Radio-resistance is a principal culprit in radiation therapy for hepatocellular carcinoma (HCC). Insights on the regulation genes of radio-resistance and underlying mechanisms in HCC are awaiting profound investigation. This study is designed to explore the role and mechanism of WD repeat and HMG-box DNA binding protein 1 (WDHD1) in HCC progression. WDHD1 mRNA level was detected using real-time quantitative polymerase chain reaction (RT-qPCR). WDHD1, ubiquitin-specific protease 28 (USP28), E-cadherin, N-cadherin, and vimentin protein levels were determined by Western blot. Cell viability, cell cycle progression, migration, invasion, and apoptosis were assessed using the cell counting kit-8 (CCK-8) assay, flow cytometry, wound healing assay, and Transwell assay. The radio-sensitivity of HCC cells was analyzed using a colony formation assay. After UbiBrowser database analysis, the interaction between USP28 and WDHD1 was verified using GST pull-down and Co-immunoprecipitation (CoIP) assay. Xenograft assay was used to test the effect of USP28 on radio-sensitivity in vivo. WDHD1 and USP28 were highly expressed in HCC patients and cell lines. Moreover, WDHD1 knockdown could repress HCC cell proliferation, migration, invasion, epithelial to mesenchymal transition (EMT), and enhance the radiosensitivity. Mechanistically, USP28 mediated the deubiquitination and stabilization of WDHD1 through its direct interaction. USP28 silencing increased the radiosensitivity of HCC in vivo. USP28 contributed to HCC development and radio-resistance through deubiquitinating WDHD1, providing a promising therapeutic target for HCC treatment.