title: Multiple cytoplasmic signals direct the intracellular trafficking of chicken kidney AE1 anion exchangers in MDCK cells.
abstract: AE1/F(c) receptor chimeras have been used to define the sequences that direct the basolateral sorting, recycling and cytoskeletal association of the chicken AE1-4 anion exchanger in MDCK cells. These analyses revealed that amino acids 1-63 of AE1-4 were sufficient to redirect a cytoplasmic tailless murine IgG F(c)RII B2 receptor from the apical to the basolateral membrane of MDCK cells, where F(c)1-63 associated with elements of the actin cytoskeleton. In contrast to F(c)1-63, chimeras containing amino acids 1-37 (F(c)1-37) or 38-63 (F(c)38-63) of AE1-4 accumulated in intracellular membrane compartments that overlapped late endosomes and the trans-Golgi network (TGN), respectively. Internalization assays indicated that the patterns of localization observed for F(c)1-37 and F(c)38-63 resulted from the recycling of these chimeras from the cell surface. These assays further indicated that F(c)1-37 and F(c)38-63 each possess a basolateral sorting activity. Mutagenesis studies revealed that the endocytic and basolateral sorting activities in F(c)1-37 are dependent upon serine 25, which is located in a sequence similar to a sorting signal in the polymeric immunoglobulin receptor. In addition, the sorting activities associated with F(c)38-63 were dependent upon tyrosine 47 and leucine 50. These residues resided within the sequence, YVEL, which matches the YXXPhi motif (where X is any amino acid and Phi is a hydrophobic residue) that functions as an endocytic and TGN recycling signal for other membrane proteins. Our data indicate that amino acids 1-63 of AE1-4 contain sorting and cytoskeletal binding activities that account for most of the properties previously associated with AE1-4 in MDCK cells. Furthermore, the alternative localization patterns exhibited by chimeras containing various combinations of these activities suggest that interplay between these cytoplasmic activities is critical for specifying AE1-4 localization in epithelial cells.
authors.affiliation: Department of Molecular Sciences, University of Tennessee Health Science Center, 858 Madison Avenue, Memphis, Tennessee 38163, USA. jcox@utmem.edu
authors.forename: Tracy L
authors.initials: TL
authors.lastname: Adair-Kirk
authors.forename: Frank C
authors.initials: FC
authors.lastname: Dorsey
authors.forename: John V
authors.initials: JV
authors.lastname: Cox
mesh_terms: D000595:Amino Acid Sequence / Q000502:physiology; D000596:Amino Acids / Q000378:metabolism; D000818:Animals; D001457:Anion Exchange Protein 1, Erythrocyte / Q000378:metabolism*; D000838:Anions / Q000378:metabolism*; D002460:Cell Line; D002645:Chickens / Q000378:metabolism; D003593:Cytoplasm / Q000378:metabolism*; D003599:Cytoskeleton / Q000378:metabolism; D004285:Dogs; D011992:Endosomes / Q000378:metabolism; D004847:Epithelial Cells / Q000378:metabolism*; D005455:Fluorescent Antibody Technique; D007424:Intracellular Fluid / Q000378:metabolism*; D007425:Intracellular Membranes / Q000378:metabolism; D007668:Kidney / Q000378:metabolism; D008969:Molecular Sequence Data; D011485:Protein Binding / Q000502:physiology; D021381:Protein Transport / Q000502:physiology*; D011993:Recombinant Fusion Proteins; D015398:Signal Transduction / Q000502:physiology
chemical_list: D000596:Amino Acids; D001457:Anion Exchange Protein 1, Erythrocyte; D000838:Anions; D011993:Recombinant Fusion Proteins