Gain and loss of function of ALS-related mutations of TARDBP (TDP-43) cause motor deficits in vivo

TLDR

TDP‑43 aggregates are found in multiple neurodegenerative diseases, and TARDBP mutations have been identified in both sporadic and familial ALS. The study aimed to determine whether TARDBP mutations A315T, G348C, and A382T are pathogenic. The authors expressed these mutants in COS‑1 and Neuro2A cells, primary motor neuron cultures, and zebrafish embryos to assess localization, toxicity, and motor phenotypes. Mutant TDP‑43 localized to nuclei in cell lines but was neurotoxic and aggregated in motor neurons, and overexpression in zebrafish caused shortened axons, abnormal branching, and swimming deficits, while knockdown produced a similar phenotype rescued by wild‑type TDP‑43, indicating both toxic gain‑of‑function and loss‑of‑function mechanisms.

Abstract

TDP-43 has been found in inclusion bodies of multiple neurological disorders, including amyotrophic lateral sclerosis, frontotemporal dementia, Parkinson's disease and Alzheimer's disease. Mutations in the TDP-43 encoding gene, TARDBP, have been subsequently reported in sporadic and familial ALS patients. In order to investigate the pathogenic nature of these mutants, the effects of three consistently reported TARDBP mutations (A315T, G348C and A382T) were tested in cell lines, primary cultured motor neurons and living zebrafish embryos. Each of the three mutants and wild-type (WT) human TDP-43 localized to nuclei when expressed in COS1 and Neuro2A cells by transient transfection. However, when expressed in motor neurons from dissociated spinal cord cultures these mutant TARDBP alleles, but less so for WT TARDBP, were neurotoxic, concomitant with perinuclear localization and aggregation of TDP-43. Finally, overexpression of mutant, but less so of WT, human TARDBP caused a motor phenotype in zebrafish (Danio rerio) embryos consisting of shorter motor neuronal axons, premature and excessive branching as well as swimming deficits. Interestingly, knock-down of zebrafisfh tardbp led to a similar phenotype, which was rescued by co-expressing WT but not mutant human TARDBP. Together these approaches showed that TARDBP mutations cause motor neuron defects and toxicity, suggesting that both a toxic gain of function as well as a novel loss of function may be involved in the molecular mechanism by which mutant TDP-43 contributes to disease pathogenesis.