The M muscarinic receptor is present in situ as a ligand-regulated mixture of monomers and oligomeric complexes.

The quaternary organization of rhodopsin-like G protein-coupled receptors in native tissues is unknown. To address this we generated mice in which the M muscarinic acetylcholine receptor was replaced with a C-terminally monomeric enhanced green fluorescent protein (mEGFP)-linked variant. Fluorescence imaging of brain slices demonstrated appropriate regional distribution, and using both anti-M and anti-green fluorescent protein antisera the expressed transgene was detected in both cortex and hippocampus only as the full-length polypeptide.

Chemokine receptor CXCR4 oligomerization is disrupted selectively by the antagonist ligand IT1t.

CXCR4, a member of the family of chemokine-activated G protein-coupled receptors, is widely expressed in immune response cells. It is involved in both cancer development and progression as well as viral infection, notably by HIV-1. A variety of methods, including structural information, have suggested that the receptor may exist as a dimer or an oligomer.

A general method to quantify ligand-driven oligomerization from fluorescence-based images.

Here, we introduce fluorescence intensity fluctuation spectrometry for determining the identity, abundance and stability of protein oligomers. This approach was tested on monomers and oligomers of known sizes and was used to uncover the oligomeric states of the epidermal growth factor receptor and the secretin receptor in the presence and absence of their agonist ligands. This method is fast and is scalable for high-throughput screening of drugs targeting protein-protein interactions.

Spatial Intensity Distribution Analysis: Studies of G Protein-Coupled Receptor Oligomerisation.

Spatial intensity distribution analysis (SpIDA) is a recently developed approach for determining quaternary structure information on fluorophore-labelled proteins of interest in situ. It can be applied to live or fixed cells and native tissue. Using confocal images, SpIDA generates fluorescence intensity histograms that are analysed by super-Poissonian distribution functions to obtain density and quantal brightness values of the fluorophore-labelled protein of interest.