Abstract
To date, the visualization of beta2-adrenergic receptor (beta2AR) trafficking has been largely limited to immunocytochemical analyses of acute internalization events of epitope-tagged receptors in various transfection systems. The development of a beta2AR conjugated with green fluorescent protein (beta2AR-GFP) provides the opportunity for a more extensive optical analysis of beta2AR sequestration, down-regulation, and recycling in cells. Here we demonstrate that stable expression of beta2AR-GFP in HeLa cells enables a detailed temporal and spatial analysis of these events. Time-dependent colocalization of beta2AR-GFP with rhodamine-labeled transferrin and rhodamine-labeled dextran following agonist exposure demonstrates receptor distribution to early endosomes (sequestration) and lysosomes (down-regulation), respectively. The observed temporal distribution of beta2AR-GFP was consistent with measures of receptor sequestration and down-regulation generated by radioligand-receptor binding assays. Cells stimulated with different beta-agonists revealed time courses of beta2AR-GFP redistribution reflective of the intrinsic activity of each agonist.
Highlights
Upon agonist stimulation, 2-adrenergic receptors (2ARs)1 are rapidly desensitized by receptor phosphorylation [1]
The time course of 2AR redistribution assessed by confocal microscopy paralleled that of 2AR sequestration measured by radioligand binding
Internalized 2ARs colocalized with transferrin receptors, suggesting that sequestered 2ARs undergo processing through endosomal compartments in a manner similar to that observed for constitutively internalized receptors
Summary
Upon agonist stimulation, 2-adrenergic receptors (2ARs)1 are rapidly desensitized by receptor phosphorylation [1]. Radioligand binding experiments using HeLa cells transiently expressing the receptor constructs demonstrated pharmacological properties of 2AR-GFP that were similar to those observed for the wild type 2AR.
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
