A G4C2 hexanucleotide repeat expansion within the first intron of chromosome 9, open reading frame 72 (C9orf72) is the most common genetic cause of amyotrophic lateral sclerosis and frontotemporal dementia. Despite being located within a non-coding region, the expanded G4C2 repeat triggers a non-canonical form of translation initiation termed repeat-associated non-AUG translation (RANT). RANT in all three reading frames of sense C9orf72 transcripts produces three dipeptide repeat-containing proteins (DRPs) that form neuronal inclusions throughout the central nervous system of mutation carriers. Exogenous expression of these DRPs is neurotoxic. However, the mechanism by which the intronic repeat becomes RAN-translated is not well understood. The goal of this project is to identify the factors required for RANT from intronic G4C2 repeat expansions and determine the effect that altering these factors has on repeat-elicited neuronal toxicity. I will accomplish these goals by utilizing a series of G4C2 RANT-specific reporters that I have generated and validated in vitro in a rabbit reticulocyte lysate and in vivo in cultured cells. Using these reporters, I will quantitatively assess the role of the upstream sequence context, the dependency on an mRNA 5? m7G cap structure, and the utilization of near-cognate codons, in RANT at G4C2 repeats in all three reading frames. I will then use primary neuronal models to delineate how sequence changes that modulate RANT efficiency alter repeat-induced decreases in neuronal survival.
In the most common genetic form of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD), an unexpected translation initiation event, termed repeat-associated non-AUG translation (RANT), produces toxic proteins that accumulate within patient neurons and contribute to disease. This proposal explores how this atypical translation process works mechanistically. Understanding the mechanism by which RANT occurs in ALS/FTD will aid in the development of therapeutic strategies aimed at preventing both this process and the diseases with which it is associated.
| Green, Katelyn M; Glineburg, M Rebecca; Kearse, Michael G et al. (2017) RAN translation at C9orf72-associated repeat expansions is selectively enhanced by the integrated stress response. Nat Commun 8:2005 |
