Templated Aggregation of TAR DNA-binding Protein of 43 kDa (TDP-43) by Seeding with TDP-43 Peptide Fibrils

Shotaro Shimonaka,Takashi Nonaka,Genjiro Suzuki,Shin-Ichi Hisanaga,Masato Hasegawa

doi:10.1074/jbc.m115.713552

Abstract

TAR DNA-binding protein of 43 kDa (TDP-43) has been identified as the major component of ubiquitin-positive neuronal and glial inclusions in frontotemporal lobar degeneration and amyotrophic lateral sclerosis. Aggregation of TDP-43 to amyloid-like fibrils and spreading of the aggregates are suggested to account for the pathogenesis and progression of these diseases. To investigate the molecular mechanisms of TDP-43 aggregation, we attempted to identify the amino acid sequence required for the aggregation. By expressing a series of deletion mutants lacking 20 amino acid residues in the C-terminal region in SH-SY5Y cells, we established that residues 274-313 in the glycine-rich region are essential for aggregation. In vitro aggregation experiments using synthetic peptides of 40 amino acids from this sequence and adjacent regions showed that peptides 274-313 and 314-353 formed amyloid-like fibrils. Transduction of these fibrils induced seed-dependent aggregation of TDP-43 in cells expressing wild-type TDP-43 or TDP-43 lacking nuclear localization signal. These cells showed different phosphorylated C-terminal fragments of TDP-43 and different trypsin-resistant bands. These results suggest that residues 274-353 are responsible for the conversion of TDP-43 to amyloid-like fibrils and that templated aggregation of TDP-43 by seeding with different peptides induces various types of TDP-43 pathologies, i.e. the peptides appear to act like prion strains.

Highlights

Frontotemporal lobar degeneration (FTLD)2 is the second most common dementing disorder in patients under 65 years of age and is characterized by progressive atrophy of the frontal and temporal lobes in the brain
These findings strongly suggest that aggregation of C-terminal regions of TAR DNA-binding protein of 43 kDa (TDP-43) plays a central role in the pathogenesis and progression of Amyotrophic lateral sclerosis (ALS) and FTLD with TDP-43 pathologies (FTLD-TDP)
The Sar-soluble TDP-43 C-terminal fragments (CTFs) levels were similar among the deletions as detected with anti-GFP (Fig. 2, left panel), but remarkable decreases of Sar-insoluble TDP-43 were detected with anti-GFP or pS409/410 in cells expressing Del 4, 5, or 8 (Fig. 2, middle and right panels)

Summary

Introduction

Frontotemporal lobar degeneration (FTLD)2 is the second most common dementing disorder in patients under 65 years of age and is characterized by progressive atrophy of the frontal and temporal lobes in the brain. It has been shown that the Sar-insoluble TDP-43 fibrils prepared from the brains of patients with ALS/FTLD have prion-like properties and can seed-dependently convert normal TDP-43 to abnormal forms [16].

Results

Conclusion