The nuclear ubiquitin ligase adaptor SPOP is a conserved regulator of C9orf72 dipeptide toxicity

Significance The G4C2 repeat expansion in the C9orf72 gene is a major cause of frontotemporal dementia and amyotrophic lateral sclerosis. Unusual translation of the repeat sequence produces two highly toxic dipeptide repeat proteins (DPRs), PRX and GRX, which accumulate in the brain tissue of individuals with these diseases. Here, we show that PR and GR toxicity in both Caenorhabditis elegans and mammalian neurons depends on the E3 ubiquitin ligase adaptor SPOP. SPOP acts through bromodomain proteins to mediate dipeptide toxicity. SPOP inhibitors, which are currently being developed to treat SPOP-dependent renal cancer, also protect neurons against DPR toxicity. Our findings identify a highly conserved and “druggable” pathway that may represent a strategy for treating these currently incurable diseases. A hexanucleotide repeat expansion in the C9orf72 gene is the most common cause of inherited amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Unconventional translation of the C9orf72 repeat produces dipeptide repeat proteins (DPRs). Previously, we showed that the DPRs PR50 and GR50 are highly toxic when expressed in Caenorhabditis elegans, and this toxicity depends on nuclear localization of the DPR. In an unbiased genome-wide RNA interference (RNAi) screen for suppressors of PR50 toxicity, we identified 12 genes that consistently suppressed either the developmental arrest and/or paralysis phenotype evoked by PR50 expression. All of these genes have vertebrate homologs, and 7 of 12 contain predicted nuclear localization signals. One of these genes was spop-1, the C. elegans homolog of SPOP, a nuclear localized E3 ubiquitin ligase adaptor only found in metazoans. SPOP is also required for GR50 toxicity and functions in a genetic pathway that includes cul-3, which is the canonical E3 ligase partner for SPOP. Genetic or pharmacological inhibition of SPOP in mammalian primary spinal cord motor neurons suppressed DPR toxicity without affecting DPR expression levels. Finally, we find that knockdown of bromodomain proteins in both C. elegans and mammalian neurons, which are known SPOP ubiquitination targets, suppresses the protective effect of SPOP inhibition. Together, these data suggest a model in which SPOP promotes the DPR-dependent ubiquitination and degradation of BRD proteins. We speculate the pharmacological manipulation of this pathway, which is currently underway for multiple cancer subtypes, could also represent an entry point for therapeutic intervention to treat C9orf72 FTD/ALS.