Biology Reference
In-Depth Information
Abstract
G protein-coupled receptors (GPCRs) are classified into a family of seven transmem-
brane receptors. Receptor oligomerization may be the key to the expression and func-
tion of these receptors, for example, ligand binding, desensitization, membrane
trafficking, and signaling. The accumulation of evidence that GPCRs form an olig-
omerization with a functional alternation may change the strategy for the discovery
of novel drugs targeting GPCRs. Identification of the oligomer is essential to eluci-
date GPCR oligomerization. GPCR oligomerizations have been demonstrated using
various biochemical approaches, which include the coimmunoprecipitation method,
fluorescence resonance energy transfer assay, and bioluminescence RET assay.
Thus, various assays are useful for the study of GPCR oligomerization, and we
should choose the best method to match the purpose. We previously targeted aden-
osine A
1
and thromboxane A
2
(TP) receptors to form a functionally novel hetero-
oligomer, since both receptors function in the same cells. This chapter describes
the methods used to detect GPCR oligomerization and alterations in the signaling
pathways, principally according to our findings on oligomerization between adeno-
sine A
1
and TP
a
receptors.
INTRODUCTION
G protein-coupled receptors (GPCRs) are classified into a family of seven transmem-
brane receptors and are the targets for various clinical drugs in the market today. In-
creasing evidence has shown that homo- and hetero-oligomerization formed by
various GPCRs play crucial roles in GPCR signaling (
Milligan, 2009
). Receptor
oligomerization may be the key to the expression and function of these receptors,
for example, ligand binding, desensitization, membrane trafficking, and signaling.
The GABA
B
receptor is known to be composed of two isoforms, the GABA
B1
and GABA
B2
subunits. Hetero-oligomerization between GABA
B1
and GABA
B2
pro-
duces the fully functional receptor at the cell surface but is nonfunctional when each
monomer is expressed individually (
Jones et al., 1998; Kaupmann et al., 1998
).
Studies on GPCR oligomerization would be valuable in understanding their roles
in pathological conditions. Oligomerization may increase the diversity of GPCR
signaling.
Identification of the oligomer is essential to elucidate GPCR oligomerization. Re-
cently, predicting the interacting partners among GPCRs has been performed in sev-
eral studies. Until now, GPCR oligomerizations have been demonstrated using
various biochemical approaches.
In the early stages, the coimmunoprecipitation method, which is one of the
biochemical methods generally used for GPCR oligomerization, was used to identify
the
b
2
-adrenergic receptor homo-oligomer (
Hebert et al., 1996
). Coimmunoprecipi-
tation strategies are simple and do not require special equipment. However, this
