Knockdown of GPC3 inhibits the proliferation of Huh7 hepatocellular carcinoma cells through down-regulation of YAP.
Glypican-3 (GPC3), a membrane-associated heparan sulfate proteoglycan, is frequently upregulated in hepatocellular carcinoma (HCC). Yes-associated protein (YAP) is also found over-expressed in HCC and has been identified as a key effector molecule in Hippo pathway, which could control the organ size in animals through the regulation of cell proliferation and apoptosis and plays an important role in the development of malignant tumors. Studies have reported that GPC3 and YAP might collaborate to regulate the development of HCC. To elucidate the role of GPC3 in the development of HCC and its relationship with YAP, siRNA technique was employed to knock down GPC3 in Huh7 HCC cells. Moreover, recombinant human YAP-1 was used to examine the effects of GPC3 on Huh7 cells. The results of flow cytometric analysis and Annexin-V-FLUOS apoptosis assay showed that knockdown of GPC3-induced apoptosis in Huh7 cells, resulting in inhibition of cell proliferation as examined by EdU incorporation assay, migration, and invasion. GPC3 knockdown also suppressed the expression of YAP in mRNA and protein levels, as examined by fluorescence quantitative PCR and Western blot analysis. Moreover, addition of recombinant human YAP-1 effectively rescued the cells from apoptosis triggered by GPC3 knockdown. Taken together, our findings suggest that GPC3 regulates HCC cell proliferation with the involvement of Hippo pathway.
Department of Hepatobiliary Surgery, Affiliated Hospital of Guangdong Medical College, Zhanjiang 524001, China. miaohuilai@tom.com
Hui-Lai
HL
Miao
Zhi-Jian
ZJ
Pan
Chang-Jiang
CJ
Lei
Ji-Yu
JY
Wen
Ming-Yi
MY
Li
Zhong-Kao
ZK
Liu
Zhi-Dong
ZD
Qiu
Man-Zhou
MZ
Lin
Nian-Ping
NP
Chen
Ming
M
Chen
D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism*; D017304:Annexin A5 / Q000032:analysis; D017209:Apoptosis / Q000235:genetics; D006528:Carcinoma, Hepatocellular / Q000235:genetics / Q000378:metabolism*; D045744:Cell Line, Tumor; D002465:Cell Movement; D049109:Cell Proliferation; D015536:Down-Regulation; D005452:Fluoresceins; D015972:Gene Expression Regulation, Neoplastic; D053673:Glypicans / Q000235:genetics*; D006801:Humans; D008113:Liver Neoplasms / Q000235:genetics / Q000378:metabolism; D009361:Neoplasm Invasiveness / Q000235:genetics; D010750:Phosphoproteins / Q000378:metabolism*; D034622:RNA Interference; D012333:RNA, Messenger / Q000096:biosynthesis; D034741:RNA, Small Interfering; D011994:Recombinant Proteins / Q000378:metabolism; D014157:Transcription Factors; D000091102:YAP-Signaling Proteins
D048868:Adaptor Proteins, Signal Transducing; D017304:Annexin A5; D005452:Fluoresceins; C105937:GPC3 protein, human; D053673:Glypicans; D010750:Phosphoproteins; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D011994:Recombinant Proteins; D014157:Transcription Factors; D000091102:YAP-Signaling Proteins; C088374:YAP1 protein, human; C097499:fluorescein carboxylate