title: Oligomerization of the UapA Purine Transporter Is Critical for ER-Exit, Plasma Membrane Localization and Turnover.
abstract: Central to the process of transmembrane cargo trafficking is the successful folding and exit from the ER (endoplasmic reticulum) through packaging in COPII vesicles. Here, we use the UapA purine transporter of Aspergillus nidulans to investigate the role of cargo oligomerization in membrane trafficking. We show that UapA oligomerizes (at least dimerizes) and that oligomerization persists upon UapA endocytosis and vacuolar sorting. Using a validated bimolecular fluorescence complementation assay, we provide evidence that a UapA oligomerization is associated with ER-exit and turnover, as ER-retained mutants due to either modification of a Tyr-based N-terminal motif or partial misfolding physically associate but do not associate properly. Co-expression of ER-retained mutants with wild-type UapA leads to in trans plasma membrane localization of the former, confirming that oligomerization initiates in the ER. Genetic suppression of an N-terminal mutation in the Tyr motif and mutational analysis suggest that transmembrane α-helix 7 affects the oligomerization interface. Our results reveal that transporter oligomerization is essential for membrane trafficking and turnover and is a common theme in fungi and mammalian cells.
authors.affiliation: Faculty of Biology, University of Athens, Panepistimioupolis 15784, Athens, Greece.
authors.forename: Olga
authors.initials: O
authors.lastname: Martzoukou
authors.affiliation: Faculty of Biology, University of Athens, Panepistimioupolis 15784, Athens, Greece.
authors.forename: Mayia
authors.initials: M
authors.lastname: Karachaliou
authors.affiliation: Faculty of Biology, University of Athens, Panepistimioupolis 15784, Athens, Greece.
authors.forename: Vassilis
authors.initials: V
authors.lastname: Yalelis
authors.affiliation: Division of Molecular Biology, Imperial College, London SW7 2AZ, UK.
authors.forename: James
authors.initials: J
authors.lastname: Leung
authors.affiliation: Division of Molecular Biology, Imperial College, London SW7 2AZ, UK.
authors.forename: Bernadette
authors.initials: B
authors.lastname: Byrne
authors.affiliation: Faculty of Biology, University of Athens, Panepistimioupolis 15784, Athens, Greece.
authors.forename: Sotiris
authors.initials: S
authors.lastname: Amillis
authors.affiliation: Faculty of Biology, University of Athens, Panepistimioupolis 15784, Athens, Greece. Electronic address: diallina@biol.uoa.gr.
authors.forename: George
authors.initials: G
authors.lastname: Diallinas
mesh_terms: D001233:Aspergillus nidulans / Q000235:genetics / Q000378:metabolism*; D001426:Bacterial Proteins / Q000235:genetics; D022181:COP-Coated Vesicles / Q000502:physiology; D002462:Cell Membrane / Q000378:metabolism; D004705:Endocytosis / Q000502:physiology; D004721:Endoplasmic Reticulum / Q000235:genetics / Q000378:metabolism; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism*; D049452:Green Fluorescent Proteins / Q000235:genetics; D008164:Luminescent Proteins / Q000235:genetics; D026901:Membrane Transport Proteins / Q000235:genetics / Q000378:metabolism*; D055503:Protein Multimerization / Q000502:physiology*; D017434:Protein Structure, Tertiary; D021381:Protein Transport / Q000235:genetics / Q000502:physiology*
chemical_list: D001426:Bacterial Proteins; D005656:Fungal Proteins; D008164:Luminescent Proteins; D026901:Membrane Transport Proteins; C083852:UAPA protein, Aspergillus nidulans; C054562:yellow fluorescent protein, Bacteria; D049452:Green Fluorescent Proteins
keywords: bimolecular fluorescence; endocytosis; membrane sorting; trafficking; transport