1 cJun N-terminal Kinase 2 (JNK2) enhances cell migration through Epidermal Growth Factor Substrate 8 (EPS8) Shreya Mitra 1,2 , Ju-Seog Lee 2 , Michael Cantrell 3 , Carla Lynn Van Den Berg 1,3§ 1 College of Pharmacy, Division of Pharmacology/Toxicology and Center for Molecular and Cellular Toxicology, 3 Institute of Cellular & Molecular Biology, University of Texas, Austin, 1 University Avenue A1915, Austin, Texas 78712-0125 2 Department of Systems Biology, Division of Cancer Medicine, University of Texas MD Anderson Cancer Center, Houston, Texas 77054 § Address correspondence to: Carla Van Den Berg, Tel.: (512) 471-5199; Fax: (512) 475-6088; E-mail: cvandenberg@mail.utexas.edu Running Title: JNK2 mediates migration through EPS8 ABSTRACT Membrane bound receptors induce biochemical signals to remodel the actin cytoskeleton and mediate cell motility. In association with receptor tyrosine kinases (RTKs), several downstream mitogen induced kinases facilitate cell migration. Herein, we show a role for cJun N-terminal kinase 2 (JNK2) in promoting mammary cancer cell migration through inhibition of Epidermal Growth Factor Substrate 8 (EPS8) expression, a key regulator of EGFR signaling and trafficking. Using jnk2-/- mice, we found that EPS8 expression is higher in Polyoma Middle T Antigen (PyVMT)jnk2-/- mammary tumors and jnk2-/- mammary glands compared to the respective jnk2+/+ controls. The inverse relationship between jnk2 and eps8 expression was also associated with cancer progression in that patients with Basal-type breast tumors expressing high jnk2 and low eps8 experienced poor disease free survival. In mammary tumor cell lines, absence of jnk2 greatly reduces cell migration which is rescued by EPS8 knockdown. Subsequent studies show that JNK2 enhances formation of the EPS8/Abi-1/Sos1 complex to augment EGFR activation of Akt and ERK, whereas; absence of JNK2 promotes ESP8 /RN-Tre association to inhibit endocytotic trafficking of the EGFR. Together, these studies unveil a critical role for JNK2 and EPS8 in RTK signaling and trafficking to convey distinctly different effects on cell migration. INTRODUCTION Eukaryotic cell migration is essential for embryonic development and tissue homeostasis. In cancer, tumor progression is associated with cell extravasation into the surrounding tissue and vasculature. Up- regulation of motility-associated genes in tumors increases sensitivity to micro- environmental cues and leads to an invasive phenotype. Since the survival rate of metastatic breast cancer remains dismal, proteins involved in cancer cell migration are promising prognostic markers or targets for treatment of metastatic disease. Cell migration commences in response to chemotactic cues which leads to transient cell polarization and accumulation of signaling complexes at the leading edge of the cell. This response is facilitated by the intrinsic vesicular trafficking machinery (1) and results in actin polymerization to form lamellipodia at the advancing end (2-4). http://www.jbc.org/cgi/doi/10.1074/jbc.M109.094441 The latest version is at JBC Papers in Press. Published on February 28, 2011 as Manuscript M109.094441 Copyright 2011 by The American Society for Biochemistry and Molecular Biology, Inc.