Deubiquitinating enzyme Josephin-2 stabilizes PHGDH to promote a cancer stem cell phenotype in hepatocellular carcinoma.

Background: Deubiquitinating enzymes (DUBs) have been shown to be possible targets for hepatocellular carcinoma (HCC) treatment.

Objective: This study was designed to reveal the effect and underlying mechanism of Josephin-2, a relatively newly defined DUB, in HCC progression.

Methods: SNU-387 and PLC/PRF/5 cells were used for in vitro functional assays. The levels of Josephin-2 and phosphoglycerate dehydrogenase (PHGDH) were determined using RT-qPCR and western blotting. Cell proliferation, migration and invasion were assessed by CCK-8, colony formation and Transwell. Spheroid-forming assay was performed to assess the cancer stem cell (CSC)-phenotype of HCC cells. A xenograft mice model was applied to verify the effect of Josephin-2 on HCC cell growth in vivo.

Results: Herein, we showed that Josephin-2 expression was negatively correlated with HCC patient survival in data from the online database. Cell experiments indicated that knockdown of Josephin-2 attenuated HCC cell malignant biological behaviors. Besides, Josephin-2 silencing also decreased the spheroid-formation while inhibited the expression of CSC biomarkers (CD133, OCT4, SOX2 and EpCAM) in HCC cells. Mechanistically, Josephin-2 had a deubiquitinating activity towards the regulation of PHGDH protein, the rate-limiting enzyme in the first step of serine biosynthesis pathway. Depletion of Josephin-2 enhanced the ubiquitination degradation of PHGDH and ultimately inhibited the proliferation and CSC-phenotype of HCC in vitro and in vivo.

Conclusion: Our work uncovered the regulatory effects of Josephin-2 on PHGDH protein stability and profiled its contribution in HCC malignant progression, which might provide a potential therapeutic target for HCC.