Ubiquitination regulates proteolytic processing of G protein-coupled receptors after their sorting to lysosomes. J Biol Chem 2009 Jul 17;284(29):19361-70
Date
05/13/2009Pubmed ID
19433584Pubmed Central ID
PMC2740561DOI
10.1074/jbc.M109.001644Scopus ID
2-s2.0-67749127754 (requires institutional sign-in at Scopus site) 76 CitationsAbstract
Ubiquitination is essential for the endocytic sorting of various G protein-coupled receptors to lysosomes. Here we identify a distinct function of this covalent modification in controlling the later proteolytic processing of receptors. Mutation of all cytoplasmic lysine residues in the murine delta-opioid receptor blocked receptor ubiquitination without preventing ligand-induced endocytosis of receptors or their subsequent delivery to lysosomes, as verified by proteolysis of extramembrane epitope tags and down-regulation of radioligand binding to the transmembrane helices. Surprisingly, a functional screen revealed that the E3 ubiquitin ligase AIP4 specifically controls down-regulation of wild type receptors measured by radioligand binding without detectably affecting receptor delivery to lysosomes defined both immunochemically and biochemically. This specific AIP4-dependent regulation required direct ubiquitination of receptors and was also regulated by two deubiquitinating enzymes, AMSH and UBPY, which localized to late endosome/lysosome membranes containing internalized delta-opioid receptor. These results identify a distinct function of AIP4-dependent ubiquitination in controlling the later proteolytic processing of G protein-coupled receptors, without detectably affecting their endocytic sorting to lysosomes. We propose that ubiquitination or ubiquitination/deubiquitination cycling specifically regulates later proteolytic processing events required for destruction of the receptor's hydrophobic core.
Author List
Hislop JN, Henry AG, Marchese A, von Zastrow MAuthor
Adriano Marchese PhD Professor in the Biochemistry department at Medical College of WisconsinMESH terms used to index this publication - Major topics in bold
Binding, CompetitiveBiotinylation
Cell Line
Electrophoresis, Polyacrylamide Gel
Endosomes
Green Fluorescent Proteins
Humans
Immunoblotting
Lysosomes
Microscopy, Fluorescence
Models, Biological
Mutation
Protein Processing, Post-Translational
Protein Transport
Radioligand Assay
Receptors, G-Protein-Coupled
Recombinant Fusion Proteins
Transfection
Ubiquitin
Ubiquitin-Protein Ligases
Ubiquitination