期刊
MOLECULAR BIOLOGY OF THE CELL
卷 26, 期 22, 页码 4087-4099出版社
AMER SOC CELL BIOLOGY
DOI: 10.1091/mbc.E15-05-0269
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资金
- National Science Foundation (NSF) [MCB-0845062]
- Oxnard Foundation
- National Institutes of Health (NIH) [P50GM085273]
- NSF [0954836]
- Max Planck Society
- Deutsche Forschungsgemeinschaft [SFB 937]
- NIH
- Division Of Physics
- Direct For Mathematical & Physical Scien [0954836] Funding Source: National Science Foundation
- Div Of Molecular and Cellular Bioscience
- Direct For Biological Sciences [0845062] Funding Source: National Science Foundation
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., Delta L747-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 Forster 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.
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