Super-resolution imaging as a method to Study GPCR dimers and higher-order oligomers
File(s)
Accepted version
Author(s)
Jonas, Kim C
Hanyaloglu, AC
Type
Chapter
Abstract
The study of G protein-coupled receptor (GPCR) dimers and higher order oligomers
has unveiled mechanisms for receptors to diversify signaling and potentially uncover
novel therapeutic targets. The functional and clinical significance of these receptorreceptor associations has been facilitated by the development of techniques and
protocols, enabling researchers to unpick their function from the molecular
interfaces, to demonstrating functional significance in vivo, in both health and
disease. Here we describe our methodology to study GPCR oligomerization at the
single molecule level via super-resolution imaging. Specifically, we have employed
photoactivated localization microscopy, with photoactivatable dyes (PD-PALM) to
visualize the spatial organization of these complexes to <10nm resolution, and the
quantitation of GPCR monomer, dimer and oligomer in both homomeric and
heteromeric forms. We provide guidelines on optimal sample preparation, imaging
parameters and necessary controls for resolving and quantifying single molecule
data. Finally, we discuss advantages and limitations of this imaging technique and its
potential future applications to the study of GPCR function.
has unveiled mechanisms for receptors to diversify signaling and potentially uncover
novel therapeutic targets. The functional and clinical significance of these receptorreceptor associations has been facilitated by the development of techniques and
protocols, enabling researchers to unpick their function from the molecular
interfaces, to demonstrating functional significance in vivo, in both health and
disease. Here we describe our methodology to study GPCR oligomerization at the
single molecule level via super-resolution imaging. Specifically, we have employed
photoactivated localization microscopy, with photoactivatable dyes (PD-PALM) to
visualize the spatial organization of these complexes to <10nm resolution, and the
quantitation of GPCR monomer, dimer and oligomer in both homomeric and
heteromeric forms. We provide guidelines on optimal sample preparation, imaging
parameters and necessary controls for resolving and quantifying single molecule
data. Finally, we discuss advantages and limitations of this imaging technique and its
potential future applications to the study of GPCR function.
Date Issued
2018-08-02
Citation
Receptor-receptor interactions in the central nervous system, 2018, pp.329-343
Publisher
Springer
Start Page
329
End Page
343
Journal / Book Title
Receptor-receptor interactions in the central nervous system
Copyright Statement
© Springer Science+Business Media, LLC, part of Springer Nature 2018
Sponsor
Biotechnology and Biological Sciences Research Council (BBSRC)
Grant Number
BB/I008004/1
Article Number
21
Date Publish Online
2018-08-02