The C-Terminal Domains of the GABA_BReceptor Subunits Mediate Intracellular Trafficking But Are Not Required for Receptor Signaling

Abstract

GABA B receptors are G-protein-coupled receptors that mediate slow synaptic inhibition in the brain and spinal cord. These receptors are heterodimers assembled from GABA B1 and GABA B2 subunits, neither of which is capable of producing functional GABA B receptors on homomeric expression. GABA B1, although able to bind GABA, is retained within the endoplasmic reticulum (ER) when expressed alone. In contrast, GABA B2 is able to access the cell surface when expressed alone but does not couple efficiently to the appropriate effector systems or produce any detectable GABA-binding sites. In the present study, we have constructed chimeric and truncated GABA B1 and GABA B2 subunits to explore further GABA B receptor signaling and assembly. Removal of the entire C-terminal intracellular domain of GABA B1 results in plasma membrane expression without the production of a functional GABA B receptor. However, coexpression of this truncated GABA B1 subunit with either GABA B2 or a truncated GABA B2 subunit in which the C terminal has also been removed is capable of functional signaling via G-proteins. In contrast, transferring the entire C-terminal tail of GABA B1 to GABA B2 leads to the ER retention of the GABA B2 subunit when expressed alone. These results indicate that the C terminal of GABA B1 mediates the ER retention of this protein and that neither of the C-terminal tails of GABA B1 or GABA B2 is an absolute requirement for functional coupling of heteromeric receptors. Furthermore although GABA B1 is capable of producing GABA-binding sites, GABA B2 is of central importance in the functional coupling of heteromeric GABA B receptors to G-proteins and the subsequent activation of effector systems.

Glutamate Receptor Trafficking: Endoplasmic Reticulum Quality Control Involves Ligand Binding and Receptor Function

“…Oligomeric proteins assembled in the proper conformation escape ER retention by association and masking of their ER retention/retrieval motifs, whereas improper conformations leave these motifs exposed to COPImediated retrieval. Such mechanisms ensuring that only proper oligomeric forms are expressed on the cell surface were shown recently to regulate trafficking of GluR6 KA receptors (S. , NMDA receptors (Okabe et al, 1999;Standley et al, 2000;Scott et al, 2001), nicotinic acetylcholine receptors (Keller et al, 2001), serotonin type-3 receptors (Boyd et al, 2002), K ϩ channels (Zerangue et al, 1999(Zerangue et al, , 2001Hough et al, 2000), Ca 2ϩ channels , and various G-protein-coupled receptors (Margeta-Mitrovic et al, 2000;Calver et al, 2001;Chan et al, 2001;Pagano et al, 2001;Grunewald et al, 2002).…”

Heterodimeric coiled-coil interactions of human GABA B receptor

“…GBR1 is trapped within the ER when expressed alone (12) but can reach the cell surface upon association with GBR2 (9,11). Mutation or removal of the ER retention signal in GBR1 results in plasma membrane expression of GBR1 (9)(10)(11). Furthermore, interaction between the coiled-coil domains of GBR1 and GBR2 masks this ER retention signal to facilitate the cell surface expression of both subunits (9)(10)(11).…”

“…Mutation or removal of the ER retention signal in GBR1 results in plasma membrane expression of GBR1 (9)(10)(11). Furthermore, interaction between the coiled-coil domains of GBR1 and GBR2 masks this ER retention signal to facilitate the cell surface expression of both subunits (9)(10)(11). Although mutation of the di-leucine motif itself is not sufficient to release GBR1 from intracellular retention, it enhances cell surface expression of various GBR1 mutants that lack the ER retention signal (9).…”

Lateral Diffusion of the GABAB Receptor Is Regulated by the GABAB2 C Terminus

“…The subunits appear to serve different functions within the fully formed receptor. GABA B1 contains the agonist binding site on its large extracellular N terminus, and the affinity of this site for agonists is increased following heterodimerization with GABA B2 (7). The GABA B2 subunit contains intracellular loops that couple the receptor to the G-protein (8 -10).…”

“…Heterodimerization of the GABA B subunits is important not only for proper receptor function but also for forward trafficking of the receptor to the cell surface (5,7). In the absence of GABA B2 , the GABA B1 subunit is retained within the endoplasmic reticulum due to the presence of a C-terminal RSRR retention motif on its C terminus.…”