scholarly journals Evaluating Cellular Impedance Assays for Detection of GPCR Pleiotropic Signaling and Functional Selectivity

2009 ◽  
Vol 14 (3) ◽  
pp. 246-255 ◽  
Author(s):  
Matthew F. Peters ◽  
Clay W. Scott
Keyword(s):  
Cell Types ◽  
D1 Receptor ◽  
G Protein Coupled Receptors ◽  
Functional Selectivity ◽  
Dynamic Impedance ◽  
Melanocortin 4 Receptor ◽  
Impedance Profile ◽  
Different Cell Types ◽  
G Protein Coupled ◽  
Receptor Conformation

G-protein—coupled receptors can couple to different signal transduction pathways in different cell types (termed cell-specific signaling) and can activate different signaling pathways depending on the receptor conformation(s) stabilized by the activating ligand (functional selectivity). These concepts offer potential for developing pathway-specific drugs that increase efficacy and reduce side effects. Despite significant interest, functional selectivity has been difficult to exploit in drug discovery, in part due to the burden of multiple assays. Cellular impedance assays use an emerging technology that can qualitatively distinguish Gs, Gi/o, and Gq signaling in a single assay and is thereby suited for studying these pharmacological concepts. Cellular impedance confirmed cell-specific Gs and Gq coupling for the melanocortin-4 receptor and dual Gi and Gs signaling with the cannabinoid-1 (CB1) receptor. The balance of Gi versus Gs signaling depended on the cell line. In CB1-HEKs, Giand Gs-like responses combined to yield a novel impedance profile demonstrating the dynamic nature of these traces. Cellspecific signaling was observed with endogenous D1 receptor in U-2 cells and SK-N-MC cells, yet the pharmacological profile of partial and full agonists was similar in both cell lines. We conclude that the dynamic impedance profile encodes valuable relative signaling information and is sufficiently robust to help evaluate cell-specific signaling and functional selectivity. ( Journal of Biomolecular Screening 2009:246-255)

scholarly journals The Biology of Vasopressin

Biomedicines ◽  
2021 ◽  
Vol 9 (1) ◽  
pp. 89
Author(s):  
Samantha Sparapani ◽  
Cassandra Millet-Boureima ◽  
Joshua Oliver ◽  
Kathy Mu ◽  
Pegah Hadavi ◽  
...  
Keyword(s):  
Parental Behavior ◽  
Cell Types ◽  
G Protein Coupled Receptors ◽  
Receptor Expression ◽  
Evolutionarily Conserved ◽  
Terrestrial Environments ◽  
Intracellular Vesicles ◽  
G Protein Coupled ◽  
Neuropsychiatric Conditions ◽  
Vasopressin Synthesis

Vasopressins are evolutionarily conserved peptide hormones. Mammalian vasopressin functions systemically as an antidiuretic and regulator of blood and cardiac flow essential for adapting to terrestrial environments. Moreover, vasopressin acts centrally as a neurohormone involved in social and parental behavior and stress response. Vasopressin synthesis in several cell types, storage in intracellular vesicles, and release in response to physiological stimuli are highly regulated and mediated by three distinct G protein coupled receptors. Other receptors may bind or cross-bind vasopressin. Vasopressin is regulated spatially and temporally through transcriptional and post-transcriptional mechanisms, sex, tissue, and cell-specific receptor expression. Anomalies of vasopressin signaling have been observed in polycystic kidney disease, chronic heart failure, and neuropsychiatric conditions. Growing knowledge of the central biological roles of vasopressin has enabled pharmacological advances to treat these conditions by targeting defective systemic or central pathways utilizing specific agonists and antagonists.

scholarly journals A Minimal Model for G Protein–Mediated Synaptic Facilitation and Depression

2003 ◽  
Vol 90 (3) ◽  
pp. 1643-1653 ◽  
Author(s):  
Richard Bertram ◽  
Jessica Swanson ◽  
Mohammad Yousef ◽  
Zhong-Ping Feng ◽  
Gerald W. Zamponi
Keyword(s):  
G Protein ◽  
Cell Types ◽  
G Protein Coupled Receptors ◽  
Β Subunit ◽  
Short Term ◽  
Paired Pulse ◽  
Synaptic Facilitation ◽  
Protein Inhibition ◽  
G Protein Coupled ◽  
High Pass

G protein–coupled receptors are ubiquitous in neurons, as well as other cell types. Activation of receptors by hormones or neurotransmitters splits the G protein heterotrimer into Gα and Gβγ subunits. It is now clear that Gβγ directly inhibits Ca2+ channels, putting them into a reluctant state. The effects of Gβγ depend on the specific β and γ subunits present, as well as the β subunit isoform of the N-type Ca2+ channel. We describe a minimal mathematical model for the effects of G protein action on the dynamics of synaptic transmission. The model is calibrated by data obtained by transfecting G protein and Ca2+ channel subunits into tsA-201 cells. We demonstrate with numerical simulations that G protein action can provide a mechanism for either short-term synaptic facilitation or depression, depending on the manner in which G protein–coupled receptors are activated. The G protein action performs high-pass filtering of the presynaptic signal, with a filter cutoff that depends on the combination of G protein and Ca2+ channel subunits present. At stimulus frequencies above the cutoff, trains of single spikes are transmitted, while only doublets are transmitted at frequencies below the cutoff. Finally, we demonstrate that relief of G protein inhibition can contribute to paired-pulse facilitation.

Compartmentalization of GPCR signalling controls unique cellular responses

2016 ◽  
Vol 44 (2) ◽  
pp. 562-567 ◽  
Author(s):  
Andrew M. Ellisdon ◽  
Michelle L. Halls
Keyword(s):  
Drug Discovery ◽  
Drug Targets ◽  
Cell Types ◽  
G Protein Coupled Receptors ◽  
Attrition Rates ◽  
Physiological Processes ◽  
Simple Chain ◽  
G Protein Coupled ◽  
Major Drug ◽  
Very High

With >800 members, G protein-coupled receptors (GPCRs) are the largest class of cell-surface signalling proteins, and their activation mediates diverse physiological processes. GPCRs are ubiquitously distributed across all cell types, involved in many diseases and are major drug targets. However, GPCR drug discovery is still characterized by very high attrition rates. New avenues for GPCR drug discovery may be provided by a recent shift away from the traditional view of signal transduction as a simple chain of events initiated from the plasma membrane. It is now apparent that GPCR signalling is restricted to highly organized compartments within the cell, and that GPCRs activate distinct signalling pathways once internalized. A high-resolution understanding of how compartmentalized signalling is controlled will probably provide unique opportunities to selectively and therapeutically target GPCRs.

scholarly journals The Biology of Prostaglandins and Their Role as a Target for Allergic Airway Disease Therapy

2020 ◽  
Vol 21 (5) ◽  
pp. 1851 ◽  
Author(s):  
Kijeong Lee ◽  
Sang Hag Lee ◽  
Tae Hoon Kim
Keyword(s):  
Nasal Polyposis ◽  
Allergic Inflammation ◽  
Cell Types ◽  
Airway Disease ◽  
G Protein Coupled Receptors ◽  
Allergic Airway Disease ◽  
Airway Diseases ◽  
Disease Therapy ◽  
Lipid Compounds ◽  
G Protein Coupled

Prostaglandins (PGs) are a family of lipid compounds that are derived from arachidonic acid via the cyclooxygenase pathway, and consist of PGD2, PGI2, PGE2, PGF2, and thromboxane B2. PGs signal through G-protein coupled receptors, and individual PGs affect allergic inflammation through different mechanisms according to the receptors with which they are associated. In this review article, we have focused on the metabolism of the cyclooxygenase pathway, and the distinct biological effect of each PG type on various cell types involved in allergic airway diseases, including asthma, allergic rhinitis, nasal polyposis, and aspirin-exacerbated respiratory disease.

scholarly journals Thrombin Ca2+-dependently stimulates protein tyrosine phosphorylation in BC3H1 muscle cells

1993 ◽  
Vol 290 (1) ◽  
pp. 27-32 ◽  
Author(s):  
S Offermanns ◽  
E Bombien ◽  
G Schultz
Keyword(s):  
Growth Factor ◽  
Tyrosine Phosphorylation ◽  
Cellular Response ◽  
Cell Types ◽  
Ionophore A23187 ◽  
Mouse Muscle ◽  
Kinase C ◽  
Epidermal Growth ◽  
Different Cell Types ◽  
G Protein Coupled

The proteinase thrombin, known to act via heptahelical G-protein-coupled receptors, is a mitogenic agent for different cell types, including the mouse muscle cell line BC3H1. In this study, the effect of thrombin on tyrosine phosphorylation was examined using anti-phosphotyrosine antibodies. Thrombin was found to induce phosphorylation of 65-70 and 110-120 kDa proteins in BC3H1 cells. The effect of thrombin was concentration-dependent, being half-maximal and maximal at concentrations of 0.03 and 1 unit/ml respectively. The thrombin-induced increase in phosphorylation was rapid (< or = 10 s) and transient, with a peak response after about 1-2 min. The effect of thrombin could be mimicked by the thrombin receptor agonist peptide SFLLRN-NH2. Preincubation of cells with pertussis toxin (PT) had no effect on thrombin-induced tyrosine phosphorylation. Epidermal growth factor, platelet-derived growth factor and insulin stimulated tyrosine phosphorylation of different proteins, among which were 65-70 and 110-120 kDa proteins. The phorbol ester 12-myristate 13-acetate (PMA) as well as the Ca2+ ionophore A23187 both stimulated tyrosine phosphorylation of proteins identical to those phosphorylated by thrombin, suggesting that activation of protein kinase C (PKC) and elevation of the cytosolic Ca2+ concentration alone are sufficient to induce tyrosine phosphorylation. However, calphostin C and other PKC inhibitors, which completely inhibited tyrosine phosphorylation induced by PMA, had no influence on the effect of thrombin, whereas loading of cells with the intracellular Ca2+ chelator bis-(O-aminophenoxy)ethane-NNN'N'-tetra-acetic acid totally blocked thrombin-stimulated tyrosine phosphorylation. Thus tyrosine phosphorylation stimulated by thrombin is an early PT-insensitive cellular response which is either directly mediated by elevation of cytosolic Ca2+ concentration or by a presently unknown mechanism that requires an elevated cytosolic Ca2+ concentration.

scholarly journals Lysophospholipids in the limelight

2002 ◽  
Vol 158 (2) ◽  
pp. 197-199 ◽  
Author(s):  
Wouter H. Moolenaar
Keyword(s):  
Growth Factor ◽  
Cell Motility ◽  
Tumor Progression ◽  
G Protein ◽  
Lysophosphatidic Acid ◽  
Cell Types ◽  
G Protein Coupled Receptors ◽  
Serum Phospholipid ◽  
A Cell ◽  
G Protein Coupled

Lysophosphatidic acid (LPA) is a serum phospholipid that evokes growth factor–like responses in many cell types through the activation of its G protein–coupled receptors. Although much is known about LPA signaling, it has remained unclear where and how bioactive LPA is produced. Umezu-Goto et al. (2002)(this issue, page 227) have purified a serum lysophospholipase D that generates LPA from lysophosphatidylcholine and found it to be identical to autotaxin, a cell motility–stimulating ectophosphodiesterase implicated in tumor progression. This result is surprising, as there was previously no indication that autotaxin could act as a phospholipase.

How ligands and signalling proteins affect G-protein-coupled receptors' conformational landscape

2013 ◽  
Vol 41 (1) ◽  
pp. 144-147 ◽  
Author(s):  
Sophie Mary ◽  
Jean-Alain Fehrentz ◽  
Marjorie Damian ◽  
Pascal Verdié ◽  
Jean Martinez ◽  
...  
Keyword(s):  
G Protein ◽  
G Protein Coupled Receptors ◽  
Effector Proteins ◽  
New Drugs ◽  
Signalling Pathways ◽  
Ghrelin Receptor ◽  
Multiple Data ◽  
Conformational Landscape ◽  
G Protein Coupled ◽  
Receptor Conformation

The dynamic character of GPCRs (G-protein-coupled receptors) is essential to their function. However, the details of how ligands and signalling proteins stabilize a receptor conformation to trigger the activation of a given signalling pathway remain largely unexplored. Multiple data, including recent results obtained with the purified ghrelin receptor, suggest a model where ligand efficacy and functional selectivity are directly related to different receptor conformations. Importantly, distinct effector proteins (G-proteins and arrestins) as well as ligands are likely to affect the conformational landscape of GPCRs in different manners, as we show with the isolated ghrelin receptor. Such modulation of the GPCR conformational landscape by pharmacologically distinct ligands and effector proteins has major implications for the design of new drugs that activate specific signalling pathways.

scholarly journals Structural Basis of Arrestin Selectivity for Active Phosphorylated G Protein-Coupled Receptors

2021 ◽  
Vol 22 (22) ◽  
pp. 12481
Author(s):  
Preethi C. Karnam ◽  
Sergey A. Vishnivetskiy ◽  
Vsevolod V. Gurevich
Keyword(s):  
G Protein ◽  
Receptor Binding ◽  
G Protein Coupled Receptors ◽  
Structural Basis ◽  
Preferential Binding ◽  
High Affinity Receptor ◽  
Functional Forms ◽  
Affinity Receptor ◽  
G Protein Coupled ◽  
Receptor Conformation

Arrestins are a small family of proteins that bind G protein-coupled receptors (GPCRs). Arrestin binds to active phosphorylated GPCRs with higher affinity than to all other functional forms of the receptor, including inactive phosphorylated and active unphosphorylated. The selectivity of arrestins suggests that they must have two sensors, which detect receptor-attached phosphates and the active receptor conformation independently. Simultaneous engagement of both sensors enables arrestin transition into a high-affinity receptor-binding state. This transition involves a global conformational rearrangement that brings additional elements of the arrestin molecule, including the middle loop, in contact with a GPCR, thereby stabilizing the complex. Here, we review structural and mutagenesis data that identify these two sensors and additional receptor-binding elements within the arrestin molecule. While most data were obtained with the arrestin-1-rhodopsin pair, the evidence suggests that all arrestins use similar mechanisms to achieve preferential binding to active phosphorylated GPCRs.

scholarly journals cAMP Signaling Regulates DNA Demethylation by Augmenting the Intracellular Labile Ferrous Iron Pool

2017 ◽  
Author(s):  
Vladimir Camarena ◽  
David W. Sant ◽  
Tyler C. Huff ◽  
Sushmita Mustafi ◽  
Ryan K. Muir ◽  
...  
Keyword(s):  
Gene Transcription ◽  
Phosphodiesterase Inhibitors ◽  
Cell Types ◽  
Dna Demethylation ◽  
G Protein Coupled Receptors ◽  
Camp Signaling ◽  
Multiple Cell ◽  
Iron Pool ◽  
Transcriptomic Changes ◽  
G Protein Coupled

AbstractIt is widely accepted that cAMP regulates gene transcription principally by activating the protein kinase A (PKA)-targeted transcription factors. Here, we show that cAMP enhances the generation of 5-hydroxymethylcytosine (5hmC) in multiple cell types. 5hmC is converted from 5-methylcytosine (5mC) by Tet methylcytosine dioxygenases, for which Fe(II) is an essential cofactor. The promotion of 5hmC was mediated by a prompt increase of the intracellular labile Fe(II) pool (LIP). cAMP enhanced the acidification of endosomes for Fe(II) release to the LIP likely through RapGEF2. The effect of cAMP on Fe(II) and 5hmC was confirmed by adenylate cyclase activators, phosphodiesterase inhibitors, and most notably by stimulation of G protein-coupled receptors (GPCR). The transcriptomic changes caused by cAMP occurred in concert with 5hmC elevation in differentially transcribed genes. Collectively, these data show a previously unrecognized regulation of gene transcription by GPCR-cAMP signaling through augmentation of the intracellular labile Fe(II) pool and DNA demethylation.

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