scholarly journals Lysophosphatidylcholine-induced Surface Redistribution Regulates Signaling of the Murine G Protein-coupled Receptor G2A

2005 ◽  
Vol 16 (5) ◽  
pp. 2234-2247 ◽  
Author(s):  
Li Wang ◽  
Caius G. Radu ◽  
Li V. Yang ◽  
Laurent A. Bentolila ◽  
Mireille Riedinger ◽  
...  
Keyword(s):  
G Protein ◽  
Fluorescent Protein ◽  
Spatial Dynamics ◽  
Surface Expression ◽  
Subcellular Fractionation ◽  
Membrane Receptors ◽  
G Protein Coupled Receptor ◽  
Erk Activation ◽  
Fractionation Analysis ◽  
G Protein Coupled

Intracellular trafficking and spatial dynamics of membrane receptors critically regulate receptor function. Using microscopic and subcellular fractionation analysis, we studied the localization of the murine G protein-coupled receptor G2A (muG2A). Evaluating green fluorescent protein-tagged, exogenously expressed as well as the endogenous muG2A, we observed that this receptor was spontaneously internalized and accumulated in endosomal compartments, whereas its surface expression was enhanced and stabilized by lysophosphatidylcholine (LPC) treatment. Monensin, a general inhibitor of recycling pathways, blocked LPC-regulated surface localization of muG2A as well as muG2A-dependent extracellular signal-regulated kinase (ERK) activation and cell migration induced by LPC treatment. Mutation of the conserved DRY motif (R→ A) enhanced the surface expression of muG2A, resulting in its resistance to monensin inhibition of ERK activation. Our data suggest that intracellular sequestration and surface expression regulated by LPC, rather than direct agonistic activity control the signaling responses of murine G2A toward LPC.

A generic microfluidic biosensor of G protein-coupled receptor activation – impedance measurements of reversible morphological changes of reverse transfected HEK293 cells on microelectrodes

RSC Advances ◽  
2015 ◽  
Vol 5 (65) ◽  
pp. 52563-52570 ◽  
Author(s):  
Saurabh K. Srivastava ◽  
Rajesh Ramaneti ◽  
Margriet Roelse ◽  
Hien Duy Tong ◽  
Elwin X. Vrouwe ◽  
...  
Keyword(s):  
G Protein ◽  
Morphological Changes ◽  
Receptor Activation ◽  
Membrane Receptors ◽  
Hek293 Cells ◽  
G Protein Coupled Receptor ◽  
Impedance Measurements ◽  
Transfected Cells ◽  
Microfluidic Biosensor ◽  
G Protein Coupled

Flowcell with micro-IDEs (250–500 μm) covered with both stable and reverse transfected cells overexpressing membrane receptors to demonstrate impedance responses to serial injections of analyte.

scholarly journals Genetic Variations in the Human G Protein-coupled Receptor Class C, Group 6, Member A (GPRC6A) Control Cell Surface Expression and Function

2016 ◽  
Vol 292 (4) ◽  
pp. 1524-1534 ◽  
Author(s):  
Stine Jørgensen ◽  
Christian Theil Have ◽  
Christina Rye Underwood ◽  
Lars Dan Johansen ◽  
Petrine Wellendorph ◽  
...  
Keyword(s):  
Cell Surface ◽  
G Protein ◽  
Control Cell ◽  
Surface Expression ◽  
Genetic Variations ◽  
Cell Surface Expression ◽  
G Protein Coupled Receptor ◽  
Group 6 ◽  
And Function ◽  
G Protein Coupled

scholarly journals G protein–coupled receptor/arrestin3 modulation of the endocytic machinery

2002 ◽  
Vol 156 (4) ◽  
pp. 665-676 ◽  
Author(s):  
Francesca Santini ◽  
Ibragim Gaidarov ◽  
James H. Keen
Keyword(s):  
G Protein ◽  
Fluorescent Protein ◽  
De Novo ◽  
Membrane Skeleton ◽  
Live Cells ◽  
G Protein Coupled Receptor ◽  
Cortical Actin ◽  
Endocytic Machinery ◽  
Green Fluorescent ◽  
G Protein Coupled

Nonvisual arrestins (arr) modulate G protein–coupled receptor (GPCR) desensitization and internalization and bind to both clathrin (CL) and AP-2 components of the endocytic coated pit (CP). This raises the possibility that endocytosis of some GPCRs may be a consequence of arr-induced de novo CP formation. To directly test this hypothesis, we examined the behavior of green fluorescent protein (GFP)-arr3 in live cells expressing β2-adrenergic receptors and fluorescent CL. After agonist stimulation, the diffuse GFP-arr3 signal rapidly became punctate and colocalized virtually completely with preexisting CP spots, demonstrating that activated complexes accumulate in previously formed CPs rather than nucleating new CP formation. After arr3 recruitment, CP appeared larger: electron microscopy analysis revealed an increase in both CP number and in the occurrence of clustered CPs. Mutant arr3 proteins with impaired binding to CL or AP-2 displayed reduced recruitment to CPs, but were still capable of inducing CP clustering. In contrast, though constitutively present in CPs, the COOH-terminal moiety of arr3, which contains CP binding sites but lacks receptor binding, did not induce CP clustering. Together, these results indicate that recruitment of functional arr3–GPCR complexes to CP is necessary to induce clustering. Latrunculin B or 16°C blocked CP rearrangements without affecting arr3 recruitment to CP. These results and earlier studies suggest that discrete CP zones exist on cell surfaces, each capable of supporting adjacent CPs, and that the cortical actin membrane skeleton is intimately involved with both the maintenance of existing CPs and the generation of new structures.

scholarly journals Serine 129 phosphorylation of membrane-associated α-synuclein modulates dopamine transporter function in a G protein–coupled receptor kinase–dependent manner

2013 ◽  
Vol 24 (11) ◽  
pp. 1649-1660 ◽  
Author(s):  
Susumu Hara ◽  
Shigeki Arawaka ◽  
Hiroyasu Sato ◽  
Youhei Machiya ◽  
Can Cui ◽  
...  
Keyword(s):  
Cell Surface ◽  
G Protein ◽  
Dopamine Transporter ◽  
Surface Expression ◽  
Hek293 Cells ◽  
Cell Surface Expression ◽  
Receptor Kinase ◽  
Wild Type ◽  
G Protein Coupled Receptor ◽  
G Protein Coupled

Most α-synuclein (α-syn) deposited in Lewy bodies, the pathological hallmark of Parkinson disease (PD), is phosphorylated at Ser-129. However, the physiological and pathological roles of this modification are unclear. Here we investigate the effects of Ser-129 phosphorylation on dopamine (DA) uptake in dopaminergic SH-SY5Y cells expressing α-syn. Subcellular fractionation of small interfering RNA (siRNA)–treated cells shows that G protein–coupled receptor kinase 3 (GRK3), GRK5, GRK6, and casein kinase 2 (CK2) contribute to Ser-129 phosphorylation of membrane-associated α-syn, whereas cytosolic α-syn is phosphorylated exclusively by CK2. Expression of wild-type α-syn increases DA uptake, and this effect is diminished by introducing the S129A mutation into α-syn. However, wild-type and S129A α-syn equally increase the cell surface expression of dopamine transporter (DAT) in SH-SY5Y cells and nonneuronal HEK293 cells. In addition, siRNA-mediated knockdown of GRK5 or GRK6 significantly attenuates DA uptake without altering DAT cell surface expression, whereas knockdown of CK2 has no effect on uptake. Taken together, our results demonstrate that membrane-associated α-syn enhances DA uptake capacity of DAT by GRKs-mediated Ser-129 phosphorylation, suggesting that α-syn modulates intracellular DA levels with no functional redundancy in Ser-129 phosphorylation between GRKs and CK2.

scholarly journals Rapid internalization and surface expression of a functional, fluorescently tagged G-protein-coupled glutamate receptor

1999 ◽  
Vol 341 (2) ◽  
pp. 415-422 ◽  
Author(s):  
Andrew J. DOHERTY ◽  
Victoria COUTINHO ◽  
Graham L. COLLINGRIDGE ◽  
Jeremy M. HENLEY
Keyword(s):  
G Protein ◽  
Functional Response ◽  
Fluorescent Protein ◽  
Single Cells ◽  
Surface Expression ◽  
Receptor Subtype ◽  
Functional Assay ◽  
Hek 293 ◽  
Excitatory Neurotransmitter ◽  
G Protein Coupled

L-Glutamate is the principal excitatory neurotransmitter in the vertebrate central nervous system, where it mediates many of its actions via G-protein-coupled metabotropic glutamate (mGlu) receptors. Since little is known about the dynamics of mGlu receptors at the plasma membrane, we have constructed a fusion protein comprising the mGlu receptor subtype 1α (mGlu1α) and green fluorescent protein (GFP). Using imaging of Ca2+ release from intracellular stores as a functional assay, the agonist pharmacology of this fluorescently tagged receptor was found to be similar to that of the wild-type receptor when expressed in HEK-293 cells. Receptor movement and function were measured simultaneously by combined imaging of Ca2+, using fura-red, and GFP fluorescence in single cells. Exposure to agonist induced a rapid loss of up to 30% of membrane-associated fluorescence, with a corresponding decrease in the functional response. Following removal of the agonist there was recovery of both the membrane fluorescence and the functional response. These data suggest that the surface expression of G-protein-coupled glutamate receptors might be rapidly regulated in response to agonist activation.

scholarly journals RACK1 Regulates the Cell Surface Expression of the G Protein-Coupled Receptor for Thromboxane A2

Traffic ◽  
2008 ◽  
Vol 9 (3) ◽  
pp. 394-407 ◽  
Author(s):  
Audrey Parent ◽  
Geneviève Laroche ◽  
Émilie Hamelin ◽  
Jean-Luc Parent
Keyword(s):  
Cell Surface ◽  
G Protein ◽  
Thromboxane A2 ◽  
Surface Expression ◽  
Cell Surface Expression ◽  
G Protein Coupled Receptor ◽  
G Protein Coupled
Sign in / Sign up

Export Citation Format

Share Document