title: Structural requirements for interactions between leucine-sorting signals and clathrin-associated adaptor protein complex AP3.
abstract: Cytoplasmic tails of LIMPII and the invariant chain contain similar leucine-based sorting signals, but the invariant chain interacts only with AP1 and AP2, whereas LIMPII interacts strongly with AP3. In a series of in vitro experiments, we investigated the effect of residues upstream of the leucine pairs and demonstrated that these residues determine adapter binding, and certain residues favor interactions with AP3. Furthermore, constructs that interacted stronger with AP3 interacted weakly with AP1 and vice versa. Exchanging residues upstream of the leucine-based signal in LIMPII with those of the invariant chain reduced LIMPII binding to AP3 in vitro, and in vivo the corresponding LIMPII mutant was rerouted via the plasma membrane like the invariant chain. These preferential interactions of different leucine signals with different AP complexes may thus be the determining step sorting proteins from the trans-Golgi network to their final destinations. Proteins that interact with AP3 are sorted directly to endosomes/lysosomes, whereas proteins that interact with AP1 are sorted via a different route. At the same time, constructs that exhibited specificity for either AP1 or AP3 might still interact with AP2, suggesting that AP2 may recognize a wider variety of leucine signals. This is consistent with the suggested role of AP2 in internalization of proteins containing general leucine-based signals, including proteins that have been missorted to the plasma membrane.
authors.affiliation: Division of Molecular Cell Biology, Department of Biology, University of Oslo, Norway.
authors.forename: Dmitrii G
authors.initials: DG
authors.lastname: Rodionov
authors.forename: Stefan
authors.initials: S
authors.lastname: Höning
authors.forename: Aleksandra
authors.initials: A
authors.lastname: Silye
authors.forename: Thomas L
authors.initials: TL
authors.lastname: Kongsvik
authors.forename: Kurt
authors.initials: K
authors.lastname: von Figura
authors.forename: Oddmund
authors.initials: O
authors.lastname: Bakke
mesh_terms: D033961:Adaptor Protein Complex 1 / Q000378:metabolism*; D033962:Adaptor Protein Complex 2 / Q000378:metabolism*; D033963:Adaptor Protein Complex 3 / Q000737:chemistry* / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D002966:Clathrin / Q000378:metabolism*; D004247:DNA / Q000378:metabolism; D004285:Dogs; D005982:Glutathione Transferase / Q000378:metabolism; D007700:Kinetics; D007930:Leucine / Q000737:chemistry* / Q000378:metabolism; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D009154:Mutation; D011233:Precipitin Tests; D011485:Protein Binding; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D020349:Surface Plasmon Resonance; D014162:Transfection
chemical_list: D033961:Adaptor Protein Complex 1; D033962:Adaptor Protein Complex 2; D033963:Adaptor Protein Complex 3; D002966:Clathrin; D011993:Recombinant Fusion Proteins; D004247:DNA; D005982:Glutathione Transferase; D007930:Leucine