Hippo pathway-independent nuclear export of the transcriptional activator YAP regulates proliferation and migration in HCC cells

Aims: The Hippo pathway is a central regulator of organ size in different organisms. Hippo signalling is regulated by direct cell-cell interaction and contact inhibition, leading to phosphorylation and cytoplasmic retention of the transcriptional co-activator YAP (yes-associated protein). Because most human cancer cells become refractory to contact inhibition, it was suggested that this pathway and especially the nuclear accumulation of YAP is involved in liver cancer development.

Methods: Expression of YAP in human normal liver, premalignant lesions (dysplastic nodules; DNs), and hepatocellular carcinoma (HCC) tissues was analyzed using tissue-microarrays. YAP levels were determined by real-time PCR and western blotting in HCCs. Functional assays (cell viability, migration, and invasion) were performed after inhibition of hippo signalling constituents by siRNA in different HCC cell lines.

Results: Expression analysis revealed a significant induction of YAP transcript and protein levels in ~66% of human HCC compared to normal livers. A strong expression for YAP was detectable in most DNs and HCCs while only a weak staining was observed in normal liver. Nuclear YAP expression significantly correlated with tumor dedifferentiation (r=0.7, p<0.001) and proliferation (r=0.55, p<0.001). YAP predominantly localized in the nucleus of HCC cells while no protein levels were detectable in primary human hepatocytes. Reduced YAP concentrations significantly decreased tumor cell viability, migration, and matrigel invasion. Surprisingly, knock-down of different hippo pathway kinases did not result in a nuclear accumulation of YAP.

Conclusion: Overexpression and nuclear enrichment of YAP is frequently observed in human hepatocarcinogenesis and facilitates tumor growth and HCC cell dissemination. Importantly, nuclear translocation of YAP is uncoupled from hippo signalling, suggesting independence of pro-tumorigenic YAP activity from contact inhibition and growth control.