TY - JOUR

T1 - Protein interacting with C kinase 1 (PICK1) reduces reinsertion rates of interaction partners sorted to Rab11-dependent slow recycling pathway

AU - Madsen, Kenneth Lindegaard

AU - Thorsen, Thor Seneca

AU - Rahbek-Clemmensen, Troels

AU - Eriksen, Jacob Møller

AU - Gether, Ulrik

PY - 2012/4/6

Y1 - 2012/4/6

N2 - The scaffolding protein PICK1 (protein interacting with C kinase 1) contains an N-terminal PSD-95/Discs large/ZO-1 (PDZ) domain and a central lipid-binding Bin/amphiphysin/Rvs (BAR) domain. PICK1 is thought to regulate trafficking of its PDZ binding partners but different and even opposing functions have been suggested. Here, we apply ELISA-based assays and confocal microscopy in HEK293 cells with inducible PICK1 expression to assess in an isolated system the ability of PICK1 to regulate trafficking of natural and engineered PDZ binding partners. The dopamine transporter (DAT), which primarily sorts to degradation upon internalization, did not form perinuclear clusters with PICK1, and PICK1 did not affect DAT internalization/recycling. However, transfer of the PICK1-binding DAT C terminus to the β(2)-adrenergic receptor, which sorts to recycling upon internalization, led to formation of PICK1 co-clusters in Rab11-positive compartments. Furthermore, PICK1 inhibited Rab11-mediated recycling of the receptor in a BAR and PDZ domain-dependent manner. In contrast, transfer of the DAT C terminus to the δ-opioid receptor, which sorts to degradation, did not result in PICK1 co-clusters or any change in internalization/recycling. Further support for a role of PICK1 determined by its PDZ cargo was obtained for the PICK1 interaction partner prolactin-releasing peptide receptor (GPR10). GPR10 co-localized with Rab11 and clustered with PICK1 upon constitutive internalization but co-localized with the late endosomal marker Rab7 and did not cluster with PICK1 upon agonist-induced internalization. Our data suggest a selective role of PICK1 in clustering and reducing the recycling rates of PDZ domain binding partners sorted to the Rab11-dependent recycling pathway.

AB - The scaffolding protein PICK1 (protein interacting with C kinase 1) contains an N-terminal PSD-95/Discs large/ZO-1 (PDZ) domain and a central lipid-binding Bin/amphiphysin/Rvs (BAR) domain. PICK1 is thought to regulate trafficking of its PDZ binding partners but different and even opposing functions have been suggested. Here, we apply ELISA-based assays and confocal microscopy in HEK293 cells with inducible PICK1 expression to assess in an isolated system the ability of PICK1 to regulate trafficking of natural and engineered PDZ binding partners. The dopamine transporter (DAT), which primarily sorts to degradation upon internalization, did not form perinuclear clusters with PICK1, and PICK1 did not affect DAT internalization/recycling. However, transfer of the PICK1-binding DAT C terminus to the β(2)-adrenergic receptor, which sorts to recycling upon internalization, led to formation of PICK1 co-clusters in Rab11-positive compartments. Furthermore, PICK1 inhibited Rab11-mediated recycling of the receptor in a BAR and PDZ domain-dependent manner. In contrast, transfer of the DAT C terminus to the δ-opioid receptor, which sorts to degradation, did not result in PICK1 co-clusters or any change in internalization/recycling. Further support for a role of PICK1 determined by its PDZ cargo was obtained for the PICK1 interaction partner prolactin-releasing peptide receptor (GPR10). GPR10 co-localized with Rab11 and clustered with PICK1 upon constitutive internalization but co-localized with the late endosomal marker Rab7 and did not cluster with PICK1 upon agonist-induced internalization. Our data suggest a selective role of PICK1 in clustering and reducing the recycling rates of PDZ domain binding partners sorted to the Rab11-dependent recycling pathway.

KW - Carrier Proteins

KW - Cell Line

KW - Dopamine Plasma Membrane Transport Proteins

KW - Endocytosis

KW - Humans

KW - Interleukin-2 Receptor alpha Subunit

KW - Nuclear Proteins

KW - Protein Binding

KW - Protein Interaction Domains and Motifs

KW - Protein Transport

KW - Receptors, AMPA

KW - Receptors, G-Protein-Coupled

KW - Receptors, Opioid

KW - Recombinant Fusion Proteins

KW - rab GTP-Binding Proteins

U2 - 10.1074/jbc.M111.294702

DO - 10.1074/jbc.M111.294702

M3 - Journal article

C2 - 22303009

VL - 287

SP - 12293

EP - 12308

JO - Journal of Biological Chemistry

JF - Journal of Biological Chemistry

SN - 0021-9258

IS - 15

ER -