title: Carboxyl-terminal truncations of human anion exchanger impair its trafficking to the plasma membrane.
abstract: The human anion exchanger AE1 (Band 3) is an abundant glycoprotein localized in plasma membrane of red cells and is responsible for the electro-neutral exchange of chloride for bicarbonate. In order to determine the role of the carboxyl-terminal tail of AE1 in its expression, function and trafficking to the plasma membrane, we generated a series of five constructs encoding truncation mutants missing the last 5 (Delta5), 11 (Delta11), 15 (Delta15), 20 (Delta20) or 35 (Delta35) amino-acids. In transiently transfected HEK 293 cells, immunoblotting of whole cell extracts showed that all the proteins were expressed at the same level as full-length AE1, except Delta20 and particularly Delta35, which showed a reduced expression. Furthermore, the last 15 amino-acids were not required for AE1 folding in the membrane, since Delta5, Delta11 and Delta15 were able to bind to an inhibitor affinity matrix, while Delta20 and Delta35 exhibited poor binding. Immunofluorescence and deglycosylation results showed that Delta15 and Delta11 were retained intracellularly, whereas a lower amount of Delta5 compared with WT trafficked to the plasma membrane. These results indicate that an intact C-terminal tail of human AE1 is important for efficient AE1 trafficking to the plasma membrane.
authors.affiliation: Departments of Biochemistry and Medicine, CIHR Group in Membrane Biology, University of Toronto, Toronto, Ontario M5S 1A8, Canada.
authors.forename: Emmanuelle
authors.initials: E
authors.lastname: Cordat
authors.forename: Jing
authors.initials: J
authors.lastname: Li
authors.forename: Reinhart A F
authors.initials: RA
authors.lastname: Reithmeier
mesh_terms: D000595:Amino Acid Sequence; D001457:Anion Exchange Protein 1, Erythrocyte / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D002462:Cell Membrane / Q000737:chemistry / Q000378:metabolism*; D004721:Endoplasmic Reticulum / Q000737:chemistry; D006801:Humans; D008969:Molecular Sequence Data; D021381:Protein Transport; D017384:Sequence Deletion
chemical_list: D001457:Anion Exchange Protein 1, Erythrocyte