EGFR endocytosis down-regulates binding to EphA2 at the plasma membrane.

The epidermal growth factor receptor (EGFR) is a membrane protein that is essential to growth, differentiation, and survival in healthy cells. Misregulation of EGFR causes these pathways to function abnormally, leading to tumorigenesis. There are several therapeutics targeting EGFR at the clinic. Unfortunately, EGFR-driven tumors often develop resistance to therapeutics, hampering long-term cancer treatment with EGFR inhibitors. Overexpression of the receptor EphA2 is a common mechanism of EGFR therapeutic resistance. Importantly, EGFR and EphA2 interact at the plasma membrane, but the factors that control this interaction are poorly understood. Here, we investigate how EGFR and EphA2 interact functionally and physically. Immunofluorescence and proximity ligation data indicate that EGFR-EphA2 interactions rapidly decrease after EGFR activation with EGF. Inhibition of endocytosis blocked the effect of EGF on co-localization between both receptors, indicating that endocytosis of EGFR impairs its ability to form hetero-complexes with EphA2. Pharmacological inhibition of EGFR with tyrosine kinase inhibitors was only partially able to block this effect. Our data indicate that endocytosis is an important negative regulatory factor that impacts the levels of EGFR-EphA2 complexes. This information has implications for EGFR drug resistance in cancer.