Molecular basis for multimerization in the activation of the epidermal growth factor receptor

Abstract

The epidermal growth factor receptor (EGFR) is activated by dimerization, but activation also generates higher-order multimers, whose nature and function are poorly understood. We have characterized ligand-induced dimerization and multimerization of EGFR using single-molecule analysis, and show that multimerization can be blocked by mutations in a specific region of Domain IV of the extracellular module. These mutations reduce autophosphorylation of the C-terminal tail of EGFR and attenuate phosphorylation of phosphatidyl inositol 3-kinase, which is recruited by EGFR. The catalytic activity of EGFR is switched on through allosteric activation of one kinase domain by another, and we show that if this is restricted to dimers, then sites in the tail that are proximal to the kinase domain are phosphorylated in only one subunit. We propose a structural model for EGFR multimerization through self-association of ligand-bound dimers, in which the majority of kinase domains are activated cooperatively, thereby boosting tail phosphorylation.DOI: http://dx.doi.org/10.7554/eLife.14107.001

Activation of the EGF Receptor by Ligand Binding and Oncogenic Mutations: The “Rotation Model”

“…More recently, EGFR-eGFP expressed in Xenopus oocytes at low levels, ~1 to ~5 molecules per μm 2 , existed in monomeric form when analyzed by photobleaching [56]. This is inconsistent with previous results in which EGFR expressed in CHO cells at low levels, ~3 molecules per μm 2 , was present in dimeric form [43].…”

“…Ligand-induced EGFR tetramerization through interaction of a region within Subdomain IV of the extracellular domain seems to be essential for full activation of the receptor [56,57]. In this model, phosphorylation of tyrosine residues in the C-terminal tail, proximal to the kinase domain, is facilitated by a continuous array of "activator" and "receiver" kinase domains of the receptor tetramers and oligomers.…”

“…Therefore, a Qdotconjugated EGF or Fab is likely to report EGFR dimers as monomers. Upon activation, the EGFR dimers become tetramers and oligomers through interaction between EGFR itself [56,57] as well as interaction with its effectors such as AP-2 and Shc1. This complex seems to interact with actin filaments for clustering while it moves toward sites for clathrin-dependent and -independent endocytosis [10].…”

EGF and NRG induce phosphorylation of HER3/ERBB3 by EGFR using distinct oligomeric mechanisms

“…In some studies, such indications came from the experiments in which FRET was detected between two EGF ligand molecules; because of the structural constraints of their interaction, this transfer is most logically explained by receptor oligomerization (17)(18)(19)(20)(21). Other studies imaged fluorescently labeled EGFR itself using image correlation microscopy (ICS) (22,23), fluorescence correlation spectroscopy (FCS) (24,25), stochastic optical reconstruction microscopy (STORM) (26), FRET, fluorescence lifetime imaging microscopy (FLIM) (27,28), spatial mapping of the receptor by immuno-electron microscopy (29,30), or number and brightness analysis (31). This spectrum of experimental approaches all led to a unifying conclusion that EGFR is organized in larger clusters in response to ligand binding.…”

“…This spectrum of experimental approaches all led to a unifying conclusion that EGFR is organized in larger clusters in response to ligand binding. More recent studies provide evidence that these higher-order multimers are needed to achieve the complete spectrum of EGFR phosphorylation (21,25).…”