Regulated GPCR Trafficking to the Plasma Membrane: General Issues and the CCR5 Chemokine Receptor Example - GPCR Signalling Complexes: Synthesis, Assembly, Trafficking and Specificity - page 95

and subsequent lysosomal targeting of the activated receptors, whereas the intracellular
pool was protected and translocated, instead, to the plasma membrane, leading to
the recovery of thrombin responsiveness. However, despite the interest raised by the
observation of a signal-regulated export of PAR1, the current model of unregulated
cell surface targeting was not challenged for other GPCRs, since PAR1 is activated
by a unique irreversible proteolytic mechanism, which unmasks a tethered ligand,
whereas the b2-adrenoceptor used as control in the same study was not found in
cytoplasmic stores.
In the late 1990s, a series of apparently disconnected observations converged to
suggest that the regulated export GPCRs to the plasma membrane from the Golgi
and ER was actually a much wider phenomenon than anticipated previously.
First, studies in both primary cells and cells expressing exogenous receptors,
revealed the existence of additional GPCRs, which display a similar phenomenon of
translocation to the cell surface from the cytoplasm as for PAR1. Consistent with a
sustained clinical efficacy in patients, agonists active on the dopamine D2 L receptor
were found to promote up-regulation of surface receptors in Sf9 cells due to the
redistribution of pre-existing pools (Ng et al. 1997 ). Similarly, regulated pools of
intracellular endogenous dopamine D1 receptors were reported in tubular renal cells.
In these cells, receptor recruitment from cytosolic stores to the plasma membrane was
elicited by agonist activation of cell surface D1 receptors (Brismar et al. 1998 ) or via
atrial natriuretic peptide-dependent heterologous activation (Holtback et al. 1999 ) .
An analogous phenomenon was also reported for a 1A -adrenoceptors in response to
neuropeptide Y stimulation (Holtback et al. 1999 ) . Finally, continued agonist
exposure of the human somatostatin type 1 receptor (hSSTR1) stably expressed in
Chinese hamster ovary-K1 (CHO-K1) cells was reported to induce protein synthesis-
independent up-regulation of functional surface receptors due to the recruitment
from a pre-existing cytoplasmic pool. This feature was dependent on molecular
signals identified in the receptor C-tail (Hukovic et al. 1999 ) .
The concept that the cell surface export of at least some GPCRs is submitted to
regulation was supported further by the concomitant observation that the capacity
of several receptors to reach the plasma membrane is markedly affected by the cell
type where they are expressed. For example, the thyrotropin-releasing hormone
receptor (TRH-R), which is predominantly located at the cell surface of pituitary
lactotrophs and AtT20 pituitary corticotrophs, is predominantly intracellular in
transfected HEK-293 and COS-7 cells, colocalizing with endoplasmic reticulum
and Golgi markers even if expressed at physiological levels (Yu and Hinkle 1997 ) .
Similarly, it was reported that olfactory receptors fail to traffic to the plasma mem-
brane in heterologous cells (McClintock et al. 1997 ) , except when overexpressed
in the baculovirus-insect sf9 cell system or in olfactory receptor neurons in vivo
(Gimelbrant et al. 1999 ). Moreover, it could be proved that intracellular retention
was dependent on some molecular signal carried by the receptors themselves, since
the phenotype could be reversed in chimeras containing intracellular domains of
GPCRs, which are predominantly localized at the cell surface (McClintock et al.
1997 ), or by receptor truncations (Gimelbrant et al. 1999 ) .