Elimination of TDP-43 inclusions linked to amyotrophic lateral sclerosis by a misfolding-specific intrabody with dual proteolytic signals

Yoshitaka Tamaki, Akemi Shodai, Toshifumi Morimura, Ryota Hikiami, Sumio Minamiyama, Takashi Ayaki, Ikuo Tooyama, Yoshiaki Furukawa, Ryosuke Takahashi, Makoto Urushitani

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9 Citations (Scopus)

Abstract

Aggregation of TAR DNA-binding protein of 43 kDa (TDP-43) is implicated in the pathogenesis of sporadic and certain familial forms of amyotrophic lateral sclerosis (ALS), suggesting elimination of TDP-43 aggregates as a possible therapeutic strategy. Here we generated and investigated a single-chain variable fragment (scFv) derived from the 3B12A monoclonal antibody (MAb) that recognises D247 of the TDP-43 nuclear export signal, an epitope masked in the physiological state. In transfected HEK293A cells, 3B12A scFv recapitulated the affinity of the full-length MAb to mislocalised TDP-43 with a defective nuclear localising signal and to a TDP-43 inclusion mimic with cysteine-to-serine substitution at RRM1. Moreover, 3B12A scFv accelerated proteasome-mediated degradation of aggregated TDP-43, likely due to an endogenous PEST-like proteolytic signal sequence in the VH domain CDR2 region. Addition of the chaperone-mediated autophagy (CMA)-related signal to 3B12A scFv induced HSP70 transcription, further enhancing TDP-43 aggregate clearance and cell viability. The 3B12A scFv also reduced TDP-43 aggregates in embryonic mouse brain following in utero electroporation while causing no overt postnatal brain pathology or developmental anomalies. These results suggest that a misfolding-specific intrabody prone to synergistic proteolysis by proteasomal and autophagic pathways is a promising strategy for mitigation of TDP-43 proteinopathy in ALS.

Original languageEnglish
Article number6030
JournalScientific Reports
Volume8
Issue number1
DOIs
Publication statusPublished - 2018 Dec 1

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Amyotrophic Lateral Sclerosis
TDP-43 Proteinopathies
Monoclonal Antibodies
Nuclear Export Signals
Single-Chain Antibodies
Electroporation
Autophagy
DNA-Binding Proteins
Brain
Proteasome Endopeptidase Complex
Protein Sorting Signals
Serine
Proteolysis
Cysteine
Epitopes
Cell Survival
Pathology
Therapeutics
Amyotrophic lateral sclerosis 1

ASJC Scopus subject areas

  • General

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Elimination of TDP-43 inclusions linked to amyotrophic lateral sclerosis by a misfolding-specific intrabody with dual proteolytic signals. / Tamaki, Yoshitaka; Shodai, Akemi; Morimura, Toshifumi; Hikiami, Ryota; Minamiyama, Sumio; Ayaki, Takashi; Tooyama, Ikuo; Furukawa, Yoshiaki; Takahashi, Ryosuke; Urushitani, Makoto.

In: Scientific Reports, Vol. 8, No. 1, 6030, 01.12.2018.

Research output: Contribution to journalArticle

Tamaki, Y, Shodai, A, Morimura, T, Hikiami, R, Minamiyama, S, Ayaki, T, Tooyama, I, Furukawa, Y, Takahashi, R & Urushitani, M 2018, 'Elimination of TDP-43 inclusions linked to amyotrophic lateral sclerosis by a misfolding-specific intrabody with dual proteolytic signals', Scientific Reports, vol. 8, no. 1, 6030. https://doi.org/10.1038/s41598-018-24463-3
Tamaki, Yoshitaka ; Shodai, Akemi ; Morimura, Toshifumi ; Hikiami, Ryota ; Minamiyama, Sumio ; Ayaki, Takashi ; Tooyama, Ikuo ; Furukawa, Yoshiaki ; Takahashi, Ryosuke ; Urushitani, Makoto. / Elimination of TDP-43 inclusions linked to amyotrophic lateral sclerosis by a misfolding-specific intrabody with dual proteolytic signals. In: Scientific Reports. 2018 ; Vol. 8, No. 1.
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