scholarly journals One gene - two proteins: The C-terminus of the prototypical M2 muscarinic receptor localizes to the mitochondria

2021 ◽  
Author(s):  
Francesco Petragnano ◽  
Irene Fasciani ◽  
Ziming Wang ◽  
Ruairidh Edwards ◽  
Narasimha Telugu ◽  
...  
Keyword(s):  
G Protein ◽  
Acetylcholine Receptors ◽  
Muscarinic Acetylcholine Receptors ◽  
Entry Site ◽  
Internal Ribosome Entry ◽  
C Terminus ◽  
M2 Muscarinic Receptor ◽  
Carboxyl Terminal ◽  
Transmembrane Regions ◽  
G Protein Coupled

Muscarinic acetylcholine receptors are prototypical G protein-coupled receptors activated by the endogenous neurotransmitter acetylcholine. We show here that the carboxyl terminal fragment of the muscarinic M2 receptor, containing the transmembrane regions VI and VII (M2tail), is expressed by virtue of an internal ribosome entry site. The M2tail fragment, whose expression is upregulated in cells undergoing integrated stress, response, does not follow the normal route to the plasma membrane, but is almost exclusively sorted to mitochondria: here it controls oxygen consumption, cell proliferation and the formation of reactive oxygen species via reduction of oxidative phosphorylation. The expression of the carboxyl-terminal of a G protein-coupled receptor, capable of regulating mitochondrial function, constitutes a hitherto unknown mechanism that cells may use for controlling their metabolism under variable environmental conditions.

scholarly journals Molecular basis of signaling specificity between GIRK channels and GPCRs

eLife ◽  
2018 ◽  
Vol 7 ◽  
Author(s):  
Kouki K Touhara ◽  
Roderick MacKinnon
Keyword(s):  
G Protein ◽  
Acetylcholine Receptors ◽  
Resonance Energy Transfer ◽  
Resonance Energy ◽  
Muscarinic Acetylcholine Receptors ◽  
K Channel ◽  
Girk Channels ◽  
Signaling Specificity ◽  
Slow Heart Rate ◽  
G Protein Coupled

Stimulated muscarinic acetylcholine receptors (M2Rs) release Gβγ subunits, which slow heart rate by activating a G protein-gated K+ channel (GIRK). Stimulated β2 adrenergic receptors (β2ARs) also release Gβγ subunits, but GIRK is not activated. This study addresses the mechanism underlying this specificity of GIRK activation by M2Rs. K+ currents and bioluminescence resonance energy transfer between labelled G proteins and GIRK show that M2Rs catalyze Gβγ subunit release at higher rates than β2ARs, generating higher Gβγ concentrations that activate GIRK and regulate other targets of Gβγ. The higher rate of Gβγ release is attributable to a faster G protein coupled receptor – G protein trimer association rate in M2R compared to β2AR. Thus, a rate difference in a single kinetic step accounts for specificity.

scholarly journals Development of a Homogeneous High-Throughput Live-Cell G-Protein-Coupled Receptor Binding Assay

2008 ◽  
Vol 13 (8) ◽  
pp. 748-754 ◽  
Author(s):  
Paul H. Lee ◽  
Steven C. Miller ◽  
Carlo van Staden ◽  
Evan F. Cromwell
Keyword(s):  
G Protein ◽  
Receptor Binding ◽  
Acetylcholine Receptors ◽  
Binding Assay ◽  
Chinese Hamster Ovary Cells ◽  
Scale Up ◽  
Chinese Hamster ◽  
Live Cell ◽  
Muscarinic Acetylcholine Receptors ◽  
G Protein Coupled

The measurement of ligand receptor binding parameters for G-protein-coupled receptors is indispensable in the drug discovery process. Traditional ligand receptor binding assays require scale-up of cells and membrane preparations, which is an expensive and time-consuming process. In this report, the authors describe the development of a homogeneous live-cell binding assay for GPCRs using a fluorophore-labeled nonpeptide ligand. The model assay used Cy3B-labeled telenzepine and Chinese hamster ovary cells expressing M1 muscarinic acetylcholine receptors. This homogeneous live-cell fluorescence binding assay format is superior to the traditional binding methods because it measures binding of a ligand to intact receptors on living cells. The assay requires no washing or separation steps, thereby allowing a real-time kinetic readout for the determination of ligand association and dissociation from the intact receptors. The results also suggest that miniaturization is feasible without compromising the data quality. ( Journal of Biomolecular Screening 2008:748-754)

Steroids as the novel class of high-affinity allosteric modulators of muscarinic receptors

2021 ◽  
Author(s):  
Eva Dolejší ◽  
Eszter Szánti-Pintér ◽  
Nikolai Chetverikov ◽  
Dominik Nelic ◽  
Alena Randáková ◽  
...  
Keyword(s):  
Muscarinic Receptors ◽  
Acetylcholine Receptors ◽  
Allosteric Modulation ◽  
G Protein Coupled Receptors ◽  
Muscarinic Acetylcholine Receptors ◽  
Allosteric Modulators ◽  
The Novel ◽  
Stress Control ◽  
Structure Activity ◽  
G Protein Coupled

Abstract The membrane cholesterol was found to bind and modulate the function of several G-protein coupled receptors including muscarinic acetylcholine receptors. We investigated the binding of 20 steroidal compounds including neurosteroids and steroid hormones to muscarinic receptors. Corticosterone, progesterone, and some neurosteroids bound to muscarinic receptors with an affinity of 100 nM or greater. We established a structure-activity relationship for steroid-based allosteric modulators of muscarinic receptors. Further, we show that corticosterone and progesterone allosterically modulate the functional response of muscarinic receptors to acetylcholine at physiologically relevant concentrations. It can play a role in stress control or in pregnancy, conditions where levels of these hormones dramatically oscillate. Allosteric modulation of muscarinic receptors via the cholesterol-binding site represents a new pharmacological approach at diseases associated with altered cholinergic signalling.

scholarly journals Yeast-Based Directed-Evolution For High-Throughput Structural Stabilization of G Protein-Coupled Receptors (GPCRs)

2021 ◽  
Author(s):  
May Meltzer ◽  
Zvagelsky Tatiana ◽  
Niv Papo ◽  
Stanislav Engel
Keyword(s):  
G Protein ◽  
Resistant Cell ◽  
Single Step ◽  
G Protein Coupled Receptors ◽  
Structural Basis ◽  
Diffusion Method ◽  
Structural Stabilization ◽  
C Terminus ◽  
Screening Platform ◽  
G Protein Coupled

Abstract The immense potential of G protein-coupled receptors (GPCRs) as targets for drug discovery is not fully realized due to the enormous difficulties associated with structure elucidation of these profoundly unstable membrane proteins. The existing methods of GPCR stability-engineering are cumbersome and low-throughput; in addition, the scope of GPCRs that could benefit from these techniques is limited. Here, we presented a yeast-based screening platform for a single-step isolation of GRCR variants stable in the presence of short-chain detergents, a feature essential for their successful crystallization using vapor diffusion method. The detergent-resistant cell wall of yeast provides a unique compartmentalization opportunity to physically link the receptor phenotype to its encoding DNA, and thus enable discovery of stable GPCR variants with unprecedent efficiency. The scope of mutations identified by the method offers important insights into the structural basis of GPCR stability, questioning the inherent instability of the GPCR scaffold, and revealing the potential role of the C-terminus in receptor stabilization.

Sign in / Sign up

Export Citation Format

Share Document