Two mutations G335D and Q343R within the amyloidogenic core region of TDP-43 influence its aggregation and inclusion formation

Abstract

TDP-43 is a DNA/RNA binding protein associated with TDP-43 proteinopathies. Many mutations have been identified in the flexible C-terminal region, which is implicated in the disease pathology. We investigated four point mutations in the amyloidogenic core region (residues 311–360) of TDP-43 by biochemical and spectroscopic methods. We found that the G335D mutation enhances the aggregation and inclusion formation of TDP-43 and this mutant in TDP-35 (the C-terminal fragment of 35 kDa) exaggerates the antagonist effect on RNA processing by endogenous TDP-43; whereas Q343R gives an opposite effect. As a comparison, M337V and Q331K have very little impact on the aggregation and inclusion formation of TDP-43 or TDP-35. NMR structural analysis showed that the G335D mutant in the core region forms a loop linker between the two α-helices and promotes α-to-β transition, but Q343R loses the second helix and consequently the structural transformation. Thus, the propensity of structural transformation in the amyloidogenic core of TDP-43 determines its aggregation and inclusion formation. This study may provide a molecular mechanism of the TDP-43 proteinopathies caused by genetic mutations.

ALS Mutations Disrupt Phase Separation Mediated by α-Helical Structure in the TDP-43 Low-Complexity C-Terminal Domain

“…Some but not all ALS associated mutations examined here are reported to alter aggregation of TDP-43(Jiang et al, 2016). In our hands, between 30 minutes and ~4 hours after LLPS induction, turbidity changes (Figure S4A, right).…”

“…Unlike FUS and hnRNP A1, TDP-43 CTD contains a conserved region of with both helical propensity and ALS mutations. Although some TDP-43 variants increase aggregation (Johnson et al, 2009), two mutations (Q331K and M337V) can induce motor neuron defects in mice in the absence of inclusion formation(Arnold et al, 2013; D'Alton et al, 2014) and do not effect TDP-43 aggregation(Jiang et al, 2016). Therefore, it is possible that ALS mutations disrupt normal TDP-43 function by altering the ability of the protein to participate in functional complexes mediated by phase separation.…”

Atomic-level evidence for packing and positional amyloid polymorphism by segment from TDP-43 RRM2

“…Specifically, segment 286–331 has been shown to form amyloid fibrils and confer neurotoxicity on primary cortical neurons 41 . Residues 311–360 have been proposed to be the amyloid core through NMR and CD studies 42,43 . Finally, it has been shown that residues 341–367 can drive pathological aggregation of TDP-43 in neuronal cell lines 44 , residues 342–366 transition from a random coil to a β-sheet 45 , and that deletion of residues 318–343 delays aggregation of full-length TDP-43 (ref.…”

An acridine derivative, [4,5-bis{(N-carboxy methyl imidazolium)methyl}acridine] dibromide, shows anti-TDP-43 aggregation effect in ALS disease models

“…As the aggregation of TDP-43 fragments has been previously shown to be accompanied by change into amyloid-like β-sheet rich conformation1241, we therefore examined the secondary structural features by far-UV circular dichroism (CD). As expected, TDP-43 2C protein aggregates displayed a negative peak at ~218 nm in the far-UV CD spectrum thus indicating a β-sheet rich structure.…”