title: Lysosomal Targeting of Cystinosin Requires AP-3.
abstract: Cystinosin is a lysosomal cystine transporter defective in cystinosis, an autosomal recessive lysosomal storage disorder. It is composed of seven transmembrane (TM) domains and contains two lysosomal targeting motifs: a tyrosine-based signal (GYDQL) in its C-terminal tail and a non-classical motif in its fifth inter-TM loop. Using the yeast two-hybrid system, we showed that the GYDQL motif specifically interacted with the μ subunit of the adaptor protein complex 3 (AP-3). Moreover, cell surface biotinylation and total internal reflection fluorescence microscopy revealed that cystinosin was partially mislocalized to the plasma membrane (PM) in AP-3-depleted cells. We generated a chimeric CD63 protein to specifically analyze the function of the GYDQL motif. This chimeric protein was targeted to lysosomes in a manner similar to cystinosin and was partially mislocalized to the PM in AP-3 knockdown cells where it also accumulated in the trans-Golgi network and early endosomes. Together with the fact that the surface levels of cystinosin and of the CD63-GYDQL chimeric protein were not increased when clathrin-mediated endocytosis was impaired, our data show that the tyrosine-based motif of cystinosin is a 'strong' AP-3 interacting motif responsible for lysosomal targeting of cystinosin by a direct intracellular pathway.
authors.affiliation: Inserm U1163, Laboratory of Hereditary Kidney Diseases, Paris, France.|Paris Descartes-Sorbonne Paris Cité University, Imagine Institute, Paris, France.
authors.forename: Zuzanna
authors.initials: Z
authors.lastname: Andrzejewska
authors.affiliation: Inserm U1163, Laboratory of Hereditary Kidney Diseases, Paris, France.|Paris Descartes-Sorbonne Paris Cité University, Imagine Institute, Paris, France.
authors.forename: Nathalie
authors.initials: N
authors.lastname: Névo
authors.affiliation: Inserm U1163, Laboratory of Hereditary Kidney Diseases, Paris, France.|Paris Descartes-Sorbonne Paris Cité University, Imagine Institute, Paris, France.
authors.forename: Lucie
authors.initials: L
authors.lastname: Thomas
authors.affiliation: Inserm U1163, Laboratory of Hereditary Kidney Diseases, Paris, France.|Paris Descartes-Sorbonne Paris Cité University, Imagine Institute, Paris, France.
authors.forename: Anne
authors.initials: A
authors.lastname: Bailleux
authors.affiliation: Inserm U1163, Laboratory of Hereditary Kidney Diseases, Paris, France.|Paris Descartes-Sorbonne Paris Cité University, Imagine Institute, Paris, France.
authors.forename: Véronique
authors.initials: V
authors.lastname: Chauvet
authors.affiliation: Inserm U1163, Laboratory of Hereditary Kidney Diseases, Paris, France.|Paris Descartes-Sorbonne Paris Cité University, Imagine Institute, Paris, France.
authors.forename: Alexandre
authors.initials: A
authors.lastname: Benmerah
authors.affiliation: Inserm U1163, Laboratory of Hereditary Kidney Diseases, Paris, France.|Paris Descartes-Sorbonne Paris Cité University, Imagine Institute, Paris, France.|Assistance Publique - Hôpitaux de Paris (AP-HP), Department of Genetics, Necker Hospital, Paris,, France.
authors.forename: Corinne
authors.initials: C
authors.lastname: Antignac
mesh_terms: D033963:Adaptor Protein Complex 3 / Q000378:metabolism*; D026921:Amino Acid Transport Systems, Neutral / Q000737:chemistry / Q000378:metabolism*; D004705:Endocytosis; D006367:HeLa Cells; D006801:Humans; D008247:Lysosomes / Q000378:metabolism*; D021382:Protein Sorting Signals; D021381:Protein Transport; D060149:Tetraspanin 30 / Q000378:metabolism
chemical_list: D033963:Adaptor Protein Complex 3; D026921:Amino Acid Transport Systems, Neutral; C111445:CTNS protein, human; D021382:Protein Sorting Signals; D060149:Tetraspanin 30
keywords: adaptor protein complex; cystinosin; cystinosis; lysosomal storage disorder; lysosomal targeting; transmembrane protein; tyrosine-based motif