Fig. 4: YAP may mediate the aggressive phenotype of ESR1-L HCC. | Experimental & Molecular Medicine

Fig. 4: YAP may mediate the aggressive phenotype of ESR1-L HCC.

From: YAP inactivation in estrogen receptor alpha-positive hepatocellular carcinoma with less aggressive behavior

Fig. 4

a A genetic network of Sig ESR1-L genes was constructed showing physical interactions (pink), genetic interactions (green), and pathways (blue) by using GeneMANIA software in Cytoscape (version 3.4.1). Cell cycle-related genes and YAP target genes in Sig ESR1-L are indicated. b Plot showing the gradually increase in ESR1 expression (top). Heatmap showing the expression of YAP target genes and cell cycle-related genes in Sig ESR1-L (bottom). Cell cycle groups, such as G0/early G1-, G1/S-, G2/M-, and mitosis-related groups, are indicated. c GSEA results showing the enrichment of YAP activation-related signatures (e.g., YAP1_UP and YAP_CONSERVED_SIG). d Bar plots showing the frequency of transcription factors binding to the promoters of the top-ranked genes in Sig ESR1-L (n = 50). This database was investigated using GeneHancer (https://www.genecards.org/). e The amount of YAP protein that translocated from the cytosol into the nucleus was assessed by Western blotting after nucleocytoplasmic fractionation. GAPDH and histone H3 were used as nuclear and cytoplasmic control markers, respectively (left). The amount of YAP protein that was translocated from the cytosol into the nucleus was assessed by Western blotting after nucleocytoplasmic fractionation using ImageJ (1.8.0_172) (right). f Confocal immunofluorescence of YAP and 4′,6-diamidino-2-phenylindole (DAPI). DAPI (blue) and YAP (green) were detected by confocal microscopy as described in the Materials and Methods. g Dual-luciferase assays showed that the enhanced normalized relative luminescence unit (RLU) value of the 8xGTIIC construct was decreased in ERα-expressing HCC cell lines. The pGL3.0-basic vector was used as a control (Ctrl). Statistical significance is indicated (Ctrl vs. ERα; *p < 0.05, **p < 0.01, and ***p < 0.001, Student’s t-test). h Bar plots showing the expression of YAP target genes (e.g., MYBL2, CDC20, and TOP2A) in ERα (+) and ERα (−) liver cancer cell lines (HepG2 and Hep3B). The expression levels of each gene were normalized to the expression level of ACTB. The data were presented as the mean ± SD values (***p < 0.001, Student’s t-test).

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