RASSF1A-mediated amphiregulin regulation in hepatocellular carcinoma via Hippo pathway
간세포암종에서 Hippo pathway를 통한 RASSF1A의 AREG 발현 조절
Dept. of Medical Science/석사
Ras association domain family 1 isoform A (RASSF1A) is a tumor suppressor and its methylation is involved in many human cancers including hepatocellular carcinoma (HCC). RASSF1A induces tumor suppressive effect by phosphorylation and activation of Hippo pathway kinase cascade.The activated kinase cascade leads to inhibition of yes-associated protein (YAP), a potent transcription coactivator playing a critical role as an oncogeniccomponent inducing tumorigenic effect in the Hippo pathway. Amphiregulin (AREG) is one of the target genes of YAPthat induces cell proliferation and anti-apoptoticeffect on cancer cells. Here, we demonstratethe regulation of AREG by RASSF1Aoverexpression through the Hippo pathway. Methylation of RASSF1A was examined in HepG2, Hep3B, Huh7, SK-Hep1, and PLC/PRF/5 HCC cells. Overexpression of RASSF1A reduced cell proliferation significantly in Hep3B and SK-Hep1 cells at 24h, 48h, and 72h and PLC/PRF/5 cells at 48h and 72h (p<0.05). RASSF1A overexpression also induced apoptosis in SK-Hep1 (p< 0.05) and PLC/PRF/5 cells. Phosphorylation of MST1/2 and YAP, which is a component of the Hippo pathway, was detected in Hep3B, SK-Hep1, and PLC/PRF/5 cells with overexpressed RASSF1A. As a result, we have also examined reduced AREG expressionin Hep3B, SK-Hep1, and PLC/PRF/5 cellswith overexpressed RASSF1A. Furthermore, in human liver tissues, RASSF1A expression was higher in non-tumor liver tissues than in HCC tissues,but AREG and YAP expressionswere higher in HCC tissues than in non-tumor liver tissues.Our results suggest that RASSF1A expression in HCC cells inhibit cell proliferation and induce apoptosis. Increased phosphorylation of MST and YAP in HCC cells with overexpressed RASSF1A, revealedthe activation of the Hippo pathway. We have also found the decrease of AREG levels in HCC cells with overexpressed RASSF1A, explaining the inhibition of YAP, which is a transcription coactivator of AREG. From the in vivo study results, we have showed that the detection of RASSF1A, YAP, and AREG in HCC tissues support the regulation of AREG by RASSF1A. Thus, our results support that RASSF1A-mediated downregulation of AREG activates the tumor suppressive Hippo pathway by phosphorylation of YAP.