Poly-dipeptides encoded by the C9ORF72 repeats block global protein translation

Abstract

The expansion of the GGGGCC hexanucleotide repeat in the non-coding region of the Chromosome 9 open-reading frame 72 (C9orf72) gene is the most common genetic cause of frontotemporal dementia (FTD) and amyotrophic lateral sclerosis (ALS). This genetic alteration leads to the accumulation of five types of poly-dipeptides translated from the GGGGCC hexanucleotide repeat. Among these, poly-proline-arginine (poly-PR) and poly-glycine-arginine (poly-GR) peptides are known to be neurotoxic. However, the mechanisms of neurotoxicity associated with these poly-dipeptides are not clear. A proteomics approach identified a number of interacting proteins with poly-PR peptide, including mRNA-binding proteins, ribosomal proteins, translation initiation factors and translation elongation factors. Immunostaining of brain sections from patients with C9orf72 ALS showed that poly-GR was colocalized with a mRNA-binding protein, hnRNPA1. In vitro translation assays showed that poly-PR and poly-GR peptides made insoluble complexes with mRNA, restrained the access of translation factors to mRNA, and blocked protein translation. Our results demonstrate that impaired protein translation mediated by poly-PR and poly-GR peptides plays a role in neurotoxicity and reveal that the pathways altered by the poly-dipeptides-mRNA complexes are potential therapeutic targets for treatment of C9orf72 FTD/ALS.

C9orf72 Dipeptide Repeats Impair the Assembly, Dynamics, and Function of Membrane-Less Organelles

“…Despite efforts to define the basis of repeat expansion toxicity in C9-ALS/FTD, the field has struggled to integrate seemingly disparate findings, such as evidence of impaired nucleolar function (Kwon et al, 2014; Wen et al, 2014), disturbances in nucleocytoplasmic transport (Freibaum et al, 2015; Jovičić et al, 2015; Zhang et al, 2015), altered RNA splicing (Prudencio et al, 2015), impaired trafficking of RNA granules (Alami et al, 2014; Schweizer Burguete et al, 2015) and impaired translation (Kanekura et al, 2016). Furthermore, despite extensive clinicopathological overlap in ALS/FTD caused by mutation in C9ORF72 and RNA-binding proteins, a cohesive view of pathogenesis has been lacking.…”

Regulation of mRNA Translation in Neurons—A Matter of Life and Death

“…Phase separation may contribute to the defects in nucleocytoplasmic transport and nucleolar function associated with C9ORF72polyGly-Arg and -Pro-Arg (Freibaum et al, 2015; Jovičić et al, 2015; Kwon et al, 2014; Tao et al, 2015). Interestingly, these proteins also interact nonspecifically with mRNAs, blocking their interactions with translation factors and thus diminishing global translation (Kanekura et al, 2016). Less is known regarding the interactomes of other repeat-encoded proteins.…”

Evidence that C9ORF72 Dipeptide Repeat Proteins Associate with U2 snRNP to Cause Mis-splicing in ALS/FTD Patients

“…Understanding the mechanisms by which the repeat RNAs and/or the DPRs cause neurodegeneration is a focus of intense investigation. Among the possibilities being examined are that the repeat RNAs or DPR proteins interfere with essential cellular processes (Taylor et al, 2016), such as nucleocytoplasmic trafficking (Freibaum et al, 2015; Jovicic et al, 2015; Zhang et al, 2015), protein synthesis (Kanekura et al, 2016), mitochondria function (Lopez-Gonzalez et al, 2016) and pre-mRNA splicing (Conlon et al, 2016; Cooper-Knock et al, 2014; Kwon et al, 2014; Prudencio et al, 2015). In addition, pathogenesis may involve DPR protein-mediated disruption of the dynamics and assembly of membrane-less organelles such as nucleoli, stress granules, and nuclear speckle domains (Lee et al, 2016).…”