title: Flat clathrin coats on endosomes mediate degradative protein sorting by scaffolding Hrs in dynamic microdomains.
abstract: Endocytosed membrane proteins that are destined for degradation in lysosomes are ubiquitylated and recognised by sorting complexes on endosome membranes. The ubiquitin-binding sorting component Hrs as well as ubiquitylated cargo are enriched in a characteristic flat clathrin coat on the endosome membrane. The function of clathrin within this coat has not been investigated. Here, we show that both clathrin and the clathrin-box motif of Hrs are required for the clustering of Hrs into restricted microdomains. The C-terminus of Hrs, which contains the clathrin-box, is sufficient to redirect a phosphatidylinositol(3)-phosphate-binding protein into the Hrs- and clathrin-containing microdomains. Although these microdomains show little lateral diffusion in the membrane, they are dynamic structures that exchange Hrs and clathrin with similar kinetics, and acquire the downstream sorting component Tsg101. The clathrin-mediated clustering is essential for the function of Hrs in degradative protein sorting. We conclude that clathrin is responsible for concentrating Hrs in endosomal microdomains specialised for recognition of ubiquitylated membrane proteins, thus enabling efficient sorting of cargo into the degradative pathway.
authors.affiliation: Department of Biochemistry, Institute for Cancer Research, The Norwegian Radium Hospital and The University of Oslo, Montebello, N-0310 Oslo, Norway.
authors.forename: Camilla
authors.initials: C
authors.lastname: Raiborg
authors.forename: Jørgen
authors.initials: J
authors.lastname: Wesche
authors.forename: Lene
authors.initials: L
authors.lastname: Malerød
authors.forename: Harald
authors.initials: H
authors.lastname: Stenmark
mesh_terms: D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism / Q000648:ultrastructure; D002478:Cells, Cultured; D002966:Clathrin / Q000378:metabolism*; D022163:Clathrin-Coated Vesicles / Q000378:metabolism / Q000648:ultrastructure; D004268:DNA-Binding Proteins / Q000378:metabolism; D056827:Endosomal Sorting Complexes Required for Transport; D011992:Endosomes / Q000378:metabolism* / Q000648:ultrastructure; D004815:Epidermal Growth Factor / Q000378:metabolism; D066246:ErbB Receptors / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D007700:Kinetics; D021962:Membrane Microdomains / Q000378:metabolism* / Q000648:ultrastructure; D008565:Membrane Proteins / Q000378:metabolism; D010750:Phosphoproteins / Q000378:metabolism*; D021381:Protein Transport; D013997:Time Factors; D014157:Transcription Factors / Q000378:metabolism; D033921:Vesicular Transport Proteins / Q000378:metabolism; D000099177:Tsg101 Protein
chemical_list: D002966:Clathrin; D004268:DNA-Binding Proteins; D056827:Endosomal Sorting Complexes Required for Transport; D004815:Epidermal Growth Factor; D066246:ErbB Receptors; D008565:Membrane Proteins; D010750:Phosphoproteins; D014157:Transcription Factors; D033921:Vesicular Transport Proteins; D000099177:Tsg101 Protein; C093846:early endosome antigen 1; C096164:hepatocyte growth factor-regulated tyrosine kinase substrate