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09:07
Detecting the Ligand-binding Domain Dimerization Activity of Estrogen Receptor Alpha Using the Mammalian Two-Hybrid Assay
Published on: December 19, 2018
ErbB1 dimerization is promoted by domain co-confinement and stabilized by ligand binding
Shalini T Low-Nam1, Keith A Lidke, Patrick J Cutler
1Department of Pathology, University of New Mexico School of Medicine, Albuquerque, New Mexico, USA.
Nature Structural & Molecular Biology
|October 25, 2011
Summary
Ligand binding stabilizes human epidermal growth factor receptor 1 (erbB1) dimers, making them long-lived and less mobile. This receptor dimerization and reduced mobility are crucial for erbB1 signaling, independent of kinase activity.
Area of Science:
- Cell biology
- Molecular signaling
- Biophysics
Background:
- The role of ligand occupancy and dimerization in epidermal growth factor receptor 1 (erbB1) signaling remains debated.
- Understanding the dynamics of erbB1 dimerization is key to elucidating its signaling mechanisms.
Purpose of the Study:
- To investigate the contribution of ligand occupancy and dimerization to erbB1 signaling.
- To quantify the off-rate (k(off)) and mobility of erbB1 dimers in living cells.
Main Methods:
- Utilized two-color quantum-dot tracking to visualize human erbB1 homodimerization.
- Employed a three-state hidden Markov model to analyze kinetic parameters and transition rates between receptor states.
- Measured dimer off-rates and receptor mobility under various conditions, including ligand binding and kinase activity blockade.
Main Results:
- Ligand-bound erbB1 dimers are long-lived with an off-rate independent of kinase activity.
- Unliganded erbB1 dimers exhibit a significantly faster off-rate (over four times).
- Receptor dimerization markedly reduces mobility (over six-fold decrease), a process influenced by kinase activity and the actin cytoskeleton.
Conclusions:
- Ligand binding stabilizes erbB1 dimers, impacting their kinetics and mobility.
- The reduced mobility of signaling-competent erbB1 dimers is linked to both signal propagation and the cortical cytoskeleton.
- These findings highlight the importance of dimerization dynamics and cytoskeletal interactions in erbB1-mediated signaling.
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