title: Plasma membrane calcium pump (PMCA) isoform 4 is targeted to the apical membrane by the w-splice insert from PMCA2.
abstract: Local Ca(2+) signaling requires proper targeting of the Ca(2+) signaling toolkit to specific cellular locales. Different isoforms of the plasma membrane Ca(2+) pump (PMCA) are responsible for Ca(2+) extrusion at the apical and basolateral membrane of polarized epithelial cells, but the mechanisms and signals for differential targeting of the PMCAs are not well understood. Recent work demonstrated that the alternatively spliced w-insert in PMCA2 directs this pump to the apical membrane. We now show that inserting the w-insert into the corresponding location of the PMCA4 isoform confers apical targeting to this normally basolateral pump. Mutation of a di-leucine motif in the C-tail thought to be important for basolateral targeting did not enhance apical localization of the chimeric PMCA4(2w)/b. In contrast, replacing the C-terminal Val residue by Leu to optimize the PDZ ligand site for interaction with the scaffolding protein NHERF2 enhanced the apical localization of PMCA4(2w)/b, but not of PMCA4x/b. Functional studies showed that both apical PMCA4(2w)/b and basolateral PMCA4x/b handled ATP-induced Ca(2+) signals with similar kinetics, suggesting that isoform-specific functional characteristics are retained irrespective of membrane targeting. Our results demonstrate that the alternatively spliced w-insert provides autonomous apical targeting information in the PMCA without altering its functional characteristics.
authors.affiliation: Department of Molecular Cell Biology, National Blood Center, Diószegi u. 64, Budapest, Hungary.
authors.forename: Géza
authors.initials: G
authors.lastname: Antalffy
authors.forename: Amy S
authors.initials: AS
authors.lastname: Mauer
authors.forename: Katalin
authors.initials: K
authors.lastname: Pászty
authors.forename: Luca
authors.initials: L
authors.lastname: Hegedus
authors.forename: Rita
authors.initials: R
authors.lastname: Padányi
authors.forename: Agnes
authors.initials: A
authors.lastname: Enyedi
authors.forename: Emanuel E
authors.initials: EE
authors.lastname: Strehler
mesh_terms: D017398:Alternative Splicing / Q000502:physiology*; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D020013:Calcium Signaling / Q000502:physiology*; D002460:Cell Line; D002462:Cell Membrane / Q000201:enzymology* / Q000235:genetics; D004285:Dogs; D006801:Humans; D007527:Isoenzymes / Q000096:biosynthesis / Q000235:genetics; D053499:Plasma Membrane Calcium-Transporting ATPases / Q000096:biosynthesis* / Q000235:genetics
chemical_list: D007527:Isoenzymes; D053499:Plasma Membrane Calcium-Transporting ATPases; D002118:Calcium