Investigating the Homotypic and Heterotypic Interactions of ErbB Receptor Tyrosine Kinases

Abstract

Human epidermal growth factor receptor family (HER1-4 or ErbB1-4) is vital to cellular homeostasis, and any dysregulation of the receptors leads to serious diseases such as cancer. HER family receptors share similar structures: a ligand-binding extracellular domain, transmembrane domain, juxtamembrane domain, kinase domain, and a C-terminal tail. HER2 has an extracellular domain with an intrinsically closed conformation that does not bind to ligands, and HER3 has a catalytically inactive kinase domain. HER family receptors form an asymmetric dimer to activate the downstream signaling upon ligand binding. With their structural features and their roles in tumorigenesis, researchers have vastly studied EGFR homodimerization and HER1-HER2/ HER2-HER3 heterodimerization as therapeutic targets. However, collective understanding on the communication and the regulation of the HER family network is still elusive due to the difficulties of quantitative assessments of direct receptor-receptor interactions. Here we use pulsed interleaved excitation fluorescence cross correlation spectroscopy (PIE-FCCS) to assess the direct homotypic and heterotypic receptor-receptor interactions of the HER family in live cells before and after ligand stimulation. PIE-FCCS allows us to evaluate the receptor-receptor interactions with quantitative dynamic parameters such as diffusion coefficient, density, binding affinity, and degree of protein co-localization. Accompanied by biochemical functional assays, this study will shed light on the regulatory effect of HER family interactions on downstream signaling.