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research article

Visualizing odorant receptor trafficking in living cells down to the single-molecule level

Jacquier, Valérie  
•
Prummer, Michael  
•
Segura, Jean-Manuel  
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2006
Proceedings Of The National Academy Of Sciences Of The United States Of America (PNAS)

Despite the importance of trafficking for regulating G protein-coupled receptor signaling, for many members of the seven transmembrane helix protein family, such as odorant receptors, little is known about this process in live cells. Here, the complete life cycle of the human odorant receptor OR17-40 was directly monitored in living cells by ensemble and single-molecule imaging, using a double-labeling strategy. While the overall, intracellular trafficking of the receptor was visualized continuously by using a GFP tag, selective imaging of cell surface receptors was achieved by pulse-labeling an acyl carrier protein tag. We found that OR17-40 efficiently translocated to the plasma membrane only at low expression, whereas at higher biosynthesis the receptor accumulated in intracellular compartments. Receptors in the plasma membrane showed high turnover resulting from constitutive internalization along the clathrin pathway, even in the absence of ligand. Single-molecule microscopy allowed monitoring of the early, dynamic processes in odorant receptor signaling. Although mobile receptors initially diffused either freely or within domains of various sizes, binding of an agonist or an antagonist increased partitioning of receptors into small domains of approximately 190 nm, which likely are precursors of clathrin-coated pits. The binding of a ligand, therefore, resulted in modulation of the continuous, constitutive internalization. After endocytosis, receptors were directed to early endosomes for recycling. This unique mechanism of continuous internalization and recycling of OR17-40 might be instrumental in allowing rapid recovery of odor perception.

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Type
research article
DOI
10.1073/pnas.0603942103
Web of Science ID

WOS:000240968100015

Author(s)
Jacquier, Valérie  
Prummer, Michael  
Segura, Jean-Manuel  
Pick, Horst  
Vogel, Horst  
Date Issued

2006

Publisher

National Academy of Sciences

Published in
Proceedings Of The National Academy Of Sciences Of The United States Of America (PNAS)
Volume

103

Issue

39

Start page

14325

End page

14330

Subjects

cell signaling

•

G protein-coupled receptors

•

single-particle tracking

•

in vivo protein labeling

•

fluorescence imaging

Editorial or Peer reviewed

REVIEWED

Written at

EPFL

EPFL units
LCPPM  
Available on Infoscience
November 2, 2006
Use this identifier to reference this record
https://infoscience.epfl.ch/handle/20.500.14299/235448
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