title: Binding of "AT4 receptor" ligands to insulin regulated aminopeptidase (IRAP) in intact Chinese hamster ovary cells.
abstract: Insulin regulated aminopeptidase (IRAP) recognises "AT(4)-receptor" ligands like angiotensin IV (Ang IV) and peptidomimetics like AL-11. The metabolic stability and high affinity of [(3)H]AL-11 for catalytically active IRAP allowed its detection in Chinese hamster ovary (CHO-K1) cell membranes in the absence of chelators (Demaegdt et al., 2009). Here, we show that, contrary to [(3)H]Ang IV, [(3)H]AL-11 displays high affinity and specificity for IRAP in intact CHO-K1 cells as well. After binding to IRAP at the surface, [(3)H]AL-11 is effectively internalized by an endocytotic process. Unexpectedly, surface binding and internalization of [(3)H]AL-11 was not affected by pretreating the cells with Ang IV but declined with AL-11. In the latter case surface expression of IRAP even increased. After elimination of simpler explanations, it is proposed that metabolically stable "AT(4)-receptor" ligands undergo semi-continuous cycling between the cell surface and endosomal compartments. The in vivo efficacy of stable and unstable "AT(4)-receptor" ligands could therefore differ.
authors: Department of Molecular and Biochemical Pharmacology, Research Group of Experimental Pharmacology, Vrije Universiteit Brussel, Pleinlaan 2, 1050 Brussels, Belgium.
authors: Heidi
authors: H
authors: Demaegdt
authors: Paul
authors: P
authors: Gard
authors: Jean-Paul
authors: JP
authors: De Backer
authors: Aneta
authors: A
authors: Lukaszuk
authors: Erzsébet
authors: E
authors: Szemenyei
authors: Géza
authors: G
authors: Tóth
authors: Dirk
authors: D
authors: Tourwé
authors: Georges
authors: G
authors: Vauquelin
mesh_terms: D000804:Angiotensin II / Q000031:analogs & derivatives* / Q000494:pharmacology; D000818:Animals; D001667:Binding, Competitive; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D010122:Cystinyl Aminopeptidase / Q000378:metabolism*; D057809:HEK293 Cells; D006454:Hemoglobins / Q000494:pharmacology; D006801:Humans; D008024:Ligands; D010446:Peptide Fragments / Q000494:pharmacology; D057786:Peptidomimetics / Q000494:pharmacology*; D011485:Protein Binding; D021381:Protein Transport; D011869:Radioligand Assay
chemical_list: D006454:Hemoglobins; D008024:Ligands; D010446:Peptide Fragments; D057786:Peptidomimetics; D000804:Angiotensin II; C012877:angiotensin II, des-Asp(1)-des-Arg(2)-Ile(5)-; C074309:LVV-hemorphin-7; D010122:Cystinyl Aminopeptidase; C105964:leucyl-cystinyl aminopeptidase