Lipid Raft-Mediated Mechanotransduction Drives OSCC Aggressiveness via EGFR/p-FAK/ZRF1 Axis

Abstract

Mechanical stresses generated through tissue stiffness and cytoskeletal dynamics are critical to cellular behavior, especially in cancer development. Mechanosensitive proteins in plasma membrane lipid rafts transmit these stresses, affecting intracellular signaling pathways. We here examined the effects of matrix stiffness on oral squamous cell carcinoma (OSCC) cells FaDu and SCC9, cultured on chitosan-coated dishes with increasing concentrations (1%, 2%, and 3%) to mimic the increase in stiffness. Improved stiffness was associated with increased cell migration, as supported by actin cytoskeleton reorganization immunofluorescence imaging and wound-healing migration assays. Interference with lipid rafts by methyl-β-cyclodextrin (MβCyD) disrupted mechanotransduction, inducing dysregulation of EGFR/Ezrin/phospho-FAK (p-FAK) signaling pathway and actin cytoskeleton disruption. FAK inhibition by PF-573228 mimicked these effects, highlighting FAK's pivotal role in cytoskeletal integrity and force sensing. Both manipulations inhibited epithelial-mesenchymal transition (EMT), characterized by loss of mesenchymal markers and decreased migratory ability. In contrast, overexpression of FAK rescued EMT phenotypes. Mechanistically, we identified an epigenetic modifier, Zuotin-related factor 1 (ZRF1) as a downstream effector of the EGFR/Ezrin/p-FAK pathway. MβCD and PF-573228 treatments reduced ZRF1 nuclear location, whereas FAK overexpression increased it. Co-immunoprecipitation verified ZRF1-p-FAK interaction, with ZRF1 also undergoing tyrosine phosphorylation. siRNA-mediated ZRF1 knockdown reduced EMT markers, whereas ZRF1 overexpression enhanced them, indicating ZRF1's regulatory role in EMT and epigenetic reprogramming. Collectively, our findings delineate the EGFR/Ezrin/p-FAK/ZRF1 axis as a key mechanotransduction pathway driving OSCC aggressiveness, offering novel therapeutic avenues for targeting cancer aggressiveness through mechanotransduction interference.