title: Targeting of G protein-coupled receptors to the basolateral surface of polarized renal epithelial cells involves multiple, non-contiguous structural signals.
abstract: Truncations and chimeras of the alpha2A-adrenergic receptor (alpha2AAR) were evaluated to identify membrane domains responsible for its direct basolateral targeting in Madin-Darby canine kidney cells. An alpha2AAR truncation, encoding transmembrane (TM) regions 1-5, was first delivered basolaterally, but within minutes appeared apically, and at steady-state was primarily lateral in its immunocytochemical localization. A TM 1-5 truncation with the third intracellular loop revealed more intense lateral localization than for the TM 1-5 structure, consistent with the role of the third intracellular loop in alpha2AAR stabilization. Addition of TM 6-7 of A1 adenosine receptor (A1AdoR) to alpha2AARTM1-5 creates a chimera, alpha2AARTM1-5/A1AdoRTM6-7, which was first delivered apically, resulting either from loss of alpha2AAR sorting information in TM 6-7 or acquisition of apical trafficking signals within A1AdoRTM6-7. Evidence that alpha2AARTM6-7 imparts basolateral targeting information is revealed by the significant basolateral localization of the A1AdoRTM1-5/alpha2AARTM6-7 and A1AdoRTM1-5/alpha2AARTM6-7+i3 chimeras, in contrast to the dominant apical localization of A1AdoR. These results reveal that sequences within TM 1-5 and within TM 6-7 of the alpha2AAR confer basolateral targeting, providing the first evidence that alpha2AAR basolateral localization is not conferred by a single region but by non-contiguous membrane-embedded or proximal sequences.
authors.affiliation: Department of Pharmacology, Vanderbilt University Medical Center, Nashville, Tennessee 37232-6600, USA.
authors.forename: C
authors.initials: C
authors.lastname: Saunders
authors.forename: J R
authors.initials: JR
authors.lastname: Keefer
authors.forename: C A
authors.initials: CA
authors.lastname: Bonner
authors.forename: L E
authors.initials: LE
authors.lastname: Limbird
mesh_terms: D000818:Animals; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D016764:Cell Polarity / Q000502:physiology*; D004285:Dogs; D004847:Epithelial Cells / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000378:metabolism*; D007668:Kidney / Q000378:metabolism*; D008958:Models, Molecular; D017433:Protein Structure, Secondary; D018341:Receptors, Adrenergic, alpha-2 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D018047:Receptors, Purinergic P1 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D017384:Sequence Deletion; D015398:Signal Transduction; D014162:Transfection
chemical_list: D018341:Receptors, Adrenergic, alpha-2; D018047:Receptors, Purinergic P1; D011993:Recombinant Fusion Proteins; D019204:GTP-Binding Proteins