Diffusion and Oligomerization States of the Muscarinic M1 Receptor in Live Cells − The Impact of Ligands and Membrane Disruptors
Bibliographic record
Abstract
ABSTRACT G protein-coupled receptors (GPCRs) are a major gateway to cellular signaling, which respond to ligands binding at extracellular sites through allosteric conformational changes that modulate their interactions with G proteins and arrestins at intracellular sites. High-resolution structures in different liganded states together with spectroscopic studies and molecular dynamics simulations have revealed a rich conformational landscape of GPCRs. However, their supramolecular spatio-temporal distribution is also thought to play a significant role in receptor activation and signaling bias within the native cell membrane environment. Here, we applied single-molecule fluorescence techniques, including single-particle tracking, single-molecule photobleaching and fluorescence correlation spectroscopy, to characterize the diffusion and oligomerization behavior of the muscarinic M 1 receptor (M 1 R) in live cells. Control samples included the monomeric protein CD86 and fixed cells, and experiments performed in the presence of different orthosteric M 1 R ligands and of several compounds known to change the fluidity and organization of the lipid bilayer. M 1 receptors exhibit Brownian diffusion characterized by three diffusion constants: confined/immobile (∼0.01 μm 2 /s), slow (∼0.04 μm 2 /s), and fast (∼0.14 μm 2 /s), whose populations were found to be modulated by both orthosteric ligands and membrane disruptors. The lipid raft disruptor C6-ceramide led to significant changes for CD86 while leaving the diffusion of M 1 R unchanged, indicating that M 1 receptors do not partition in lipid rafts. The extent of receptor oligomerization was found to be promoted by increasing the level of expression, and the binding of orthosteric ligands; in particular the agonist carbachol elicited a large increase in the fraction of M 1 R oligomers. This study provides new insights into the balance between conformational and environmental factors that define the movement and oligomerization states of GPCRs in live cells under close-to-native conditions.
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How this classification was reachedexpand
Full frame machine prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. The Gemma side is a direct model label for every work in the frame, read from the title-only record. The Codex side is a classifier learned from the 10,348 direct Codex labels and calibrated to design-weighted sample rates; fields without enough sample support carry no Codex call. Candidate is the union of the two sides; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels.
Distilled classifier scores by category (both heads)
| Category | Codex | Gemma |
|---|---|---|
| Metaresearch | 0.000 | 0.000 |
| Meta-epidemiology (narrow) | 0.000 | 0.000 |
| Meta-epidemiology (broad) | 0.000 | 0.000 |
| Bibliometrics | 0.000 | 0.000 |
| Science and technology studies | 0.000 | 0.000 |
| Scholarly communication | 0.000 | 0.000 |
| Open science | 0.000 | 0.000 |
| Research integrity | 0.000 | 0.000 |
| Insufficient payload (model declined to judge) | 0.001 | 0.000 |
Machine scores (provisional)
The two teacher heads of the student model, read on this work. A score orders the frame for review; it never asserts a category, and the validation status ships verbatim with every row.
Baseline scores from an immature model (maturity gate not passed, 7 training rounds). Scores rank; they never assert a category.
score_only:v0-immature-baseline · verbatim from the scoring run: score_only means the number may rank works, and no category label ships from itClassification
machine, unvalidatedMachine predicted; a candidate call from one source (direct Gemma or distilled Codex), not a consensus.
How this classification was reached, model by model and score by score, is at the end of the page under "How this classification was reached".