Enhanced dimerization drives ligand-independent activity of mutant epidermal growth factor receptor in lung cancer.
Mutations within the epidermal growth factor receptor (EGFR/erbB1/Her1) are often associated with tumorigenesis. In particular, a number of EGFR mutants that demonstrate ligand-independent signaling are common in non-small cell lung cancer (NSCLC), including 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 molecular mechanisms by which these mutations confer constitutive activity remain unresolved. Using multiple subdiffraction-limit imaging modalities, we reveal the altered receptor structure and interaction kinetics of NSCLC-associated EGFR mutants. We applied two-color single 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-independent dimers. Two-color superresolution localization microscopy confirmed ligand-independent aggregation of EGFR mutants. Live-cell Förster resonance energy transfer measurements revealed that the L858R kinase mutation alters ectodomain structure such that unliganded mutant EGFR adopts an extended, dimerization-competent conformation. Finally, mutation of the putative dimerization arm confirmed a critical role for ectodomain engagement 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.
Department of Pathology and Cancer Research and Treatment Center, University of New Mexico, Albuquerque, NM 87131.
Christopher C
CC
Valley
Laboratory of Cellular Dynamics, Max Planck Institute for Biophysical Chemistry, 37077 Göttingen, Germany.
Donna J
DJ
Arndt-Jovin
III. Institute of Physics, Georg-August University of Göttingen, 37077 Göttingen, Germany.
Narain
N
Karedla
Department of Pathology and Cancer Research and Treatment Center, University of New Mexico, Albuquerque, NM 87131.
Mara P
MP
Steinkamp
III. Institute of Physics, Georg-August University of Göttingen, 37077 Göttingen, Germany.
Alexey I
AI
Chizhik
Theoretical Biology and Biophysics Group, Theoretical Division, Los Alamos National Laboratory, Los Alamos, NM 87545.
William S
WS
Hlavacek
Department of Pathology and Cancer Research and Treatment Center, University of New Mexico, Albuquerque, NM 87131.
Bridget S
BS
Wilson
Department of Physics and Astronomy, University of New Mexico, Albuquerque, NM 87131.
Keith A
KA
Lidke
Department of Pathology and Cancer Research and Treatment Center, University of New Mexico, Albuquerque, NM 87131 DLidke@salud.unm.edu.
Diane S
DS
Lidke
D000818:Animals; D016466:CHO Cells; D002289:Carcinoma, Non-Small-Cell Lung / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D049109:Cell Proliferation; D002471:Cell Transformation, Neoplastic; D003412:Cricetulus; D066255:EGF Family of Proteins / Q000235:genetics / Q000378:metabolism*; D066246:ErbB Receptors / Q000235:genetics / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D008175:Lung Neoplasms / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D018613:Microscopy, Confocal; D009154:Mutation; D010766:Phosphorylation; D066329:Protein Aggregates; D047428:Protein Kinase Inhibitors; D055503:Protein Multimerization; D015398:Signal Transduction
D066255:EGF Family of Proteins; D066329:Protein Aggregates; D047428:Protein Kinase Inhibitors; C512478:EGFR protein, human; D066246:ErbB Receptors