title: Actin-Sorting Nexin 27 (SNX27)-Retromer Complex Mediates Rapid Parathyroid Hormone Receptor Recycling.
abstract: The G protein-coupled parathyroid hormone receptor (PTHR) regulates mineral-ion homeostasis and bone remodeling. Upon parathyroid hormone (PTH) stimulation, the PTHR internalizes into early endosomes and subsequently traffics to the retromer complex, a sorting platform on early endosomes that promotes recycling of surface receptors. The C terminus of the PTHR contains a type I PDZ ligand that binds PDZ domain-containing proteins. Mass spectrometry identified sorting nexin 27 (SNX27) in isolated endosomes as a PTHR binding partner. PTH treatment enriched endosomal PTHR. SNX27 contains a PDZ domain and serves as a cargo selector for the retromer complex. VPS26, VPS29, and VPS35 retromer subunits were isolated with PTHR in endosomes from cells stimulated with PTH. Molecular dynamics and protein binding studies establish that PTHR and SNX27 interactions depend on the PDZ recognition motif in PTHR and the PDZ domain of SNX27. Depletion of either SNX27 or VPS35 or actin depolymerization decreased the rate of PTHR recycling following agonist stimulation. Mutating the PDZ ligand of PTHR abolished the interaction with SNX27 but did not affect the overall rate of recycling, suggesting that PTHR may directly engage the retromer complex. Coimmunoprecipitation and overlay experiments show that both intact and mutated PTHR bind retromer through the VPS26 protomer and sequentially assemble a ternary complex with PTHR and SNX27. SNX27-independent recycling may involve N-ethylmaleimide-sensitive factor, which binds both PDZ intact and mutant PTHRs. We conclude that PTHR recycles rapidly through at least two pathways, one involving the ASRT complex of actin, SNX27, and retromer and another possibly involving N-ethylmaleimide-sensitive factor.
authors.affiliation: From the Laboratory for GPCR Biology, Department of Pharmacology and Chemical Biology, and.
authors.forename: Jennifer C
authors.initials: JC
authors.lastname: McGarvey
authors.affiliation: From the Laboratory for GPCR Biology, Department of Pharmacology and Chemical Biology, and.
authors.forename: Kunhong
authors.initials: K
authors.lastname: Xiao
authors.affiliation: the Department of Biological Sciences, Carnegie Mellon University, Pittsburgh, Pennsylvania 15213.
authors.forename: Shanna L
authors.initials: SL
authors.lastname: Bowman
authors.affiliation: From the Laboratory for GPCR Biology, Department of Pharmacology and Chemical Biology, and.
authors.forename: Tatyana
authors.initials: T
authors.lastname: Mamonova
authors.affiliation: From the Laboratory for GPCR Biology, Department of Pharmacology and Chemical Biology, and.
authors.forename: Qiangmin
authors.initials: Q
authors.lastname: Zhang
authors.affiliation: From the Laboratory for GPCR Biology, Department of Pharmacology and Chemical Biology, and.
authors.forename: Alessandro
authors.initials: A
authors.lastname: Bisello
authors.affiliation: From the Laboratory for GPCR Biology, Department of Pharmacology and Chemical Biology, and.
authors.forename: W Bruce
authors.initials: WB
authors.lastname: Sneddon
authors.affiliation: From the Laboratory for GPCR Biology, Department of Pharmacology and Chemical Biology, and.
authors.forename: Juan A
authors.initials: JA
authors.lastname: Ardura
authors.affiliation: From the Laboratory for GPCR Biology, Department of Pharmacology and Chemical Biology, and.
authors.forename: Frederic
authors.initials: F
authors.lastname: Jean-Alphonse
authors.affiliation: From the Laboratory for GPCR Biology, Department of Pharmacology and Chemical Biology, and.
authors.forename: Jean-Pierre
authors.initials: JP
authors.lastname: Vilardaga
authors.affiliation: the Department of Biological Sciences, Carnegie Mellon University, Pittsburgh, Pennsylvania 15213.
authors.forename: Manojkumar A
authors.initials: MA
authors.lastname: Puthenveedu
authors.affiliation: From the Laboratory for GPCR Biology, Department of Pharmacology and Chemical Biology, and the Department of Structural Biology, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania 15261 and paf10@pitt.edu.
authors.forename: Peter A
authors.initials: PA
authors.lastname: Friedman
mesh_terms: D000199:Actins / Q000737:chemistry / Q000378:metabolism*; D000818:Animals; D016466:CHO Cells; D003412:Cricetulus; D011992:Endosomes / Q000378:metabolism; D057809:HEK293 Cells; D006801:Humans; D053858:Metabolic Networks and Pathways; D056004:Molecular Dynamics Simulation; D046912:Multiprotein Complexes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D052066:N-Ethylmaleimide-Sensitive Proteins / Q000378:metabolism; D054731:PDZ Domains; D011485:Protein Binding; D021122:Protein Subunits; D021381:Protein Transport; D059748:Proteolysis; D044168:Receptor, Parathyroid Hormone, Type 1 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D058305:Sorting Nexins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
chemical_list: D000199:Actins; D046912:Multiprotein Complexes; D021122:Protein Subunits; D044168:Receptor, Parathyroid Hormone, Type 1; D011994:Recombinant Proteins; C522885:SNX27 protein, human; D058305:Sorting Nexins; D052066:N-Ethylmaleimide-Sensitive Proteins
keywords: G protein-coupled receptor (GPCR); endosome; parathyroid hormone; receptor recycling; sorting nexin (SNX)