A novel interaction between adrenergic receptors and the alpha-subunit of eukaryotic initiation factor 2B.
The alpha-subunit of eukaryotic initiation factor 2B (eIF-2B), a guanine nucleotide exchange protein that functions in regulation of translation, was observed to associate with the carboxyl-terminal cytoplasmic domains of the alpha2A- and alpha2B-adrenergic receptors in a yeast two-hybrid screen of a cDNA library prepared from 293 cells. This protein association was confirmed in vitro by affinity chromatography and was shown to be specific for a subset of G protein-coupled receptors, including the alpha2A-, alpha2B-, alpha2C-, and beta2-adrenergic receptors, but not the vasopressin (V2) receptor. Association of these proteins in vivo was confirmed by specific co-immunoprecipitation of eIF-2Balpha with full-length beta2-adrenergic receptors expressed in transfected 293 cells and by fluorescence microscopy showing co-localization of these proteins in intact cells. Remarkably, eIF-2Balpha co-localized with receptors exclusively in regions of the plasma membrane that are in contact with the extracellular medium, but failed to associate with membranes making cell-cell contacts. Overexpression of eIF-2Balpha in 293 cells caused a small (approximately 15%) but significant enhancement of beta2-adrenergic receptor-mediated activation of adenylyl cyclase, without affecting forskolin or V2 receptor-mediated activation. These observations suggest a new role for a previously identified guanine nucleotide exchange protein in membrane biology and cell signaling.
Klein
U
U
Department of Division of Cardiovascular Medicine, Stanford University Medical School, Stanford, California 94305, USA.
Ramirez
M T
MT
Kobilka
B K
BK
von Zastrow
M
M
D000595:Amino Acid Sequence; D019204:GTP-Binding Proteins / Q000378:metabolism; D020662:Guanine Nucleotide Exchange Factors; D008969:Molecular Sequence Data; D011506:Proteins / Q000378:metabolism*; D018343:Receptors, Adrenergic, beta-2 / Q000378:metabolism*
D020662:Guanine Nucleotide Exchange Factors; D011506:Proteins; D018343:Receptors, Adrenergic, beta-2; D019204:GTP-Binding Proteins