Journal
ACS CHEMICAL NEUROSCIENCE
Volume 13, Issue 17, Pages 2599-2612Publisher
AMER CHEMICAL SOC
DOI: 10.1021/acschemneuro.1c00873
Keywords
TDP-43; hyperphosphorylation; amyloid; oligomer; fibril; toxicity; neurodegenerative disease
Funding
- Career Development Award
- Academia Sinica [CDA-106-L01]
- Ministry of Science and Technology, Taiwan [MOST 108-2113-M-001027, MOST 109-2113-M-001-011]
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This study characterizes the effects of hyperphosphorylation on TDP-43 proteinopathies, finding that hyperphosphorylation promotes fibril formation and toxicity attributed to TDP-43 oligomers. These results contribute to a better understanding and potential therapeutic development for TDP-43-related diseases.
TDP-43 proteinopathies cover a range of neurodegenerative diseases, including frontotemporal lobar degeneration (FTLD) and amyotrophic lateral sclerosis (ALS). Hyperphosphorylated TDP-43 was found within the inclusion bodies in disease lesions; however, the role of hyperphosphorylation and the toxic species are still ambiguous. To characterize the hyperphosphorylation effect of TDP-43, here, we employed five serine mutations implicated in the diseases at serine locations 379, 403, 404, 409, and 410 in the C-terminus to aspartate (SSD) and to alanine (SSA). We systematically characterized the conformation, liquid-liquid phase separation, oligomerization, and fibrillization of TDP-43 variants. Results revealed that the recombinant TDP-43 variants readily formed structurally similar spherical oligomers, as evidenced by circular dichroism spectroscopy, fluorescence spectroscopy, the TDP-43 oligomer-specific antibody assay, dynamic light scattering, and transmission electron microscopy. After incubation, only the phosphor-mimic SSD TDP-43 formed thioflavin-positive amyloid fibrils, whereas wild-type and SSA TDP-43 formed amorphous aggregates. We also examined membrane disruption, the cytotoxicity of human neuroblastoma, and the synaptic loss of primary neurons induced by oligomers and large aggregates of TDP-43. The results showed that all oligomeric TDP-43 variants were toxic regardless of hyperphosphorylation, but the fibrils and amorphous aggregates were not. Overall, our results demonstrated the hyperphosphorylation effect on fibril formation and the toxicity attributed from TDP-43 oligomers. This study facilitates the understanding and therapeutic development for TDP-43 proteinopathies.
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