EWS::FLI1 gene fusions underlie an aggressive subset of Ewing sarcoma (EWS). DNA replication stress is abundant in EWS::FLI1-driven EWS, and the mechanism by which EWS cells survive that stress is unknown. In their study, Mallard and colleagues used RT-qPCR arrays and shRNA-mediated EWS::FLI1 ablation to demonstrate that EWS::FLI1 facilitates RNA and protein expression of ubiquitin-specific peptidase 1 (USP1), a deubiquitylase. ChIP RT-qPCR and public ChIP-sequencing and CUT&RUN data analysis suggested EWS::FLI1 localizes to the USP1 promoter and directly promotes its transcription. Genetic and pharmacologic USP1 inhibition slowed S-phase entry and progression and induced cell death in EWS::FLI1-expressing EWS cells. Tandem mass tag mass spectrometry using USP1 shRNA-expressing EWS cells revealed that USP1 augments anti-apoptotic Survivin expression, which was confirmed by Western blotting after genetic or pharmacologic USP1 inhibition. Accordingly, USP1 inhibition enhanced activities of caspases 3, 7, and 9 in EWS::FLI1-expressing EWS cells, and sensitized the cells...

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