Role of GLI-1 in epidermal growth factor-induced invasiveness of ARCaP ^e prostate cancer cells

Epidermal growth factor (EGF) signaling and Hedgehog (HH) signaling are both involved in prostate cancer (PCa) progression, yet the mechanisms through which these two pathways are synergistically linked require elucidation. In the present study, we aimed to ascertain how EGF and the HH signaling transcription factor GLI-1 are linked in prostate cancer invasiveness. ARCaP human prostate cancer cells, which included ARCaP_E and ARCaP_M cells, were used as a model in the present study. The expression of EGF receptor (EGFR) and the HH signaling transcriptional factor GLI-1 were detected in ARCaP _E cells by immunofluorescence, and the ARCaP_E cells were treated with human recombinant EGF protein (hrEGF) for 4 consecutive days in vitro. Transwell invasion assays were performed in the ARCaP_E cells following treatment with DMSO (vehicle control), hrEGF, GATN61 (GLI-1-specific inhibitor), hrEGF plus GANT61 and in the ARCaP_M cells. The expression of phosphorylated extracellular signal regulated kinase (p-ERK), total ERK and GLI-1 was detected by western blotting in ARCaP_E cells at different time-points following treatment with hrEGF. The expression of EGFR and GLI-1 was detected in ARCaP_E cells, which exhibited a cobblestone-like morphology, while after treatment with hrEGF, the cell morphology was altered to a spindle-shaped mesenchymal cell morphology. Transwell invasion assays demonstrated that hrEGF dramatically enhanced the invasive capability of the ARCaP_E cells (P<0.05). Additionally, western blot assay demonstrated that the expression levels of p-ERK and GLI-1 in ARCaP_E cells increased in a time-dependent manner after treatment with hrEGF (P<0.05); however, the expression levels of total ERK in the cells remained relatively unchanged. It also demonstrated that the GLI-1 inhibitor GANT61 could reverse the enhanced invasive effect induced by EGF in ARCaP_E cells (P<0.05). Our preliminary in vitro study showed that EGF signaling may increase the invasive capability of ARCaP_E human prostate cancer cells via upregulation of p-ERK and the HH signaling transcriptional factor GLI-1. Additionally, this enhanced cell invasive effect was reversed by a GLI-1-specific inhibitor in vitro. Consequently, it indicates that both EGF and HH signaling are synergistically involved in the progression of human prostate cancer ARCaP cells, and GLI-1 may be one of the important effectors, which is activated by EGF downstream signaling, to promote the invasiveness of ARCaP _E prostate cancer cells.