Abstract
Tumour invasion and metastasis is the most common cause of death from cancer. For epithelial cells to invade surrounding tissues and metastasise, an epithelial-mesenchymal transition (EMT) is required. We have demonstrated that FGFR1 expression is increased in bladder cancer and that activation of FGFR1 induces an EMT in urothelial carcinoma (UC) cell lines. Here, we created an in vitro FGFR1-inducible model of EMT, and used this model to identify regulators of urothelial EMT. FGFR1 activation promoted EMT over a period of 72 hours. Initially a rapid increase in actin stress fibres occurred, followed by an increase in cell size, altered morphology and increased migration and invasion. By using site-directed mutagenesis and small molecule inhibitors we demonstrated that combined activation of the mitogen activated protein kinase (MAPK) and phospholipase C gamma (PLCγ) pathways regulated this EMT. Actin stress fibre formation was regulated by PLCγ activation, and was also important for the increase in cell size, migration and altered morphology. MAPK activation regulated migration and E-cadherin expression, indicating that combined activation of PLCγand MAPK is required for a full EMT. We used expression microarrays to assess changes in gene expression downstream of these signalling cascades. COX-2 was transcriptionally upregulated by FGFR1 and caused increased intracellular prostaglandin E2 levels, which promoted migration. In conclusion, we have demonstrated that FGFR1 activation in UC cells lines promotes EMT via coordinated activation of multiple signalling pathways and by promoting activation of prostaglandin synthesis.
Highlights
Epithelial to mesenchymal transition (EMT) is a process that was observed initially in embryonic development but more recently has been implicated as a mechanism for cancer metastasis [1,2]
Using site-directed mutagenesis and small molecule inhibitors we have identified signalling pathways activated by FGFR1 that contribute to EMT, including the mitogen activated protein kinase (MAPK), PLCc and COX-2 signalling pathways
FGFR1 was ectopically expressed in a polyclonal population of 94-10 cells by retroviral transduction and expression confirmed by western blotting (Figure S1D))
Summary
Epithelial to mesenchymal transition (EMT) is a process that was observed initially in embryonic development but more recently has been implicated as a mechanism for cancer metastasis [1,2]. Tumour invasion and metastasis is the major cause of death in cancer patients, the biological mechanisms of metastasis remain incompletely understood. The majority of adult solid tumours are derived from an epithelial lineage. Epithelial cells form layers of cells that are closely adjoined by specialised membrane structures and such cells are generally non-motile under normal conditions. For epithelial cancer cells to invade into surrounding tissues and establish secondary tumours at distant sites they must lose cell-cell adhesions and polarity and increase their motility. Understanding the complex mechanisms that drive these changes in EMT is key to developing therapeutic strategies to both prevent and treat metastasis
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