FHOD1 is upregulated in clinical oral SCC, as well as <i>in vitro</i> in SCC cells with EMT features.

<p>A) In normal or non-neoplastic stratified squamous epithelium, no FHOD1 can be detected (top). In invasive squamous cell carcinomas, moderate to strong FHOD1 immunoreactivity is seen in spindle-shaped cells at the invasive front that on morphological grounds have undergone EMT (bottom; details in inset). In the tumour bulk, which consists of cells with an epithelial morphology, only weak immunoreactivity is present. Scale bar: 200 μm. B) Western blot analysis of cell lines show that the epithelial SCC cell line UT-SCC-43A does not express detectable FHOD1, while both the spontaneous EMT cell line UT-SCC-43B and the Snail-induced EMT cell line 43A-SNA express FHOD1. UT-SCC-43A expresses the epithelial marker E-cadherin but not N-cadherin, whereas UT-SCC43-B and 43A-SNA express N-cadherin but not E-cadherin. C) F-actin organization of UT-SCC-43A (upper panel) is typically epithelial, with distinct cell submembraneous filaments and scant stress fibres. UT-SCC-43B shows features of mesenchymal organization (middle panel). Cell-cell contacts are few, the cells are elongated and contain lamellopodia, filopodia and stress fibres. The insert (bottom panel) shows that a fraction of FHOD1 co-localizes with stress fibres in UT-SCC-43B cells (arrowheads). Nuclei are stained with DAPI (blue). D) FHOD1 upregulation in UT-SCC-43B cells is dependent of PI3K signalling. Treatment with MEK 1/2 inhibitor U0126 reduces phosphorylation of ERK 1/2 but does not influence FHOD1 expression. In contrast, PI3K inhibition by LY294992 markedly reduces FHOD1 expression. The reduction of p-Akt indicates that the pathway is efficiently inhibited.