title: Plasma membrane compartmentalization of D2 dopamine receptors.
abstract: Plasma membrane microcompartments could allow different signaling pathways to operate more efficiently and prevent cross-talk. We utilized a novel in-cell biotin transfer assay to demonstrate that the majority of plasma membrane-expressed D2 dopamine receptor (D2R) is microcompartmentalized within detergent-resistant structures. Conversely, a minority of D2R existed in a detergent-soluble form and interacted in a relatively unrestricted manner with other cellular proteins. The microcompartmentalization of D2R had functional consequences because dopamine-induced internalization of D2R was largely restricted to the compartmentalized receptor. The D2R-containing microcompartments did not correspond to putative detergent-resistant lipid raft structures. First, the detergent-insoluble D2R structures were significantly denser than detergent-resistant membrane fragments containing flotillin, a widely utilized lipid raft marker protein. Second, the detergent solubility of D2R was unaffected by treatment of cells with the cholesterol chelating agent, methyl-β-cyclodextrin, that is thought to disrupt lipid rafts. Finally, the in-cell biotinylation assay did not provide any evidence for the membrane compartmentalization of peptide motifs thought to target to lipid rafts. Thus, our observations form one of the first demonstrations, in living cells, of plasma membrane microcompartments defined by the ability of the compartment structure to broadly restrict the interaction of resident molecules with other cellular proteins.
authors.affiliation: Department of Biomedical and Pharmaceutical Sciences, University of Rhode Island, Kingston, Rhode Island 02881, USA.
authors.forename: Meenakshi
authors.initials: M
authors.lastname: Sharma
authors.forename: Jeremy
authors.initials: J
authors.lastname: Celver
authors.forename: J Christopher
authors.initials: JC
authors.lastname: Octeau
authors.forename: Abraham
authors.initials: A
authors.lastname: Kovoor
mesh_terms: D020816:Amino Acid Motifs; D000818:Animals; D057809:HEK293 Cells; D006801:Humans; D021962:Membrane Microdomains / Q000235:genetics / Q000378:metabolism*; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D051379:Mice; D017448:Receptors, Dopamine D2 / Q000235:genetics / Q000378:metabolism*; D047392:beta-Cyclodextrins / Q000494:pharmacology
chemical_list: D008565:Membrane Proteins; D017448:Receptors, Dopamine D2; D047392:beta-Cyclodextrins; C106354:flotillins; C108732:methyl-beta-cyclodextrin
keywords: 7-Helix Receptor; Arrestin; Cell Compartmentation; Dopamine Receptors; G Protein-coupled Receptor; Plasma Membrane; Plasma Membrane Compartmentalization; Receptor Internalization; Receptor Regulation