Epstein–Barr virus LMP1 induces focal adhesions and epithelial cell migration through effects on integrin-α5 and N-cadherin

摘要

Epstein–Barr virus (EBV) is a γ-herpesvirus associated with human epithelial and B-cell malignancies. The EBV latent membrane protein (LMP) 1 is expressed in nasopharyngeal carcinoma (NPC) and promotes oncogenic intracellular signaling mechanisms. LMP1 also promotes a pro-migratory phenotype through potential effects on cell surface proteins, as expression of LMP1 induces an epithelial–mesenchymal transition (EMT) in epithelial cell lines. In this study, LMP1 was examined for potential effects on cadherin and integrin surface interactions, and assessed for biological effects on adhesion and motility to fibronectin. Expression of LMP1 in the non-tumorigenic epithelial cell line MCF10a induced an EMT-associated cadherin switch. The induced N-cadherin was ligated and localized to the cell surface as determined by triton-solubility and immunofluorescence assays. In addition, LMP1 induced the assembly of focal adhesions (FAs) with increased production of fibronectin in MCF10a and NP460hTERT-immortalized nasopharyngeal cells. Biochemical enrichment of fibronectin-associated proteins indicated that LMP1 selectively promoted the recruitment of integrin-α5 and Src family kinase proteins to FA complexes. Neutralizing antibodies to N-cadherin and integrin-α5, but not integrin-αV, blocked the adhesion and transwell motility of MCF10a cells to fibronectin induced by LMP1. LMP1-induced transwell motility was also decreased by Src inhibition with the PP2 kinase inhibitor and short hairpin RNAs. These studies reveal that LMP1 has multiple mechanisms to promote the adhesive and migratory properties of epithelial cells through induction of fibronectin and modulation of cell surface interactions involving integrin-α5 and N-cadherin, which may contribute to the metastatic potential of NPC.