INSULIN-LIKE GROWTH FACTOR RECEPTOR-1 IS A MEDIATOR OF YAP-DEPENDENT ONCOGENIC FUNCTIONS

摘要

Objectives: Investigate the downstream target gene of Yes-Associated Protein (YAP) in hepatocellular carcinoma (HCC) and their functional and molecular link. Methods: Insulin-like growth factor receptor-1 (IGF1R), a potential downstream target gene of YAP, was screened out from the transcriptional profile of YAP over-expressing hepatic cell MIHA-YAP by using comparative cDNA microarray. The regulatory role of YAP with IGF1R was then validated by 1) qPCR and Western blotting examinations of IGF1R mRNA and protein levels in genetic modified MIHA, Huh7 and PLC/PRF/5 cells, 2) Chromatin immunoprecipitation (ChIP) of IGF1R promoter region in YAP-chromatin fragment complex. As the functions of IGF1R depend on its autophosphorylation, picropodophyllin was used to inhibit IGF1R activation in YAP overexpressing Huh7 cells. The cell viability and mobility were examined by using MTT assay and wound healing assay to clarify the functional connections between YAP and IGF1R. Finally, the regulation of YAP with IGF1R was demonstrated in 86 cases of HCC clinical specimens by using cDNA microarray. Results: qPCR and Western blotting showed the mRNA and protein levels of IGF1R were changed in commitment with YAP expression in MHA, Huh7 and PLC/PRF/5 cells. ChIP assay showed partial IGF1R promoter region can be amplified from YAP-chromatin fragment complex. When inhibiting IGF1R activation, YAP overexpressing Huh7 cells lost their advantages in cell proliferation and mobility. Furthermore, a significantly positive correlation of YAP and IGF1R mRNA expression was found in human HCC samples. Conclusions: IGF1R is a downstream target of YAP, mediating YAP-dependent oncogenic functions in HCC.