Hrs recruits clathrin to early endosomes.
The hepatocyte growth factor-regulated tyrosine kinase substrate, Hrs, has been implicated in intracellular trafficking and signal transduction. Hrs contains a phosphatidylinositol 3-phosphate-binding FYVE domain that contributes to its endosomal targeting. Here we show that Hrs and EEA1, a FYVE domain protein involved in endocytic membrane fusion, are localized to different regions of early endosomes. We demonstrate that Hrs co-localizes with clathrin, and that the C-terminus of Hrs contains a functional clathrin box motif that interacts directly with the terminal beta-propeller domain of clathrin heavy chain. A massive recruitment of clathrin to early endosomes was observed in cells transfected with Hrs, but not with Hrs lacking the C-terminus. Furthermore, the phosphatidylinositol 3-kinase inhibitor wortmannin caused the dissociation of both Hrs and clathrin from endosomes. While overexpression of Hrs did not affect endocytosis and recycling of transferrin, endocytosed epidermal growth factor and dextran were retained in early endosomes. These results provide a molecular mechanism for the recruitment of clathrin onto early endosomes and suggest a function for Hrs in trafficking from early to late endosomes.
Raiborg
C
C
Department of Biochemistry, Institute for Cancer Research, The Norwegian Radium Hospital, Montebello, N-0310 Oslo, Norway.
Bache
K G
KG
Mehlum
A
A
Stang
E
E
Stenmark
H
H
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002460:Cell Line; D002966:Clathrin / Q000737:chemistry* / Q000378:metabolism / Q000502:physiology*; D006224:Cricetinae; D003911:Dextrans / Q000378:metabolism; D004705:Endocytosis; D056827:Endosomal Sorting Complexes Required for Transport; D011992:Endosomes / Q000502:physiology* / Q000648:ultrastructure; D004815:Epidermal Growth Factor / Q000378:metabolism; D046911:Macromolecular Substances; D008545:Melanoma; D018129:Phosphatidylinositol Phosphates / Q000378:metabolism; D010750:Phosphoproteins / Q000737:chemistry / Q000378:metabolism / Q000502:physiology*; D011487:Protein Conformation; D021122:Protein Subunits; D021381:Protein Transport; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D014168:Transferrin / Q000378:metabolism; D014407:Tumor Cells, Cultured
D002966:Clathrin; D003911:Dextrans; D056827:Endosomal Sorting Complexes Required for Transport; D046911:Macromolecular Substances; D018129:Phosphatidylinositol Phosphates; D010750:Phosphoproteins; D021122:Protein Subunits; D014168:Transferrin; C096164:hepatocyte growth factor-regulated tyrosine kinase substrate; C055525:phosphatidylinositol 3-phosphate; D004815:Epidermal Growth Factor