Dynamic in Situ Confinement Triggers Ligand-Free Neuropeptide Receptor Signaling

Area of Science:

• Cell biology
• Molecular signaling
• Biophysics

Background:

• Membrane receptor clustering is crucial for cell-cell communication.
• The precise physiological roles of receptor clustering in cell signaling are not fully understood.

Purpose of the Study:

• To investigate the function of receptor clustering in cell signaling.
• To develop a method for inducing and studying receptor clustering in situ.

Main Methods:

• Development of a dynamic platform using a chelator nanotool to induce in situ clustering of histidine-tagged neuropeptide Y2 receptors (Y2R).
• Analysis of Y2R dynamics within clusters, including multivalent interactions and dynamic exchange.
• Measurement of downstream signaling events, such as cytosolic calcium increase and cell migration, in response to clustering and ligand stimulation.
• Investigation of arrestin-3 recruitment as a downstream effector.

Main Results:

• Induced Y2R clustering led to ligand-independent signaling, evidenced by increased cytosolic calcium and cell migration.
• Pre-clustering of Y2R amplified the calcium and motility response to ligand-induced activation, suggesting sensitization to lower ligand concentrations.
• Ligand-independent and ligand-induced signaling pathways differed in arrestin-3 recruitment; arrestin-3 was recruited only upon ligand presence.

Conclusions:

• In situ receptor clustering can trigger ligand-independent signaling pathways.
• Receptor clustering plays a key role in sensitizing cellular responses to external stimuli, such as neuropeptides.
• The developed platform enables exploration of diverse receptor activation mechanisms and signaling modalities.