Enhanced dimerization drives ligand-independent activity of mutant epidermal growth factor receptor in lung cancer

2015 
Mutations within the epidermal growth factor receptor (EGFR/erbB1/Her1) are often associated with tumorigenesis. In particular, a number of EGFR mutants that demon- strate ligand-independent signaling are common in non-small cell lung cancer (NSCLC), in- cluding kinase domain mutations L858R (also called L834R) and exon 19 deletions (e.g., ΔL747-P753insS), which collectively make up nearly 90% of mutations in NSCLC. The mole- cular mechanisms by which these mutations confer constitutive activity remain unresolved. Using multiple subdiffraction-limit imaging modalities, we reveal the altered receptor struc- ture and interaction kinetics of NSCLC-associated EGFR mutants. We applied two-color sin- gle quantum dot tracking to quantify receptor dimerization kinetics on living cells and show that, in contrast to wild-type EGFR, mutants are capable of forming stable, ligand-indepen- dent dimers. Two-color superresolution localization microscopy confirmed ligand-indepen - dent aggregation of EGFR mutants. Live-cell Forster resonance energy transfer measure- ments revealed that the L858R kinase mutation alters ectodomain structure such that unliganded mutant EGFR adopts an extended, dimerization-competent conformation. Final- ly, mutation of the putative dimerization arm confirmed a critical role for ectodomain engage - ment in ligand-independent signaling. These data support a model in which dysregulated activity of NSCLC-associated kinase mutants is driven by coordinated interactions involving both the kinase and extracellular domains that lead to enhanced dimerization.
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