Ubiquitin-specific peptidase 39 promotes hepatocyte proliferation by targeting the Wnt/β-catenin signaling pathway

Ubiquitin-specific peptidase 39 (USP39) is a deubiquitinating enzyme implicated in maintaining cellular homeostasis, yet its role in physiological hepatocyte proliferation remains unclear. This study investigated whether USP39 regulated hepatocyte proliferation via the Wnt/β-catenin signaling pathway.

NCTC1469 and BNL CL.2 mouse hepatocyte cells were treated with hepatocyte growth factor (HGF) to establish an in vitro cell model mimicking liver regeneration. Cell viability and proliferation were evaluated using CCK-8, colony formation, and EdU assays; meanwhile, the mRNA and protein expression levels of USP39 were determined by quantitative real-time PCR (qRT-PCR) and Western blotting (WB), respectively. After USP39 knockdown/overexpression or treatment with Wnt inhibitor IWR-1, WB was performed to detect the level of key proteins in the Wnt/β-catenin pathway. To investigate the underlying mechanism, we measured β-catenin ubiquitination and half-life using Co-immunoprecipitation (IP), WB, and proteasome inhibitor MG132.

HGF treatment significantly upregulated USP39 expression. USP39 promoted NCTC1469 and BNL CL.2 cell proliferation by activating the Wnt/β-catenin pathway, but such effect was reversed by IWR-1. Additionally, USP39 inhibited β-catenin ubiquitination, prolonged its protein half-life, reduced its degradation. Importantly, this stabilizing effect of USP39 on β-catenin was mediated through the proteasome pathway.

This study confirmed the key regulatory role of USP39 in hepatocyte proliferation in vitro, revealing that USP39 stabilizes Wnt/β-catenin by regulating its ubiquitination and activating the Wnt/β-catenin pathway. These findings identify USP39 as a novel regulator of liver proliferation and link it to physiological liver regeneration, offering new insights for understanding liver proliferation mechanisms and developing therapeutic targets for liver diseases.