title: PDE6δ-mediated sorting of INPP5E into the cilium is determined by cargo-carrier affinity.
abstract: The phosphodiesterase 6 delta subunit (PDE6δ) shuttles several farnesylated cargos between membranes. The cargo sorting mechanism between cilia and other compartments is not understood. Here we show using the inositol polyphosphate 5'-phosphatase E (INPP5E) and the GTP-binding protein (Rheb) that cargo sorting depends on the affinity towards PDE6δ and the specificity of cargo release. High-affinity cargo is exclusively released by the ciliary transport regulator Arl3, while low-affinity cargo is released by Arl3 and its non-ciliary homologue Arl2. Structures of PDE6δ/cargo complexes reveal the molecular basis of the sorting signal which depends on the residues at the -1 and -3 positions relative to farnesylated cysteine. Structure-guided mutation allows the generation of a low-affinity INPP5E mutant which loses exclusive ciliary localization. We postulate that the affinity to PDE6δ and the release by Arl2/3 in addition to a retention signal are the determinants for cargo sorting and enrichment at its destination.
authors.affiliation: Max Planck Institute of Molecular Physiology, Otto-Hahn-Strasse 11, 44227 Dortmund, Germany.
authors.forename: Eyad Kalawy
authors.initials: EK
authors.lastname: Fansa
authors.affiliation: Max Planck Institute of Molecular Physiology, Otto-Hahn-Strasse 11, 44227 Dortmund, Germany.
authors.forename: Stefanie Kristine
authors.initials: SK
authors.lastname: Kösling
authors.affiliation: Max Planck Institute of Molecular Physiology, Otto-Hahn-Strasse 11, 44227 Dortmund, Germany.
authors.forename: Eldar
authors.initials: E
authors.lastname: Zent
authors.affiliation: Max Planck Institute of Molecular Physiology, Otto-Hahn-Strasse 11, 44227 Dortmund, Germany.
authors.forename: Alfred
authors.initials: A
authors.lastname: Wittinghofer
authors.affiliation: CR-UK Beatson Institute, Garscube Estate Switchback Road, Glasgow G61 1BD, UK.
authors.forename: Shehab
authors.initials: S
authors.lastname: Ismail
mesh_terms: D020727:ADP-Ribosylation Factors / Q000737:chemistry / Q000378:metabolism; D000818:Animals; D002460:Cell Line; D002923:Cilia / Q000378:metabolism*; D054707:Cyclic Nucleotide Phosphodiesterases, Type 6 / Q000378:metabolism*; D005454:Fluorescence Polarization; D049452:Green Fluorescent Proteins / Q000378:metabolism; D006160:Guanosine Triphosphate / Q000378:metabolism; D000072181:Inositol Polyphosphate 5-Phosphatases; D007700:Kinetics; D051379:Mice; D008954:Models, Biological; D020559:Monomeric GTP-Binding Proteins / Q000378:metabolism; D050505:Mutant Proteins / Q000378:metabolism; D009479:Neuropeptides / Q000378:metabolism; D010744:Phosphoric Monoester Hydrolases / Q000378:metabolism*; D011485:Protein Binding; D017368:Protein Prenylation; D021382:Protein Sorting Signals; D017433:Protein Structure, Secondary; D021381:Protein Transport; D000076205:Ras Homolog Enriched in Brain Protein
chemical_list: D050505:Mutant Proteins; D009479:Neuropeptides; D021382:Protein Sorting Signals; D000076205:Ras Homolog Enriched in Brain Protein; C490213:Rheb protein, mouse; D049452:Green Fluorescent Proteins; D006160:Guanosine Triphosphate; D010744:Phosphoric Monoester Hydrolases; D000072181:Inositol Polyphosphate 5-Phosphatases; D054707:Cyclic Nucleotide Phosphodiesterases, Type 6; C490248:Arl3 protein, mouse; D020727:ADP-Ribosylation Factors; D020559:Monomeric GTP-Binding Proteins