Functional analysis of Zyxin in cell migration and invasive potential of oral squamous cell carcinoma cells

Abstract

Zyxin is an evolutionarily conserved protein that has been implicated in the regulation of actin assembly and is mainly located at focal adhesions. However, the biological roles of Zyxin in cancer cells are incompletely understood. We analyzed the functions of Zyxin in cell migration and the invasive potential of OSCC. Zyxin expression was examined using eight OSCC cell lines with two different cell morphologies (6 epithelial type and 2 fibroblastic type). To knockdown Zyxin expression, OSCC cells were transfected with Zyxin siRNA and control siRNA. The cell lines were studied by western blot analysis, immunocytochemical analysis and cell migration and invasion assay. Epithelial type OSCC cells showed a high level of E-cadherin expression and a low level of Zyxin expression. N-cadherin as well as Zyxin were strongly expressed in fibroblastic type OSCC cells. Expression levels of LPP and TRIP6, members of the human Zyxin family, did not differ between epithelial type and fibroblastic type. Knockdown of Zyxin expression by siRNA in fibroblastic type OSCC cells was associated with cell morphological changes from spindle (fibroblastic) to polygonal (epithelial) shape and significantly inhibited cell growth as well as cell migration and invasion. Expression levels of Rac1 and Cdc42 were weaker in Zyxin siRNA-treated fibroblastic type OSCC cells than in control siRNA-treated cells, but the expression of RhoA did not differ significantly. Treatment of fibroblastic type OSCC cells with Rac1 inhibitor decreased the expression of Zyxin mRNA and protein. Zyxin is suggested to promote growth, migration and invasiveness of fibroblastic type OSCC cells by upregulating Rac1 and Cdc42.

Molecular analysis of keratocystic odontogenic tumor cell lines derived from sporadic and basal cell nevus syndrome patients

“…For western blotting, iKCOT1 and sKCOT1 were harvested or incubated with low (0.65 mM) or high (1.23 mM) calcium medium for 10-12 days. Gel electrophoresis and western blotting were performed as described previously (18). Signals were detected by chemiluminescence using a Pierce SuperSignal western blotting kit (Thermo Fisher Scientific).…”

Transcriptional Network Analysis Reveals that AT1 and AT2 Angiotensin II Receptors Are Both Involved in the Regulation of Genes Essential for Glioma Progression

“…Another gene encoding the protein Zyxin (Zyx) was also regulated at both time intervals. Zyx concentrates at focal adhesions, regulates actin assembly and was previously described to be relevant for cell migration and invasion [28]. In addition, the Nek2 gene, which codes for a centrosomal kinase, was also observed in the common genes list obtained from Ang II x Control comparisons.…”

The Hippo pathway transcriptional co-activator, YAP, confers resistance to cisplatin in human oral squamous cell carcinoma

“…Western blotting. Cell lysates were subjected to western blotting as described previously (17). The primary antibodies used were rabbit polyclonal antibodies against YAP, phospho-YAP (serine-127), TEAD, LATS1, LATS2, AKT, and phospho-AKT (serine-473) (Cell Signaling Technology, Boston, mA, USA), glutathione S-transferase (GST)-π (Calbiochem, USA), ATP7B, ERCC1, P73 (Santa Cruz Biotechnology, Santa Cruz, CA, USA).…”

“…Transfection of siRNAs was conducted as described previously (17). Cells were cultured in DmEm supplemented with 10% FBS for 24 h and transfected with 5 µm siRNA using Thermo Scientific DharmaFECT Transfection reagents (Roche, Indianapolis, IN, USA) according to the manufacturer's instructions.…”