The most prevalent form of familial amyotrophic lateral sclerosis (ALS) involves the expansion of the GGGGCC (G4C2) hexanucleotide repeat found in the first intron of a gene designated C9orf72 unaffected patients bear 15-30 copies of the G4C2 repeat sequence. Affected patients exhibit expansions to upwards of 1,000 or more of the repeats. Various concepts have emerged concerning the molecular basis of disease pathophysiology, including impediments to expression of the C9orf72 gene itself, expression of putatively toxic sense and anti-sense transcripts of the repeats, expression of putatively toxic repeat-associated, and non-ATG (RAN) translation products. Among the five poly-dipeptides encoded by sense and anti-sense transcripts of the expanded repeat (GA, GP, GR, PA and PR), two have emerged to display significant toxicity – GR and PR. Recent studies have reported the ability of the GR and PR poly-dipeptides to impede nucleo: cytosolic transport, pre-mRNA splicing and rRNA biogenesis. Missing to date are comprehensive studies of the intracellular targets of GR and PR toxicity. Here, the research team has used a photocrosslinking approach in order to identify proteins bound by the PR poly-dipeptide. Observations outlined herein give evidence favoring interaction of PR with many RNA binding proteins containing low complexity sequences. They further report interaction of PR with the polypeptide constituents of intermediate filament proteins. Such studies indicate that PR toxicity results from widespread impediments to cell organization and function. Two complementary approaches were used in search of the intracellular targets of the toxic PR poly-dipeptide encoded by the repeat sequences expanded in the C9orf72 form of amyotrophic lateral sclerosis. The top categories of PR-bound proteins include constituents of non-membrane invested cellular organelles and intermediate filaments. PR targets are enriched for the inclusion of low complexity (LC) sequences.
Evidence is presented indicating that LC sequences represent the direct target of PR binding and that interaction between the PR poly-dipeptide and LC domains is polymer-dependent. These studies indicate that PR-mediated toxicity may result from broad impediments to the dynamics of cell structure and information flow from gene to message to protein.