Interaction of the amyloid precursor like protein 1 with the alpha2A-adrenergic receptor increases agonist-mediated inhibition of adenylate cyclase.
Alpha2-adrenergic receptor agonists exert potent analgesic and sedative/hypnotic effects. In addition, they have been shown to be neuroprotective, but the mechanisms of these actions are still poorly defined. To isolate proteins that may control alpha2-adrenergic receptor function or trafficking, we performed a two-hybrid screen using the carboxy-terminal fourth intracellular tail of the alpha2A-adrenergic receptor as bait. This screen identified the amyloid precursor like protein 1 (APLP1), a homologue of the beta-amyloid precursor protein involved in Alzheimer's disease, as alpha2A-adrenergic receptor-binding protein. GST affinity chromatography revealed that APLP1 specifically interacts with all three human alpha2-adrenergic receptor subtypes and deletion mutant analysis confined the APLP1 domain involved in binding to alpha2-adrenergic receptors to the 13 amino acid residues Ser599-Ala611. Coimmunoprecipitations of transiently transfected cells with epitope-tagged APLP1 and alpha2-adrenergic receptors confirmed the interaction. Agonist treatment tended to increase the amount of alpha2A-adrenergic receptor associated with APLP1 while coimmunoprecipitations were not affected by the state of receptor phosphorylation or cotransfection of arrestin-3. Confocal laser microscopy showed that APLP1 causes a considerable shift of the alpha2A-adrenergic receptor localization from plasma membrane to intracellular compartments. Furthermore, cotransfection of alpha2A-adrenergic receptor and APLP1 into HEK293 cells significantly increased norepinephrine mediated inhibition of adenylate cyclase activity. These results suggest a possible role of APLP1 in regulation of alpha2A-adrenergic receptor trafficking. Moreover, we speculate that this interaction may present one mechanism by which alpha2-adrenergic receptor agonists exert their neuroprotective effects.
Department of Anaesthesiology and Intensive Care Medicine, University Hospital Schleswig-Holstein, Campus Kiel, Schwanenweg 21, 24105 Kiel, Germany. weber@anaesthesie.uni-kiel.de
Bernd
B
Weber
Christian
C
Schaper
Jens
J
Scholz
Berthold
B
Bein
Cornelia
C
Rodde
Peter
P
H Tonner
D000067956:Adenylyl Cyclase Inhibitors; D000595:Amino Acid Sequence; D016564:Amyloid beta-Protein Precursor / Q000737:chemistry / Q000378:metabolism*; D000818:Animals; D019390:Arrestins / Q000378:metabolism; D019556:COS Cells; D002522:Chlorocebus aethiops; D005982:Glutathione Transferase / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D008969:Molecular Sequence Data; D010766:Phosphorylation; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D018341:Receptors, Adrenergic, alpha-2 / Q000737:chemistry / Q000378:metabolism*; D020798:Two-Hybrid System Techniques
C547768:ADRA2A protein, human; C489697:APLP1 protein, human; D000067956:Adenylyl Cyclase Inhibitors; D016564:Amyloid beta-Protein Precursor; D019390:Arrestins; D018341:Receptors, Adrenergic, alpha-2; C082333:arrestin3; D005982:Glutathione Transferase