Elimination of TDP-43 inclusions linked to amyotrophic lateral sclerosis by a
misfolding-specific intrabody with dual proteolytic signals
#MMPMID29662239
Tamaki Y
; Shodai A
; Morimura T
; Hikiami R
; Minamiyama S
; Ayaki T
; Tooyama I
; Furukawa Y
; Takahashi R
; Urushitani M
Sci Rep
2018[Apr]; 8
(1
): 6030
PMID29662239
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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.
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