We examined the impact of crosstalk between the insulin receptor (INSR) and G protein-coupled receptor (GPCR) signaling pathways on the regulation of Yes-associated Protein (YAP) localization, phosphorylation and transcriptional activity in the context of human pancreatic ductal adenocarcinoma (PDAC). Stimulation of PANC-1 or MiaPaCa-2 cells with insulin and neurotensin, a potent mitogenic combination of agonists for these cells, promoted striking YAP nuclear localization and decreased YAP phosphorylation at Ser127 and Ser397. Challenging PDAC cells with either insulin or neurotensin alone modestly induced the expression of YAP/TEAD-regulated genes, including Connective Tissue Growth Factor (CTGF), Cysteine-rich angiogenic inducer 61 (CYR61) and CXCL5 whereas the combination of neurotensin and insulin induced a marked increase in the level of expression of these genes. In addition, siRNA-mediated knockdown of YAP/TAZ prevented the increase in the expression of these genes. A small-molecule inhibitor (A66), selective for the p110alpha subunit of PI3K, abrogated the increase in phosphatidylinositol 3,4,5-trisphosphate (PIP3) production and the expression of CTGF, CYR61 and CXCL5 induced by neurotensin and insulin. Furthermore, treatment of PDAC cells with protein kinase D (PKD) family inhibitors (CRT0066101 or kb NB 142-70) or with siRNAs targeting the PKD family abrogated the increase of CTGF, CYR61 and CXCL5 mRNA levels in response to insulin and neurotensin stimulation. Thus, PI3K and PKD mediate YAP activation in response to insulin and neurotensin in pancreatic cancer cells. Implications: Inhibitors of PI3K or PKD disrupt crosstalk between insulin receptor and GPCR signaling systems by blocking YAP/TEAD-regulated gene expression in pancreatic cancer cells.
- Received January 13, 2017.
- Revision received March 9, 2017.
- Accepted March 27, 2017.
- Copyright ©2017, American Association for Cancer Research.