2018
In Receptor-Receptor Interactions in the Central Nervous System (pp. 329-343). Humana Press

Super-resolution imaging as a method to study GPCR dimers and higher-order oligomers

Authors:

Jonas, K. C., & Hanyaloglu, A. C.

Keywords:

GPCR, dimer, oligomer, super-resolution, PALM, single molecule imaging

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 receptor–receptor 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 <10 nm 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.