Multi-drug resistance is the major cause of chemotherapy failure in hepatocellular carcinoma (HCC). YAP, a critical effector of the Hippo pathway, has been shown to contribute to the progression, metastasis and invasion of cancers. However, the potential role of YAP in mediating drug resistance remains obscure. RT-qPCR and western blot were used to assess YAP expression in HCC cell lines. CCK-8 assays, flow cytometry, a xenograft tumour model, immunochemistry and GFP-mRFP-LC3 fusion proteins were utilized to evaluate the effect of YAP on multi-drug resistance, intracellular ROS production and the autophagy of HCC cells in vitro and in vivo. Autophagy inhibitor and rescue experiments were carried out to elucidate the mechanism by which YAP promotes chemoresistance in HCC cells. We found that BEL/FU, a typical HCC cell line with chemoresistance, exhibited overexpression of YAP. Moreover, the inhibition of YAP by shRNA or verteporfin conferred the sensitivity of BEL/FU cells to chemotherapeutic agents through autophagy-related cell death in vitro and in vivo. Mechanistically, YAP silencing significantly enhanced autophagic flux by increasing RAC1-driven ROS, which contributed to the inactivation of mTOR in HCC cells. In addition, the antagonist of autophagy reversed the enhanced effect of YAP silencing on cell death under treatment with chemotherapeutic agents. Our findings suggested that YAP upregulation endowed HCC cells with multi-drug resistance via the RAC1-ROS-mTOR pathway, resulting in the repression of autophagy-related cell death. The blockade of YAP may serve as a promising novel therapeutic strategy for overcoming chemoresistance in HCC.

Holliday junction recognition protein (HJURP) has been implicated in many cancers including hepatocellular carcinoma (HCC). However, the underlying mechanism by which HJURP promotes HCC cell proliferation remains unclear. RT-qPCR and immunohistochemistry were used to detect HJURP expression in HCC and adjacent tumor tissues and HCC cell lines. The localization of p21 were determined by immunofluorescence and western blot. Co-immunoprecipitation and western blot were used to validate the p21 stability and signaling pathways affected by HJURP. The effects of HJURP on HCC cell proliferation were assessed both in vivo and in vitro. The ERK1/2 pathway inhibitor U0126 and AKT pathway agonist SC-79 were used to treat HCC cell lines for further mechanistic investigations. HJURP expression was higher in HCC tissues than in para-tumor tissues. Moreover, ectopic HJURP expression facilitated the proliferation of HCC cells, whereas the depletion of HJURP resulted in decreased cell growth in vitro and in vivo. Furthermore, the effects of HJURP silencing were reversed by p21 knockdown. Likewise, p21 overexpression inhibited cell growth ability mediated by HJURP elevation. Mechanistically, HJURP destabilized p21 via the MAPK/ERK1/2 and AKT/GSK3β pathways, which regulated the nucleus-cytoplasm translocation and ubiquitin-mediated degradation of p21. Clinically, high HJURP expression was correlated with unfavorable prognoses in HCC individuals. Our data revealed that HJURP is an oncogene that drives cell cycle progression upstream of p21 in HCC. These findings may provide a potential therapeutic and prognostic target for HCC.

Plant homeodomain finger protein 8 (PHF8) serves an activator of epithelial-mesenchymal transition (EMT) and is implicated in various tumors. However, little is known about PHF8 roles in hepatocellular carcinoma (HCC) and regulating E-cadherin expression. PHF8 expression pattern was investigated by informatic analysis and verified by RT-qPCR and immunochemistry in HCC tissues and cell lines. CCK8, xenograft tumor model, transwell assay, and tandem mCherry-GFP-LC3 fusion protein assay were utilized to assess the effects of PHF8 on proliferation, metastasis and autophagy of HCC cells in vitro and in vivo. ChIP, immunoblot analysis, rescue experiments and inhibitor treatment were used to clarify the mechanism by which PHF8 facilitated EMT, metastasis and autophagy. PHF8 upregulation was quite prevalent in HCC tissues and closely correlated with worse overall survival and disease-relapse free survival. Furthermore, PHF8-knockdown dramatically suppressed cell growth, migration, invasion and autophagy, and the expression of SNAI1, VIM, N-cadherin and FIP200, and increased E-cadherin level, while PHF8-overexpression led to the opposite results. Additionally, FIP200 augmentation reversed the inhibited effects of PHF8-siliencing on tumor migration, invasion and autophagy. Mechanistically, PHF8 was involved in transcriptionally regulating the expression of SNAI1, VIM and FIP200, rather than N-cadherin and E-cadherin. Noticeably, E-cadherin degradation could be accelerated by PHF8-mediated FIP200-dependent autophagy, a crucial pathway complementary to transcriptional repression of E-cadherin by SNAI1 activation. These findings suggested that PHF8 played an oncogenic role in facilitating FIP200-dependent autophagic degradation of E-cadherin, EMT and metastasis in HCC. PHF8 might be a promising target for prevention, treatment and prognostic prediction of HCC.