The Role of Alternative Epidermal Growth Factor Receptor Trafficking in Driving Cancer Progression

Abstract

The Epidermal Growth Factor Receptor (EGFR) is associated with a variety of cancers, including brain, lung, cervix, renal and breast. It is part of a family of receptors known as the ErbB receptors (ErbB1/EGFR, ErbB2/HER2, ErbB3/HER3, and ErbB4/HER4), transmembrane proteins found on epithelial cells responsible for a multitude of signaling events. In cancers, EGFR is frequently mutated or improperly expressed, upregulated in more than 50 percent of basal-like cancers. Mutations commonly promote constitutive activation or increase receptor recycling. In basal-like breast cancers such as triple negative breast cancer (TNBC), named for the lack of hormone receptors (estrogen and progesterone) and the HER2 receptor, EGFR is highly upregulated and associated with a variety of oncogenic activity, including increased proliferation and migration, and inhibition of cell death. Changes in these pathways are predicated on altered trafficking and activation of EGFR, events driven by variation in stimuli and interacting partners, such as other ErbB family members or oncogenic adaptor proteins such as MUC1, a member of the mucin family. In TNBC, upon stimulus with epidermal growth factor (EGF), EGFR colocalizes with MUC1 in intracellular vesicles distributed throughout the cytoplasm. These intracellular vesicles are associated with early endosomes, as indicated by the presence of early endosome antigen 1 (EEA1). Association with MUC1 prolongs the presence of EGFR in these vesicles, as EGFR's stay is significantly reduced in cells lacking MUC1. Retention in these vesicles by MUC1 inhibits trafficking of EGFR to the lysosome for degradation and is also associated with an increase in EGF-dependent migratory ability. Introduction of late endosome inhibitors (thereby preventing lysosomal targeting) increases migration in the absence of MUC1, the same effect as in the presence of MUC1. Further, inhibition of retrograde trafficking significantly decreases the rate of migration and changes cellular distribution of filopodia corresponding to migratory ability in MUC1-containing cells. Taken together, these data indicate that MUC1 is responsible for altering EGFR trafficking by retaining EGFR in EEA1-positive vesicles for prolonged periods, allowing for increased signal transduction through retrograde trafficking of EGFR and structural reorganization promoting a migratory phenotype. Loss of the polarity protein Llgl1 is associated with alterations in EGFR trafficking, promoting highly diffuse EGFR distribution throughout the cytoplasm versus along basolateral membranes. These changes in trafficking are also associated with increases in AKT and dual-phosphorylated-ERK signal transduction, both downstream targets of activated EGFR. Altering localization of EGFR to other membranes and intracellular vesicles without inducing polarity loss through a point mutation at amino acid 667 was found to also upregulate the AKT pathway. Mislocalization driven by polarity loss or point mutation in the basolateral targeting domain is sufficient to increase migration speeds of non-cancerous epithelial cell lines in vitro. This increased oncogenic activity is likely attributed to increased nuclear localization of the transcription factor TAZ (transcription co-activator with a PDZ-binding domain), whose nuclear translocation is associated with increased stem-like properties such as migration and survival. Together, these data reveal the oncogenic potential caused by alterations in EGFR trafficking that occur when polarity is lost or EGFR is improperly associated with proteins that promote changes to canonical EGFR localization and degradation, such as MUC1.