A genome-scale RNAi screen was performed in a mammalian cell-based assay to identify modifiers of mutant huntingtin toxicity. Ontology analysis of suppressor data identified processes previously implicated in Huntington’s disease, including proteolysis, glutamate excitotoxicity, and mitochondrial dysfunction. In addition to established mechanisms, the screen identified multiple components of the RRAS signaling pathway as loss-of-function suppressors of mutant huntingtin toxicity in human and mouse cell models. Loss-of-function in orthologous RRAS pathway members also suppressed motor dysfunction in a Drosophila model of Huntington’s disease. Abnormal activation of RRAS and a down-stream effector, RAF1, was observed in cellular models and a mouse model of Huntington’s disease. We also observe co-localization of RRAS and mutant huntingtin in cells and in mouse striatum, suggesting that activation of R-Ras may occur through protein interaction. These data indicate that mutant huntingtin exerts a pathogenic effect on this pathway that can be corrected at multiple intervention points including RRAS, FNTA/B, PIN1, and PLK1. Consistent with these results, chemical inhibition of farnesyltransferase can also suppress mutant huntingtin toxicity. These data suggest that pharmacological inhibition of RRAS signaling may confer therapeutic benefit in Huntington’s disease. Author Summary Huntington’s disease (HD) is an inherited disorder caused by mutation of the gene that encodes the huntingtin protein. The specific mutation that results in disease is an increase in the copies of the amino acid glutamine in a stretch of repeated glutamines at the amino-terminus of the protein. This “expanded polyglutamine” huntingtin acquires toxic properties, presumably through mechanisms that involve its reduced solubility and aberrant interactions with other cellular proteins that do not occur with the normal protein. In this study, we sought to identify cellular processes that were involved in the toxicity conferred by the mutant huntingtin protein. We used RNA interference in order to specifically reduce the levels of individual cellular proteins and identified a number that could reduce mutant huntingtin toxicity. These modifiers clustered into functional pathways know to be involved in HD and other novel pathways. Among these modifiers, we found that the signaling protein RRAS, as well as additional members of its signaling cascade, are involved in mutant huntingtin toxicity. We further showed that a small molecule inhibitor of an enzyme involved in this pathway is effective at reducing this toxicity, indicating that the targeted inhibition of the RRAS pathway may be of therapeutic benefit in Huntington’s disease.
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