RAN translation at CGG repeats induces ubiquitin proteasome system impairment in models of fragile X-associated tremor ataxia syndrome

Abstract

Fragile X-associated tremor ataxia syndrome (FXTAS) is a neurodegenerative disorder caused by a CGG trinucleotide repeat expansion in the 5' UTR of the Fragile X gene, FMR1. FXTAS is thought to arise primarily from an RNA gain-of-function toxicity mechanism. However, recent studies demonstrate that the repeat also elicits production of a toxic polyglycine protein, FMRpolyG, via repeat-associated non-AUG (RAN)-initiated translation. Pathologically, FXTAS is characterized by ubiquitin-positive intranuclear neuronal inclusions, raising the possibility that failure of protein quality control pathways could contribute to disease pathogenesis. To test this hypothesis, we used Drosophila- and cell-based models of CGG-repeat-associated toxicity. In Drosophila, ubiquitin proteasome system (UPS) impairment led to enhancement of CGG-repeat-induced degeneration, whereas overexpression of the chaperone protein HSP70 suppressed this toxicity. In transfected mammalian cells, CGG repeat expression triggered accumulation of a UPS reporter in a length-dependent fashion. To delineate the contributions from CGG repeats as RNA from RAN translation-associated toxicity, we enhanced or impaired the production of FMRpolyG in these models. Driving expression of FMRpolyG enhanced induction of UPS impairment in cell models, while prevention of RAN translation attenuated UPS impairment in cells and suppressed the genetic interaction with UPS manipulation in Drosophila. Taken together, these findings suggest that CGG repeats induce UPS impairment at least in part through activation of RAN translation.

CGG Repeat-Associated Non-AUG Translation Utilizes a Cap-Dependent Scanning Mechanism of Initiation to Produce Toxic Proteins

“…FMRpolyG accumulates in neuronal inclusions in FXTAS patient brains and disease models (Todd et al, 2013). CGG RAN translation is required for toxicity in Drosophila and impairs protein quality control pathways (Oh et al, 2015; Todd et al, 2013), with similar findings observed in other repeat expansion disorders (Mizielinska et al, 2014; Yamakawa et al, 2015; Zhang et al, 2015). …”

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

“…However, disruption of proteostasis may be a common mechanism of toxicity for many of these aberrant translation products. Consistent with this hypothesis, several RAN products from C9ORF72 (polyGly-Ala, -Gly-Pro, -Gly-Arg, and -Pro-Arg) and FMR1polyGly have been linked to proteasomal dysfunction (Gupta et al, 2017; Oh et al, 2015; Yamakawa et al, 2015; Zhang et al, 2014b). C9ORF72polyPro-Arg is also associated with impaired autophagy (Gupta et al, 2017).…”

RAN translation—What makes it run?

“…Initiation within the 5’ UTR occurs in at least two reading frames in the absence of an AUG start codon: the GGC (+1) frame yields a polyglycine product (FMRpolyG), and the GCG (+2) frame yields a polyalanine product (FMRpolyA). FMRpolyG accumulates in ubiquitinated inclusions in patient tissue and cellular and animal disease models, is necessary to elicit toxicity in Drosophila models of disease, and induces proteasome perturbations in Drosophila and HeLa cells (Todd et al , 2013; Buijsen et al , 2014; Oh et al , 2015). In an inducible mouse model of FXTAS that expresses the FMR1 5'UTR with 90 CGG repeats, turning off transgene expression reverses the formation of neuronal FMRpolyG-positive inclusions and repeat-elicited behavioral deficits (Hukema et al ., 2015).…”