title: The C-terminal half of the α2C-adrenoceptor determines the receptor's membrane expression level and drug selectivity.
abstract: In tissues as well as in transfected cells, α2C-adrenoceptors show poorer expression levels compared to α2A-adrenoceptors. In order to characterize which regions of the α2C-adrenoceptor are involved in regulating the expression of binding-competent receptors at the plasma membrane, six chimeric α2A-/α2C-adrenoceptors were constructed. The wild-type α2A- and α2C-adrenoceptors and the six chimeric α2A-/α2C-adrenoceptors were transiently transfected into human embryonic kidney 293 (HEK293) cells, and the expression levels were investigated by radioligand binding. The results show that the C-terminal half of the α2C-adrenoceptor, ranging from the second extracellular loop to the C-terminus, is the main determinant of the low expression level of binding-competent α2C-adrenoceptors in HEK293 cell membranes. The so-called retention signal in the N-terminus of the α2C-adrenoceptor had a less profound effect on the expression levels of the chimeric receptors. For seven drugs competing for [(3)H]-RX821002 binding, the K i values were determined at the wild-type α2A- and α2C-adrenoceptors and at four of the chimeric α2A-/α2C-adrenoceptors. The results show that the α2C- over α2A-selectivity of spiroxatrine, spiperone, clozapine, MK912, and chlorpromazine, as well as the α2A- over α2C-selectivity of BRL44408, resides mainly in the C-terminal half of the receptors. To some extent, the region comprising the N-terminal half of the receptors contributed to the α2C-selectivity of spiperone, clozapine, and chlorpromazine.
authors.affiliation: Section of Pharmacology/Center for Pharmacy, Department of Clinical Science, University of Bergen, NLB, 9th floor, 5021, Bergen, Norway.
authors.forename: Jan Anker
authors.initials: JA
authors.lastname: Jahnsen
authors.forename: Staffan
authors.initials: S
authors.lastname: Uhlén
mesh_terms: D000595:Amino Acid Sequence; D001665:Binding Sites; D001667:Binding, Competitive; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism*; D057809:HEK293 Cells; D006801:Humans; D008024:Ligands; D008969:Molecular Sequence Data; D017434:Protein Structure, Tertiary; D011869:Radioligand Assay; D018341:Receptors, Adrenergic, alpha-2 / Q000737:chemistry / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D014162:Transfection
chemical_list: C547771:ADRA2B protein, human; C547774:ADRA2C protein, human; D008024:Ligands; D018341:Receptors, Adrenergic, alpha-2; D011993:Recombinant Fusion Proteins