title: Differentiation of opioid drug effects by hierarchical multi-site phosphorylation.
abstract: Differences in the ability of opioid drugs to promote regulated endocytosis of μ-opioid receptors are related to their tendency to produce drug tolerance and dependence. Here we show that drug-specific differences in receptor internalization are determined by a conserved, 10-residue sequence in the receptor's carboxyl-terminal cytoplasmic tail. Diverse opioids induce receptor phosphorylation at serine (S)375, present in the middle of this sequence, but opioids differ markedly in their ability to drive higher-order phosphorylation on flanking residues [threonine (T)370, T376, and T379]. Multi-phosphorylation is required for the endocytosis-promoting activity of this sequence and occurs both sequentially and hierarchically, with S375 representing the initiating site. Higher-order phosphorylation involving T370, T376, and T379 specifically requires GRK2/3 isoforms, and the same sequence controls opioid receptor internalization in neurons. These results reveal a biochemical mechanism differentiating the endocytic activity of opioid drugs.
authors.affiliation: Institute of Pharmacology and Toxicology, Jena University Hospital, Friedrich Schiller University Jena, Drackendorfer Straße 1, D-07747 Jena, Germany. Stefan.Schulz@mti.uni-jena.de
authors.forename: Sascha
authors.initials: S
authors.lastname: Just
authors.forename: Susann
authors.initials: S
authors.lastname: Illing
authors.forename: Michelle
authors.initials: M
authors.lastname: Trester-Zedlitz
authors.forename: Elaine K
authors.initials: EK
authors.lastname: Lau
authors.forename: Sarah J
authors.initials: SJ
authors.lastname: Kotowski
authors.forename: Elke
authors.initials: E
authors.lastname: Miess
authors.forename: Anika
authors.initials: A
authors.lastname: Mann
authors.forename: Christian
authors.initials: C
authors.lastname: Doll
authors.forename: Jonathan C
authors.initials: JC
authors.lastname: Trinidad
authors.forename: Alma L
authors.initials: AL
authors.lastname: Burlingame
authors.forename: Mark
authors.initials: M
authors.lastname: von Zastrow
authors.forename: Stefan
authors.initials: S
authors.lastname: Schulz
mesh_terms: D000701:Analgesics, Opioid / Q000494:pharmacology*; D000818:Animals; D004705:Endocytosis / Q000187:drug effects; D054769:G-Protein-Coupled Receptor Kinase 2 / Q000378:metabolism; D054770:G-Protein-Coupled Receptor Kinase 3 / Q000378:metabolism; D057809:HEK293 Cells; D006801:Humans; D051379:Mice; D009474:Neurons / Q000187:drug effects / Q000378:metabolism; D010766:Phosphorylation; D020033:Protein Isoforms / Q000378:metabolism; D011957:Receptors, Opioid / Q000378:metabolism*; D012694:Serine / Q000378:metabolism; D013912:Threonine / Q000378:metabolism
chemical_list: D000701:Analgesics, Opioid; D020033:Protein Isoforms; D011957:Receptors, Opioid; D013912:Threonine; D012694:Serine; D054770:G-Protein-Coupled Receptor Kinase 3; C496567:GRK2 protein, human; C517848:GRK3 protein, human; D054769:G-Protein-Coupled Receptor Kinase 2