TDP-43 is the major pathological protein identified in the cellular inclusions in amyotrophic lateral sclerosis and frontotemporal lobar degeneration. The pathogenic forms of TDP-43 are processed C-terminal fragments containing a truncated RNA-recognition motif (RRM2) and a glycine-rich region. Although extensive studies have focused on this protein, it remains unclear how the dimeric full-length TDP-43 is folded and assembled and how the processed C-terminal fragments are misfolded and aggregated. Here, using size-exclusion chromatography, pulldown assays, and small angle x-ray scattering, we show that the C-terminal-deleted TDP-43 without the glycinerich tail is sufficient to form a head-to-head homodimer primarily via its N-terminal domain. The truncated RRM2, as well as twoβ-strands within the RRM2, form fibrils in vitro with a similar amyloid-negative staining property to those of TDP-43 pathogenic fibrils in diseases. In addition to the glycine-rich region, the truncated RRM2, but not the intact RRM2, plays a key role in forming cytoplasmic inclusions in neuronal cells. Our data thus suggest that the process that disrupts the dimeric structure, such as the proteolytic cleavage of TDP-43 within the RRM2 that removes the N-terminal dimerization domain, may produce unassembled truncated RRM2 fragments with abnormally exposedβ-strands, which can oligomerize into high-order inclusions.
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