Zyxin Is a Transforming Growth Factor-β (TGF-β)/Smad3 Target Gene That Regulates Lung Cancer Cell Motility via Integrin α5β1

Although TGF-beta acts as a tumor suppressor in normal tissues and in early carcinogenesis, these tumor suppressor effects are lost in advanced malignancies. Single cell migration and epithelial-mesenchymal transition (EMT), both of which are regulated by TGF-beta, are critical steps in mediating cancer progression. Here, we sought to identify novel direct targets of TGF-beta signaling in lung cancer cells and have indentified the zyxin gene as a target of Smad3-mediated TGF-beta 1 signaling. Zyxin concentrates at focal adhesions and along the actin cytoskeleton; as such, we hypothesized that cytoskeletal organization, motility, and EMT in response to TGF-beta 1 might be regulated by zyxin expression. We show that TGF-beta 1 treatment of lung cancer cells caused rapid phospho-Smad3-dependent expression of zyxin. Zyxin expression was critical for the formation and integrity of cell adherens junctions. Silencing of zyxin decreased expression of the focal adhesion protein vasodilator-activated phosphoprotein (VASP), although the formation and morphology of focal adhesions remained unchanged. Zyxin-depleted cells displayed significantly increased integrin alpha 5 beta 1 levels, accompanied by enhanced adhesion to fibronectin and acquisition of a mesenchymal phenotype in response to TGF-beta 1. Zyxin silencing led to elevated integrin alpha 5 beta 1-dependent single cell motility. Importantly, these features are mirrored in the K-ras-driven mouse model of lung cancer. Here, lung tumors revealed decreased levels of both zyxin and phospho-Smad3 when compared with normal tissues. Our data thus demonstrate that zyxin is a novel functional target and effector of TGF-beta signaling in lung cancer. By regulating cell-cell junctions, integrin alpha 5 beta 1 expression, and cell-extracellular matrix adhesion, zyxin may regulate cancer cell motility and EMT during lung cancer development and progression.