PDE6δ-mediated sorting of INPP5E into the cilium is determined by cargo-carrier affinity

Nat Commun. 2016 Apr 11;7:11366. doi: 10.1038/ncomms11366.

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.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • ADP-Ribosylation Factors / chemistry
  • ADP-Ribosylation Factors / metabolism
  • Animals
  • Cell Line
  • Cilia / metabolism*
  • Cyclic Nucleotide Phosphodiesterases, Type 6 / metabolism*
  • Fluorescence Polarization
  • Green Fluorescent Proteins / metabolism
  • Guanosine Triphosphate / metabolism
  • Inositol Polyphosphate 5-Phosphatases
  • Kinetics
  • Mice
  • Models, Biological
  • Monomeric GTP-Binding Proteins / metabolism
  • Mutant Proteins / metabolism
  • Neuropeptides / metabolism
  • Phosphoric Monoester Hydrolases / metabolism*
  • Protein Binding
  • Protein Prenylation
  • Protein Sorting Signals
  • Protein Structure, Secondary
  • Protein Transport
  • Ras Homolog Enriched in Brain Protein

Substances

  • Mutant Proteins
  • Neuropeptides
  • Protein Sorting Signals
  • Ras Homolog Enriched in Brain Protein
  • Rheb protein, mouse
  • Green Fluorescent Proteins
  • Guanosine Triphosphate
  • Phosphoric Monoester Hydrolases
  • Inositol Polyphosphate 5-Phosphatases
  • Cyclic Nucleotide Phosphodiesterases, Type 6
  • Arl3 protein, mouse
  • ADP-Ribosylation Factors
  • Monomeric GTP-Binding Proteins