Interaction of G-protein-coupled receptors with synaptic scaffolding proteins

2002 ◽  
Vol 30 (4) ◽  
pp. 464-468 ◽  
Author(s):  
H.-J. Kreienkamp ◽  
M. Soltau ◽  
D. Richter ◽  
T. Böckers
Keyword(s):  
Rat Brain ◽  
Somatostatin Receptor ◽  
Pdz Domain ◽  
Interacting Protein ◽  
Transmembrane Receptors ◽  
C Terminus ◽  
Large N ◽  
Brain Extracts ◽  
The Central Nervous System

The calcium-independent receptors for latrotoxin (CIRL1-CIRL3) constitute a family of seven-transmembrane receptors with an unsually large N-terminal extracellular domain which comprises several motifs usually found in cell adhesion molecules. By yeast two-hybrid screening, we have identified the intracellular C-termini of CIRL1 and CIRL2 as interaction partners of the PDZ domain of the proline-rich synapse-associated protein (ProSAP)/somatostatin receptor-interacting protein (SSTRIP) family of postsynaptic proteins (SSTRIP, ProSAP1 and ProSAP2, also known as shank1-shank3 respectively). Overlay assays indicate that the ProSAP1/shank2 PDZ domain in particular interacts strongly with the C-terminus of CIRL1 and CIRL2. Co-immuno-precipitation of ProSAP1 and CIRL1 (but not CIRL2) from rat brain extracts indicates that this interaction also occurs in vivo in rat brain. The known postsynaptic localization of ProSAP1, as well as our observation that CIRL1 (but not CIRL2) is enriched in postsynaptic density preparations from the rat brain, suggests that CIRL1 is localized pre- as well as post-synaptically in the central nervous system.

scholarly journals Interactions of GIPC with Dopamine D2, D3 but not D4 Receptors Define a Novel Mode of Regulation of G Protein-coupled Receptors

2004 ◽  
Vol 15 (2) ◽  
pp. 696-705 ◽  
Author(s):  
Freddy Jeanneteau ◽  
Jorge Diaz ◽  
Pierre Sokoloff ◽  
Nathalie Griffon
Keyword(s):  
Rat Brain ◽  
G Protein ◽  
Plasma Membranes ◽  
Pdz Domain ◽  
G Protein Coupled Receptors ◽  
Interacting Protein ◽  
R And D ◽  
C Terminus ◽  
Endogenous Proteins ◽  
G Protein Coupled

The C-terminus domain of G protein-coupled receptors confers a functional cytoplasmic interface involved in protein association. By screening a rat brain cDNA library using the yeast two-hybrid system with the C-terminus domain of the dopamine D3 receptor (D3R) as bait, we characterized a new interaction with the PDZ domain-containing protein, GIPC (GAIP interacting protein, C terminus). This interaction was specific for the dopamine D2 receptor (D2R) and D3R, but not for the dopamine D4 receptor (D4R) subtype. Pull-down and affinity chromatography assays confirmed this interaction with recombinant and endogenous proteins. Both GIPC mRNA and protein are widely expressed in rat brain and together with the D3R in neurons of the islands of Calleja at plasma membranes and in vesicles. GIPC reduced D3R signaling, cointernalized with D2R and D3R, and sequestered receptors in sorting vesicles to prevent their lysosomal degradation. Through its dimerization, GIPC acts as a selective scaffold protein to assist receptor functions. Our results suggest a novel function for GIPC in the maintenance, trafficking, and signaling of GPCRs.

Vasopressin inhibits calcium-coupled sodium efflux system in rat brain

1994 ◽  
Vol 266 (4) ◽  
pp. R1169-R1173 ◽  
Author(s):  
F. Kanda ◽  
A. I. Arieff
Keyword(s):  
Rat Brain ◽  
Neuronal Cell ◽  
Atp Synthesis ◽  
Sodium Efflux ◽  
Rat Brain Synaptosomes ◽  
Neuronal Membranes ◽  
Brain Synaptosomes ◽  
The Central Nervous System ◽  
Na Uptake

Centrally released vasopressin plays an important role in the regulation of brain water and electrolyte composition and can affect brain intracellular pH and ATP synthesis in vivo. In this study, we evaluated the effects of [Arg8]vasopressin (AVP) on the Na(+)-Ca2+ exchanger, an important pathway in the regulation of cell Ca2+ concentration. It was found that AVP inhibited the Na(+)-Ca2+ exchanger in rat brain synaptosomes. This effect was completely blocked by the vasopressin V1-receptor antagonist d(CH2)5[(O-Me) Tyr2, Arg8]vasopressin. In addition, the vasopressin V2-receptor agonist 1-desamino-8-D-arginine vasopressin had no effect on the Na(+)-Ca2+ exchanger in rat brain synaptosomes. Depletion of intracellular Ca2+ by caffeine also had no effect on the effect of AVP on the Na(+)-Ca2+ exchanger. Na+ uptake by other pathways was also evaluated. It was found that AVP had no effect on Na+ uptake by pathways other than the Na(+)-Ca2+ exchanger. It is concluded that AVP inhibits the Na(+)-Ca2+ exchanger in neuronal membranes through vasopressin V1 receptors. Since this pathway is important in the regulation of cell volume and cytosolic Ca2+ in excitable tissue, AVP may impair neuronal cell repolarization in the central nervous system.

scholarly journals Functional Analysis of the Murine Cytomegalovirus Chemokine Receptor Homologue M33: Ablation of Constitutive Signaling Is Associated with an Attenuated Phenotype In Vivo

2007 ◽  
Vol 82 (4) ◽  
pp. 1884-1898 ◽  
Author(s):  
Ruth Case ◽  
Emma Sharp ◽  
Tau Benned-Jensen ◽  
Mette M. Rosenkilde ◽  
Nicholas Davis-Poynter ◽  
...  
Keyword(s):  
Cell Surface ◽  
Salivary Glands ◽  
Surface Expression ◽  
Receptor Activation ◽  
Murine Cytomegalovirus ◽  
Cell Surface Expression ◽  
Transmembrane Receptors ◽  
C Terminus ◽  
The Impact

ABSTRACT The murine cytomegalovirus (MCMV) M33 gene is conserved among all betaherpesviruses and encodes a homologue of seven-transmembrane receptors (7TMR) with the capacity for constitutive signaling. Previous studies have demonstrated that M33 is important for MCMV dissemination to or replication within the salivary glands. In this study, we probed N- and C-terminal regions of M33 as well as known 7TMR signature motifs in transmembrane (TM) II and TM III to determine the impact on cell surface expression, constitutive signaling, and in vivo phenotype. The region between amino acids R340 and A353 of the C terminus was found to be important for CREB- and NFAT-mediated signaling, although not essential for phosphatidylinositol turnover. Tagging or truncation of the N terminus of M33 resulted in loss of cell surface expression. Within TM II, an F79D mutation abolished constitutive signaling, demonstrating a role, as in other cellular and viral 7TMR, of TM II in receptor activation. In TM III, the arginine (but not the asparagine) residue of the NRY motif (the counterpart of the common DRY motif in cellular 7TMR) was found to be essential for constitutive signaling. Selected mutations incorporated into recombinant MCMV showed that disruption of constitutive signaling for a viral 7TMR homologue resulted in a reduced capacity to disseminate to or replicate in the salivary glands. In addition, HCMV UL33 was found to partially compensate for the lack of M33 in vivo, suggesting conserved biological roles of the UL33 gene family.

scholarly journals CHIP promotes Runx2 degradation and negatively regulates osteoblast differentiation

2008 ◽  
Vol 181 (6) ◽  
pp. 959-972 ◽  
Author(s):  
Xueni Li ◽  
Mei Huang ◽  
Huiling Zheng ◽  
Yinyin Wang ◽  
Fangli Ren ◽  
...  
Keyword(s):  
Transcriptional Activation ◽  
Osteoblast Differentiation ◽  
Degradation Mechanism ◽  
Interacting Protein ◽  
E3 Ligases ◽  
Protein Levels ◽  
C Terminus ◽  
Osteoblast Lineage

Runx2, an essential transactivator for osteoblast differentiation, is tightly regulated at both the transcriptional and posttranslational levels. In this paper, we report that CHIP (C terminus of Hsc70-interacting protein)/STUB1 regulates Runx2 protein stability via a ubiquitination-degradation mechanism. CHIP interacts with Runx2 in vitro and in vivo. In the presence of increased Runx2 protein levels, CHIP expression decreases, whereas the expression of other E3 ligases involved in Runx2 degradation, such as Smurf1 or WWP1, remains constant or increases during osteoblast differentiation. Depletion of CHIP results in the stabilization of Runx2, enhances Runx2-mediated transcriptional activation, and promotes osteoblast differentiation in primary calvarial cells. In contrast, CHIP overexpression in preosteoblasts causes Runx2 degradation, inhibits osteoblast differentiation, and instead enhances adipogenesis. Our data suggest that negative regulation of the Runx2 protein by CHIP is critical in the commitment of precursor cells to differentiate into the osteoblast lineage.

scholarly journals Cx50 requires an intact PDZ-binding motif and ZO-1 for the formation of functional intercellular channels

2011 ◽  
Vol 22 (23) ◽  
pp. 4503-4512 ◽  
Author(s):  
Zhifang Chai ◽  
Daniel A. Goodenough ◽  
David L. Paul
Keyword(s):  
Gap Junction ◽  
Hela Cells ◽  
Pdz Domain ◽  
Cell Communication ◽  
Normal Function ◽  
Scaffolding Protein ◽  
Binding Motif ◽  
Interacting Protein ◽  
And Function

The three connexins expressed in the ocular lens each contain PDZ domain–binding motifs directing a physical association with the scaffolding protein ZO-1, but the significance of the interaction is unknown. We found that Cx50 with PDZ-binding motif mutations did not form gap junction plaques or induce cell–cell communication in HeLa cells, whereas the addition of a seven–amino acid PDZ-binding motif restored normal function to Cx50 lacking its entire C-terminal cytoplasmic domain. C-Terminal deletion had a similar although weaker effect on Cx46 but little if any effect on targeting and function of Cx43. Furthermore, small interfering RNA knockdown of ZO-1 completely inhibited the formation of gap junctions by wild-type Cx50 in HeLa cells. Thus both a PDZ-binding motif and ZO-1 are necessary for Cx50 intercellular channel formation in HeLa cells. Knock-in mice expressing Cx50 with a PDZ-binding motif mutation phenocopied Cx50 knockouts. Furthermore, differentiating lens fibers in the knock-in displayed extensive intracellular Cx50, whereas plaques in mature fibers contained only Cx46. Thus normal Cx50 function in vivo also requires an intact PDZ domain–binding motif. This is the first demonstration of a connexin-specific requirement for a connexin-interacting protein in gap junction assembly.

scholarly journals Syntenin-1 Is a New Component of Tetraspanin-Enriched Microdomains: Mechanisms and Consequences of the Interaction of Syntenin-1 with CD63

2006 ◽  
Vol 26 (20) ◽  
pp. 7707-7718 ◽  
Author(s):  
Nadya Latysheva ◽  
Gairat Muratov ◽  
Sundaresan Rajesh ◽  
Matthew Padgett ◽  
Neil A. Hotchin ◽  
...  
Keyword(s):  
Amino Acids ◽  
Plasma Membrane ◽  
Tyrosine Kinases ◽  
Pdz Domain ◽  
Binding Pocket ◽  
Resonance Spectroscopy ◽  
Nmr Studies ◽  
Transmembrane Receptors ◽  
C Terminus ◽  
Elevated Expression

ABSTRACT Tetraspanins are clustered in specific microdomains (named tetraspanin-enriched microdomains, or TERM) in the plasma membrane and regulate the functions of associated transmembrane receptors, including integrins and receptor tyrosine kinases. We have identified syntenin-1, a PDZ domain-containing protein, as a new component of TERM and show that syntenin-1 specifically interacts with the tetraspanin CD63. Detailed biochemical and heteronuclear magnetic resonance spectroscopy (NMR) studies have demonstrated that the interaction is mediated by the C-terminal cytoplasmic region of the tetraspanin and the PDZ domains of syntenin-1. Upon interaction, NMR chemical shift perturbations were predominantly localized to residues around the binding pocket of PDZ1, indicating a specific mode of recognition of the cytoplasmic tail of CD63. In addition, the C terminus of syntenin-1 has a stabilizing role in the CD63-syntenin-1 association, as deletion of the last 17 amino acids abolished the interaction. The CD63-syntenin-1 complex is abundant on the plasma membrane, and the elevated expression of the wild-type syntenin-1 slows down constitutive internalization of the tetraspanin. Furthermore, internalization of CD63 was completely blocked in cells expressing a syntenin-1 mutant lacking the first 100 amino acids. Previous results have shown that CD63 is internalized via AP-2-dependent mechanisms. Hence, our data indicate that syntenin-1 can counteract the AP-2-dependent internalization and identify this tandem PDZ protein as a new regulator of endocytosis.

scholarly journals Determination of the Interaction and Pharmacological Modulation of MCHR1 Signaling by C Terminus of MRAP2 Protein

2021 ◽  
Author(s):  
Meng Wang ◽  
Yue Zhai ◽  
Xiaowei Lei ◽  
Jing Xu ◽  
Bopei Jiang ◽  
...  
Keyword(s):  
Energy Homeostasis ◽  
Transmembrane Protein ◽  
Melanocortin Receptor ◽  
Pharmacological Modulation ◽  
C Terminus ◽  
Melanin Concentrating Hormone ◽  
The Central Nervous System

Abstract Background: Melanin concentrating hormone (MCH), an orexigenic neuropeptide, is primarily secreted by the hypothalamus and acts at its receptor, the melanin-concentrating hormone receptor 1 (MCHR1), to regulate energy homeostasis and body weight. The Melanocortin Receptor Accessory Protein 2 (MRAP2), a small single transmembrane protein broadly expressed in multiple tissues, has been defined as a vital endocrine pivot of five melanocortin receptors (MC1R-MC5R) and several other GPCRs in the regulation of central neuronal appetite and peripheral energy homeostasis. However, the regulatory and relationship between MCHR1 and MRAP2 is unknown.Results: In this study, we show that MRAP2 interacts with MCHR1 and suppresses MCHR1 signaling in vitro. We also identified the C-terminal domains of MRAP2 protein required for pharmacological modulation of intracellular Ca2+ cascades and membrane transport.Conclusions: These findings elucidated the broad regulatory profile of MRAP2 protein in the central nervous system and may provide implications for the modulation of central MCHR1 function in vivo.

scholarly journals Nuclear-localized focal adhesion kinase regulates inflammatory VCAM-1 expression

2012 ◽  
Vol 197 (7) ◽  
pp. 907-919 ◽  
Author(s):  
Ssang-Taek Lim ◽  
Nichol L.G. Miller ◽  
Xiao Lei Chen ◽  
Isabelle Tancioni ◽  
Colin T. Walsh ◽  
...  
Keyword(s):  
Focal Adhesion Kinase ◽  
Focal Adhesion ◽  
Nuclear Localization ◽  
Mitogen Activated Protein Kinase ◽  
Interacting Protein ◽  
C Terminus ◽  
Tnf Α ◽  
Mitogen Activated Protein ◽  
Factor Α

Vascular cell adhesion molecule–1 (VCAM-1) plays important roles in development and inflammation. Tumor necrosis factor–α (TNF-α) and focal adhesion kinase (FAK) are key regulators of inflammatory and integrin–matrix signaling, respectively. Integrin costimulatory signals modulate inflammatory gene expression, but the important control points between these pathways remain unresolved. We report that pharmacological FAK inhibition prevented TNF-α–induced VCAM-1 expression within heart vessel–associated endothelial cells in vivo, and genetic or pharmacological FAK inhibition blocked VCAM-1 expression during development. FAK signaling facilitated TNF-α–induced, mitogen-activated protein kinase activation, and, surprisingly, FAK inhibition resulted in the loss of the GATA4 transcription factor required for TNF-α–induced VCAM-1 production. FAK inhibition also triggered FAK nuclear localization. In the nucleus, the FAK-FERM (band 4.1, ezrin, radixin, moesin homology) domain bound directly to GATA4 and enhanced its CHIP (C terminus of Hsp70-interacting protein) E3 ligase–dependent polyubiquitination and degradation. These studies reveal new developmental and anti-inflammatory roles for kinase-inhibited FAK in limiting VCAM-1 production via nuclear localization and promotion of GATA4 turnover.

scholarly journals Determination of the Interaction and Pharmacological Modulation of MCHR1 Signaling by C Terminus of MRAP2 Protein

2021 ◽  
Author(s):  
Meng Wang ◽  
Yue Zhai ◽  
Xiaowei Lei ◽  
Jing Xu ◽  
Bopei Jiang ◽  
...  
Keyword(s):  
Energy Homeostasis ◽  
Transmembrane Protein ◽  
Melanocortin Receptor ◽  
Pharmacological Modulation ◽  
C Terminus ◽  
Melanin Concentrating Hormone ◽  
The Central Nervous System

Abstract Background: Melanin concentrating hormone (MCH), an orexigenic neuropeptide, is primarily secreted by the hypothalamus and acts at its receptor, the melanin-concentrating hormone receptor 1 (MCHR1), to regulate energy homeostasis and body weight. The Melanocortin Receptor Accessory Protein 2 (MRAP2), a small single transmembrane protein broadly expressed in multiple tissues, has been defined as a vital endocrine pivot of five melanocortin receptors (MC1R-MC5R) and several other GPCRs in the regulation of central neuronal appetite and peripheral energy homeostasis. However, the regulatory and relationship between MCHR1 and MRAP2 is unknown.Results: In this study, we show that MRAP2 interacts with MCHR1 and suppresses MCHR1 signaling in vitro. We also identified the C-terminal domains of MRAP2 protein required for pharmacological modulation of intracellular Ca2+ cascades and membrane transport. Conclusions: These findings elucidated the broad regulatory profile of MRAP2 protein in the central nervous system and may provide implications for the modulation of central MCHR1 function in vivo.

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