Many G-protein-coupled receptors carry C-terminal ligand motifs for PSD-95/discs large/ZO-1 (PDZ) domains; via interaction with PDZ domain-containing scaffold proteins, this allows for integration of ...receptors into signaling complexes. However, the presence of PDZ domain proteins attached to intracellular membranes suggests that PDZ-type interactions may also contribute to subcellular sorting of receptors. The protein interacting specifically with Tc10 (PIST; also known as GOPC) is a trans-Golgi-associated protein that interacts through its single PDZ domain with a variety of cell surface receptors. Here we show that PIST controls trafficking of the interacting β1-adrenergic receptor both in the anterograde, biosynthetic pathway and during postendocytic recycling. Overexpression and knockdown experiments show that PIST leads to retention of the receptor in the trans-Golgi network (TGN), to the effect that overexpressed PIST reduces activation of the MAPK pathway by β1-adrenergic receptor (β1AR) agonists. Receptors can be released from retention in the TGN by coexpression of the plasma membrane-associated scaffold PSD-95, which allows for transport of receptors to the plasma membrane. Stimulation of β1 receptors and activation of the cAMP pathway lead to relocation of PIST from the TGN to an endosome-like compartment. Here PIST colocalizes with SNX1 and the internalized β1AR and protects endocytosed receptors from lysosomal degradation. In agreement, β1AR levels are decreased in hippocampi of PIST-deficient mice. Our data suggest that PIST contributes to the fine-tuning of β1AR sorting both during biosynthetic and postendocytic trafficking.
Background: PIST/GOPC is a Golgi-associated protein that interacts with several G-protein-coupled receptors via its single PDZ domain.
Results: PIST retains β1-adrenergic receptors in intracellular compartments and interferes with receptor degradation after endocytosis.
Conclusion: PIST stabilizes the receptor in an intracellular compartment.
Significance: PDZ proteins associated with intracellular membranes confer specific features to the subcellular targeting of interacting receptors.
PSD-95/discs large/ZO-1 (PDZ) domain proteins integrate many G-protein coupled receptors (GPCRs) into membrane associated signalling complexes. Additional PDZ proteins are involved in intracellular ...receptor trafficking. We show that three PDZ proteins (SNX27, PIST and NHERF1/3) regulate the mouse somatostatin receptor subtype 5 (SSTR5). Whereas the PDZ ligand motif of SSTR5 is not necessary for plasma membrane targeting or internalization, it protects the SSTR5 from postendocytic degradation. Under conditions of lysosomal inhibition, recycling of the SSTR5 to the plasma membrane does not depend on the PDZ ligand. However, recycling of the wild type receptor carrying the PDZ binding motif depends on SNX27 which interacts and colocalizes with the receptor in endosomal compartments. PIST, implicated in lysosomal targeting of some membrane proteins, does not lead to degradation of the SSTR5. Instead, overexpressed PIST retains the SSTR5 at the Golgi. NHERF family members release SSTR5 from retention by PIST, allowing for plasma membrane insertion. Our data suggest that PDZ proteins act sequentially on the GPCR at different stages of its subcellular trafficking.
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Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
► We search for interaction partners of the G-protein coupled receptor SSTR5. ► Using the Ras recruitment system, we identify the palmitoyl transferase ZDHHC5. ► Interaction involves helix 8 of ...SSTR5, and the N-terminal part of ZDHHC5. ► Knockdown experiments show that ZDHHC5 is required for palmitoylation of SSTR5.
Many G-protein coupled receptors are palmitoylated in their C-terminal, intracellular regions. So far no enzymes responsible for this modification have been described. We identified an interaction of the membrane proximal helix 8 of somatostatin receptor 5 (SSTR5) with the N-terminal region of the putative palmitoyltransferase ZDHHC5 using the Ras recruitment interaction screening system. ZDHHC5 and SSTR5 are colocalized at the plasma membrane and can be efficiently coimmunoprecipitated from transfected cells. Coexpression of ZDHHC5 in HEK293 cells increased palmitoylation of SSTR5 whereas knock-down of endogenous ZDHHC5 by siRNAs decreased it. Our data identify the first palmitoyltransferase for a G-protein coupled receptor.
SSTR5
physically interacts
with
ZDHHC5
by
ras recruitment system
(View interaction)
SSTR5
and
ZDHHC5
colocalize by
fluorescence microscopy
(View interaction)
SSTR5
physically interacts
with
ZDHHC5
by
pull down
(View interaction)
A hippocampal neuron in primary culture stained with anti‐PIST (red) and anti‐neuroligin‐2 (green), a postsynaptic cell adhesion protein which is exclusively located at inhibitory synapses. The ...nucleus was stained with DAPI (blue). Note the typical distribution of neuroligin‐2 immunoreactivity reminiscent of inhibitory synapses at proximal dendrites and cell body, which is not matched by the PIST immunofluorescence signal. The latter is concentrated in the perikaryon in a juxtanuclear position and appears to be absent from postsynaptic sites. The Journal of Comparative Neurology, Volume 520, Number 5, pages 889–913.
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