title: Retromer- and WASH-dependent sorting of nutrient transporters requires a multivalent interaction network with ANKRD50.
abstract: Retromer and the associated actin-polymerizing WASH complex are essential for the endocytic recycling of a wide range of integral membrane proteins. A hereditary Parkinson's-disease-causing point mutation (D620N) in the retromer subunit VPS35 perturbs retromer's association with the WASH complex and also with the uncharacterized protein ankyrin-repeat-domain-containing protein 50 (ANKRD50). Here, we firmly establish ANKRD50 as a new and essential component of the SNX27-retromer-WASH super complex. Depletion of ANKRD50 in HeLa or U2OS cells phenocopied the loss of endosome-to-cell-surface recycling of multiple transmembrane proteins seen upon suppression of SNX27, retromer or WASH-complex components. Mass-spectrometry-based quantification of the cell surface proteome of ANKRD50-depleted cells identified amino acid transporters of the SLC1A family, among them SLC1A4, as additional cargo molecules that depend on ANKRD50 and retromer for their endocytic recycling. Mechanistically, we show that ANKRD50 simultaneously engages multiple parts of the SNX27-retromer-WASH complex machinery in a direct and co-operative interaction network that is needed to efficiently recycle the nutrient transporters GLUT1 (also known as SLC2A1) and SLC1A4, and potentially many other surface proteins.
authors.affiliation: Center for Biological Systems Analysis (ZBSA), Albert Ludwigs Universitaet Freiburg, Habsburgerstrasse 49, Freiburg 79104, Germany.|Faculty of Biology, Schaenzlestrasse 1, D-79104, Freiburg, Germany.
authors.forename: Arunas
authors.initials: A
authors.lastname: Kvainickas
authors.affiliation: Center for Biological Systems Analysis (ZBSA), Albert Ludwigs Universitaet Freiburg, Habsburgerstrasse 49, Freiburg 79104, Germany.|Faculty of Biology, Schaenzlestrasse 1, D-79104, Freiburg, Germany.
authors.forename: Ana Jimenez
authors.initials: AJ
authors.lastname: Orgaz
authors.affiliation: Center for Biological Systems Analysis (ZBSA), Albert Ludwigs Universitaet Freiburg, Habsburgerstrasse 49, Freiburg 79104, Germany.|Faculty of Biology, Schaenzlestrasse 1, D-79104, Freiburg, Germany.
authors.forename: Heike
authors.initials: H
authors.lastname: Nägele
authors.affiliation: Center for Biological Systems Analysis (ZBSA), Albert Ludwigs Universitaet Freiburg, Habsburgerstrasse 49, Freiburg 79104, Germany.|Faculty of Biology, Schaenzlestrasse 1, D-79104, Freiburg, Germany.
authors.forename: Britta
authors.initials: B
authors.lastname: Diedrich
authors.affiliation: School of Biochemistry, Bristol University, University Walk, Bristol BS81TD, UK.
authors.forename: Kate J
authors.initials: KJ
authors.lastname: Heesom
authors.affiliation: Department of Biology, Fribourg University, Chemin du Musee 10, Fribourg CH-1700, Switzerland.
authors.forename: Jörn
authors.initials: J
authors.lastname: Dengjel
authors.affiliation: School of Biochemistry, Bristol University, University Walk, Bristol BS81TD, UK.
authors.forename: Peter J
authors.initials: PJ
authors.lastname: Cullen
authors.affiliation: Center for Biological Systems Analysis (ZBSA), Albert Ludwigs Universitaet Freiburg, Habsburgerstrasse 49, Freiburg 79104, Germany florian.steinberg@zbsa.de.|Faculty of Biology, Schaenzlestrasse 1, D-79104, Freiburg, Germany.
authors.forename: Florian
authors.identifier: 0000-0003-1849-0506
authors.initials: F
authors.lastname: Steinberg
mesh_terms: D020816:Amino Acid Motifs; D026962:Amino Acid Transport System ASC / Q000378:metabolism*; D001692:Biological Transport; D002462:Cell Membrane / Q000378:metabolism; D004705:Endocytosis; D011992:Endosomes / Q000378:metabolism; D066246:ErbB Receptors / Q000378:metabolism; D055785:Gene Knockdown Techniques; D051272:Glucose Transporter Type 1 / Q000378:metabolism*; D057809:HEK293 Cells; D006367:HeLa Cells; D006801:Humans; D016023:Integrins / Q000378:metabolism; D013058:Mass Spectrometry; D008840:Microfilament Proteins / Q000378:metabolism*; D010749:Phosphoprotein Phosphatases / Q000737:chemistry / Q000378:metabolism*; D011485:Protein Binding; D060066:Protein Interaction Maps; D059748:Proteolysis; D040901:Proteomics; D058305:Sorting Nexins / Q000378:metabolism; D014168:Transferrin / Q000378:metabolism; D033921:Vesicular Transport Proteins / Q000737:chemistry / Q000378:metabolism*
chemical_list: C000621799:ANKRD50 protein, human; D026962:Amino Acid Transport System ASC; D051272:Glucose Transporter Type 1; D016023:Integrins; D008840:Microfilament Proteins; C498848:SLC1A4 protein, human; D058305:Sorting Nexins; D014168:Transferrin; D033921:Vesicular Transport Proteins; C549919:WASH protein, human; D066246:ErbB Receptors; D010749:Phosphoprotein Phosphatases
keywords: Endosome; Membrane trafficking; Retromer; SNX27; WASH complex