Motor Neurons (metabolism)
• Nuclear Proteins (genetics)
• RNA Splicing
• Spinal Cord (metabolism)