To investigate mutations in OTU domain - containing protein 3 (OTUD3) and their functional mechanisms in the initiation and progression of colorectal cancer (CRC).

Gene expression profiling interactive analysis (GEPIA2) and the human protein atlas database (THPA) were used to analyze gene transcription and protein expressions. Samples from 32 patients with CRC were collected to identify OTUD3 mutants. Based on the information about mutation sitesof OTUD3 in an existing database, a plasmid vector containing the OTUD3 gene mutant was constructed. Plasmid vectors containing the phosphatase and tensin homolog deleted on chromosome 10 (PTEN) and the OTUD3 gene mutant were co-transfected into HCT116 cells. Western blotting, half-life, immunoprecipitation, ubiquitination, and hybrid algorithm molecular docking (H-DOCK) assays were employed to find out whether and why the OTUD3 mutants affected PTEN protein levels. Functional alterations in CRC cells after OTUD3 mutation were verified by CCK-8 cell proliferation, transwell cell invasion, scratch, and clonal formation assays.

OTUD3 mutations were highly frequent in CRC. OTUD3 mutants R178W and N321S resulted in the loss of function of the stable PTEN protein, leading to enhanced proliferation, invasion, migration, and survival of CRC.

In CRC, OTUD3 mutation reduces the ability to stabilize PTEN and promotes the occurrence and development of CRC.