Formation and dissociation of M 1 muscarinic receptor dimers seen by total internal reflection fluorescence imaging of single molecules
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- 20 January 2010
- journal article
- research article
- Published by Proceedings of the National Academy of Sciences in Proceedings of the National Academy of Sciences of the United States of America
- Vol. 107 (6), 2693-2698
- https://doi.org/10.1073/pnas.0907915107
Abstract
G-protein-coupled receptors (GPCRs) are the largest family of transmembrane signaling proteins in the human genome. Events in the GPCR signaling cascade have been well characterized, but the receptor composition and its membrane distribution are still generally unknown. Although there is evidence that some members of the GPCR superfamily exist as constitutive dimers or higher oligomers, interpretation of the results has been disputed, and recent studies indicate that monomeric GPCRs may also be functional. Because there is controversy within the field, to address the issue we have used total internal reflection fluorescence microscopy (TIRFM) in living cells to visualize thousands of individual molecules of a model GPCR, the M-1 muscarinic acetylcholine receptor. By tracking the position of individual receptors over time, their mobility, clustering, and dimerization kinetics could be directly determined with a resolution of similar to 30 ms and similar to 20 nm. In isolated CHO cells, receptors are randomly distributed over the plasma membrane. At any given time, similar to 30% of the receptor molecules exist as dimers, and we found no evidence for higher oligomers. Two-color TIRFM established the dynamic nature of dimer formation with M-1 receptors undergoing interconversion between monomers and dimers on the timescale of seconds.Keywords
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