β-Arrestin multimers: does a crowd help or hinder function?

2008 ◽  
Vol 413 (1) ◽  
pp. e1-e3 ◽  
Author(s):  
Kathryn Anne Defea
Keyword(s):  
Specific Binding ◽  
Receptor Signalling ◽  
Binding Partners ◽  
Biochemical Journal ◽  
Extracellular Signal ◽  
Dimerization Interface ◽  
Block Association ◽  
Self Association ◽  
Subsequent Activation ◽  
G Protein Coupled

In this issue of the Biochemical Journal, Xu et al. describe how they use a spot peptide array to identify a unique sequence within β-arrestin-2 that is required for both multimerization and ERK1/2 (extracellular-signal-related kinase 1/2) scaffolding. They provide evidence that dimers may serve as more than just ‘storage forms’ of β-arrestins, incapable of interacting with receptors but, rather, perhaps, adding to the specificity of G-protein-coupled-receptor signalling. They show that key charged residues within this dimerization interface of β-arrestin-2 block association with ERK1/2 and subsequent activation of ERK1/2 by β2-adrenergic receptors, while internalization is unaffected. They suggest that self-association may serve as a means of sheltering scaffolding sites on β-arrestins from specific binding partners to prevent constitutive activation of key signalling pathways. These studies enhance our understanding of how β-arrestins can juggle their roles as scaffolds of multiple pathways in response to diverse signals.

scholarly journals Potential for imaging the high-affinity state of the 5-HT1B receptor: a comparison of three PET radioligands with differing intrinsic activity

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Anton Lindberg ◽  
Ryosuke Arakawa ◽  
Tsuyoshi Nogami ◽  
Sangram Nag ◽  
Magnus Schou ◽  
...  
Keyword(s):  
Pet Imaging ◽  
Specific Binding ◽  
Occipital Cortex ◽  
Intrinsic Activity ◽  
High Affinity ◽  
G Protein Coupled ◽  
Dose Dependent ◽  
The Brain ◽  
Agonist Affinity States ◽  
Different Doses

Abstract Background Over the last decade, a few radioligands have been developed for PET imaging of brain 5-HT1B receptors. The 5-HT1B receptor is a G-protein-coupled receptor (GPCR) that exists in two different agonist affinity states. An agonist ligand is expected to be more sensitive towards competition from another agonist, such as endogenous 5-HT, than an antagonist ligand. It is of interest to know whether the intrinsic activity of a PET radioligand for the 5-HT1B receptor impacts on its ability to detect changes in endogenous synaptic 5-HT density. Three high-affinity 11C-labeled 5-HT1B PET radioligands with differing intrinsic activity were applied to PET measurements in cynomolgus monkey to evaluate their sensitivity to be displaced within the brain by endogenous 5-HT. For these experiments, fenfluramine was pre-administered at two different doses (1.0 and 5.0 mg/kg, i.v.) to induce synaptic 5-HT release. Results A dose-dependent response to fenfluramine was detected for all three radioligands. At the highest dose of fenfluramine (5.0 mg/kg, i.v.), reductions in specific binding in the occipital cortex increased with radioligand agonist efficacy, reaching 61% for [11C]3. The most antagonistic radioligand showed the lowest reduction in specific binding. Conclusions Three 5-HT1B PET radioligands were identified with differing intrinsic activity that could be used in imaging high- and low-affinity states of 5-HT1B receptors using PET. From this limited study, radioligand sensitivity to endogenous 5-HT appears to depend on agonist efficacy. More extensive studies are required to substantiate this suggestion.

scholarly journals Identification of P-glycoprotein co-fractionating proteins and specific binding partners in rat brain microvessels

2015 ◽  
Vol 134 (2) ◽  
pp. 200-210 ◽  
Author(s):  
Margaret E. Tome ◽  
Charles P. Schaefer ◽  
Leigh M. Jacobs ◽  
Yifeng Zhang ◽  
Joseph M. Herndon ◽  
...  
Keyword(s):  
Rat Brain ◽  
Specific Binding ◽  
Binding Partners ◽  
Brain Microvessels ◽  
P Glycoprotein

scholarly journals Evidence that aggregation of mouse sperm receptors by ZP3 triggers the acrosome reaction.

1989 ◽  
Vol 108 (6) ◽  
pp. 2163-2168 ◽  
Author(s):  
L Leyton ◽  
P Saling
Keyword(s):  
Acrosome Reaction ◽  
Specific Binding ◽  
Polyclonal Antibodies ◽  
Subsequent Treatment ◽  
Mouse Sperm ◽  
Fab Fragments ◽  
Sperm Binding ◽  
Extracellular Signal ◽  
Acrosomal Exocytosis ◽  
Rabbit Igg

In the mouse, considerable evidence indicates that initial sperm binding to the zona pellucida (ZP) is mediated by ZP3. In addition, this same glycoprotein is also responsible for inducing the acrosome reaction (AR). Whereas the O-linked oligosaccharides of ZP3 appear to mediate sperm-ZP binding, the portion of ZP3 bearing AR activity has not been defined. To try to understand the bifunctional role of ZP3 (binding and AR inducing activities), we have examined the hypothesis that ZP3 aggregates sperm receptor molecules. By analogy with findings in a variety of other extracellular signal transducing systems, including receptors for growth factors and insulin, this aggregation event could initiate the cascade resulting in the AR. To test this hypothesis, we have generated monospecific polyclonal antibodies against ZP2 and against ZP3, and examined the effects of these probes on capacitated sperm incubated in the absence or presence of various ZP protein preparations. For some experiments, we have used proteolytic fragments of ZP3, a preparation known to retain specific binding, but not AR-inducing, activity. We show here that capacitated mouse sperm, incubated with ZP glycopeptides, displayed ARs when incubated subsequently with anti-ZP3 IgG; ARs did not occur when parallel sperm samples were incubated with anti-ZP2 IgG or with anti-ZP3 Fab fragments. When capacitated sperm were treated successively, with (a) ZP3 glycopeptides, (b) anti-ZP3 Fab fragments, and (c) goat anti-rabbit IgG, ARs occurred in the majority of sperm. An alternative approach to examine this hypothesis used ZP proteins obtained from tubal eggs treated previously with bioactive phorbol diester (12-O-tetradecanoyl phorbol-13-acetate [TPA]). This preparation arrests capacitated sperm in an intermediate state of the AR. We demonstrate here that these sperm can be induced to undergo a complete AR by subsequent treatment with anti-ZP3 IgG. Together, these findings are consistent with the hypothesis under examination, and suggest that the aggregation of sperm molecules recognized by ZP3 glycopeptides or by TPA-treated ZP is sufficient to trigger the events that occur during acrosomal exocytosis.

scholarly journals cAMP-stimulated phosphorylation of diaphanous 1 regulates protein stability and interaction with binding partners in adrenocortical cells

2013 ◽  
Vol 24 (6) ◽  
pp. 848-857 ◽  
Author(s):  
Donghui Li ◽  
Eric B. Dammer ◽  
Natasha C. Lucki ◽  
Marion B. Sewer
Keyword(s):  
Oxysterol Binding Protein ◽  
Mitochondrial Movement ◽  
Binding Partners ◽  
Extracellular Signal ◽  
Camp Signaling Pathway ◽  
Integral Role ◽  
Mitochondrial Trafficking ◽  
The Stability ◽  
And Function ◽  
Β Tubulin

Diaphanous homologue 1 (DIAPH1) is a Rho effector protein that coordinates cellular dynamics by regulating microfilament and microtubule function. We previously showed that DIAPH1 plays an integral role in regulating the production of cortisol by controlling the rate of mitochondrial movement, by which activation of the adrenocorticotropin (ACTH)/cAMP signaling pathway stimulates mitochondrial trafficking and promotes the interaction between RhoA and DIAPH1. In the present study we use mass spectrometry to identify DIAPH1 binding partners and find that DIAPH1 interacts with several proteins, including RhoA, dynamin-1, kinesin, β-tubulin, β-actin, oxysterol-binding protein (OSBP)–related protein 2 (ORP2), and ORP10. Moreover, DIAPH1 is phosphorylated in response to dibutyryl cAMP (Bt2cAMP) at Thr-759 via a pathway that requires extracellular signal-related kinase (ERK). Alanine substitution of Thr-759 renders DIAPH1 more stable and attenuates the interaction between DIAPH1 and kinesin, ORP2, and actin but has no effect on the ability of the protein to interact with RhoA or β-tubulin. Finally, overexpression of a DIAPH1 T759A mutant significantly decreases the rate of Bt2cAMP-stimulated mitochondrial movement. Taken together, our findings establish a key role for phosphorylation in regulating the stability and function of DIAPH1.

scholarly journals G protein-coupled receptor signalling in the cardiac nuclear membrane: evidence and possible roles in physiological and pathophysiological function

2012 ◽  
Vol 590 (6) ◽  
pp. 1313-1330 ◽  
Author(s):  
Artavazd Tadevosyan ◽  
George Vaniotis ◽  
Bruce G. Allen ◽  
Terence E. Hébert ◽  
Stanley Nattel
Keyword(s):  
G Protein ◽  
Nuclear Membrane ◽  
G Protein Coupled Receptor ◽  
Receptor Signalling ◽  
G Protein Coupled

scholarly journals Structure of the Mouse Sex Peptide Pheromone ESP1 Reveals a Molecular Basis for Specific Binding to the Class C G-protein-coupled Vomeronasal Receptor

2013 ◽  
Vol 288 (22) ◽  
pp. 16064-16072 ◽  
Author(s):  
Sosuke Yoshinaga ◽  
Toru Sato ◽  
Makoto Hirakane ◽  
Kaori Esaki ◽  
Takashi Hamaguchi ◽  
...  
Keyword(s):  
G Protein ◽  
Molecular Basis ◽  
Specific Binding ◽  
Sex Peptide ◽  
Vomeronasal Receptor ◽  
Peptide Pheromone ◽  
Class C ◽  
G Protein Coupled

scholarly journals Unlabeled lysophosphatidic acid receptor binding in free solution as determined by a compensated interferometric reader

2020 ◽  
Vol 61 (8) ◽  
pp. 1244-1251 ◽  
Author(s):  
Manisha Ray ◽  
Kazufumi Nagai ◽  
Yasuyuki Kihara ◽  
Amanda Kussrow ◽  
Michael N. Kammer ◽  
...  
Keyword(s):  
Receptor Binding ◽  
Lysophosphatidic Acid ◽  
Specific Binding ◽  
Test Case ◽  
Free Solution ◽  
Label Free ◽  
Adhesive Properties ◽  
Lpa Receptor ◽  
System A ◽  
G Protein Coupled

Native interactions between lysophospholipids (LPs) and their cognate LP receptors are difficult to measure because of lipophilicity and/or the adhesive properties of lipids, which contribute to high levels of nonspecific binding in cell membrane preparations. Here, we report development of a free-solution assay (FSA) where label-free LPs bind to their cognate G protein-coupled receptors (GPCRs), combined with a recently reported compensated interferometric reader (CIR) to quantify native binding interactions between receptors and ligands. As a test case, the binding parameters between lysophosphatidic acid (LPA) receptor 1 (LPA1; one of six cognate LPA GPCRs) and LPA were determined. FSA-CIR detected specific binding through the simultaneous real-time comparison of bound versus unbound species by measuring the change in the solution dipole moment produced by binding-induced conformational and/or hydration changes. FSA-CIR identified KD values for chemically distinct LPA species binding to human LPA1 and required only a few nanograms of protein: 1-oleoyl (18:1; KD = 2.08 ± 1.32 nM), 1-linoleoyl (18:2; KD = 2.83 ± 1.64 nM), 1-arachidonoyl (20:4; KD = 2.59 ± 0.481 nM), and 1-palmitoyl (16:0; KD = 1.69 ± 0.1 nM) LPA. These KD values compared favorably to those obtained using the previous generation back-scattering interferometry system, a chip-based technique with low-throughput and temperature sensitivity. In conclusion, FSA-CIR offers a new increased-throughput approach to assess quantitatively label-free lipid ligand-receptor binding, including nonactivating antagonist binding, under near-native conditions.

scholarly journals An arrestin-1 surface opposite of its interface with photoactivated rhodopsin engages with enolase-1

2020 ◽  
Vol 295 (19) ◽  
pp. 6498-6508
Author(s):  
Connie Jaqueline Miranda ◽  
Nicole Fernandez ◽  
Nader Kamel ◽  
Daniel Turner ◽  
Del Benzenhafer ◽  
...  
Keyword(s):  
Amino Acids ◽  
Molecular Model ◽  
Signaling Cascades ◽  
Contact Sites ◽  
Binding Partners ◽  
Protein Partners ◽  
Fluorescence Quench ◽  
Selective Substitution ◽  
Additional Protein ◽  
G Protein Coupled

Arrestin-1 is the arrestin family member responsible for inactivation of the G protein–coupled receptor rhodopsin in photoreceptors. Arrestin-1 is also well-known to interact with additional protein partners and to affect other signaling cascades beyond phototransduction. In this study, we investigated one of these alternative arrestin-1 binding partners, the glycolysis enzyme enolase-1, to map the molecular contact sites between these two proteins and investigate how the binding of arrestin-1 affects the catalytic activity of enolase-1. Using fluorescence quench protection of strategically placed fluorophores on the arrestin-1 surface, we observed that arrestin-1 primarily engages enolase-1 along a surface that is opposite of the side of arrestin-1 that binds photoactivated rhodopsin. Using this information, we developed a molecular model of the arrestin-1–enolase-1 complex, which was validated by targeted substitutions of charge-pair interactions. Finally, we identified the likely source of arrestin's modulation of enolase-1 catalysis, showing that selective substitution of two amino acids in arrestin-1 can completely remove its effect on enolase-1 activity while still remaining bound to enolase-1. These findings open up opportunities for examining the functional effects of arrestin-1 on enolase-1 activity in photoreceptors and their surrounding cells.

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