Abstract 5024: USP8, a survival gatekeeper turned oncogenic trigger via hyperactivation of the integrated stress response

作者：Debjani Mandal, Santosh Kumar, Dhruval Bhatt, Yan Li, Kory Johnson, Maric Dragan, Prashant Chittiboina · 发表于：Cancer Research · 年份：2026 · DOI：10.1158/1538-7445.am2026-5024 · 研究领域：Ubiquitin and proteasome pathways、Protein Degradation and Inhibitors、interferon and immune responses

Abstract Eukaryotic deubiquitinase enzymes (DUBs) play a vital role in maintaining cellular homeostasis by deubiquitinating substrates marked for degradation. Of the ∼100 DUBs discovered in humans, USP8 is unique for its association with human tumors including Cushing’s disease (activating mutation), and solid cancers (genomic overexpression). The mechanisms underlying USP8 in tumorigenesis remain unknown. We first found that USP8 is essential for cell survival using CRISPR-Cas9-mediated homozygous excision and RNA interference. To circumvent obligate lethality, identify the substrates of USP8, and determine pathways dysregulated in human tumors with elevated USP8 activity, we engineered a GFP-auxin inducible degron (AID) at the endogenous USP8 locus in DLD1 cells. Additionally, we assayed both transient and long-term interactors using a promiscuous biotinylator BirA fused to a USP8 lentiviral construct. We performed cell survival, apoptosis assays, immunocytochemistry, western blot, RNAseq, and TMT labeled LC/MS, TUBE and IP-LC/MS study. We found that with auxin activation, DLD1USP8-GFP-AID had >80% reduction in USP8 within 30 minutes. At 6 hours, LC/MS revealed a profound alteration of the proteome and phosphoproteome with suppression of eukaryotic elongational initiation factor-2 (EIF2) signaling. We found that USP8 triggers the Integrated Stress Response (ISR), as measured by an ATF4 reporter assay. Consistently, USP8 overexpression led to a concordant elevation of...