UC Berkeley

Epidermal growth factor receptor (EGFR) is a critical receptor tyrosine kinase and controls cellular functions, such as proliferation and differentiation, by activating the mitogen-activated protein kinase (MAPK) pathway. Mutations and overexpression of this protein are associated with many types of cancer and thereby, is the target of many therapeutic drugs. However, despite initial patient response to anti-EGFR therapeutics, long-term treatment remains a challenge due to inevitable acquired resistance and disease progression. Secondary mutations and amplification of bypass pathways are common resistance mechanisms, but other mechanisms of resistance are complex and not fully understood. For the development of improved therapeutics, a complete understanding of EGFR activation is required.

Before activation by a soluble ligand, EGFR is primarily monomeric and in an autoinhibited form. Upon ligand binding, EGFR undergoes conformational changes that lead to dimerization and activation of its intracellular kinase domain. In the canonical view, EGFR dimerization is sufficient for signaling, but further studies demonstrated higher-order EGFR assemblies may be required. These clusters alter the stoichiometry and catalytic rates of protein binders and associated enzymes, which modulate signaling in the process. However, the biophysical basis for EGFR clustering is not well understood. In this work presented here, we discuss our findings regarding EGFR protein condensates and its effect on downstream Ras signaling. We discovered that EGFR condenses into an extended bond percolation network through recruitment and dimerization of growth factor receptor-bound protein 2 (Grb2). EGFR condensation promotes the ability of guanine nucleotide exchange factor Son of Sevenless (SOS) to activate Ras. Furthermore, tyrosine phosphorylation in EGFR activation both promotes and inhibits Ras activation through competing effects on the EGFR protein condensate. We also demonstrate kinase inhibitors can promote Ras activation under certain conditions through a double negative effect. When measured in live mammalian cells, we find EGFR condensation is enhanced by Grb2. However, direct Grb2:Grb2 interactions are not required for condensation. Instead, Grb2:SOS interactions are the main driver. We further find that Grb2:Grb2 interactions can control EGFR condensation in the absence of Grb2:SOS interactions when Grb2 is highly expressed. Together, these insights uncover new and complex mechanisms controlling EGFR signaling.