title: ESCRT-0 protein hepatocyte growth factor-regulated tyrosine kinase substrate (Hrs) is targeted to endosomes independently of signal-transducing adaptor molecule (STAM) and the complex formation with STAM promotes its endosomal dissociation.
abstract: The ESCRT-0 complex, consisting of the hepatocyte growth factor-regulated tyrosine kinase substrate (Hrs) and the signal-transducing adaptor molecule (STAM) proteins, recognizes ubiquitylated cargo during the initial step of endosomal sorting. The endosomal accumulation of overexpressed Hrs has been reported previously to be associated with endosome enlargement. In this study, we have found that co-expressing exogenous STAM1 in Hrs-overexpressing cells leads to a diffuse localization for a large part of the Hrs accumulated on endosomes and a recovery of the impaired cargo protein degradation process, thus suggesting that exogenous STAM abrogates the abnormalities of the Hrs-positive endosomes. A fluorescently labeled Hrs, introduced into the cells by membrane permeabilization, exhibited endosomal localization in the absence of STAM1 and gradually dissociated from the endosomes upon the sequential addition of recombinant STAM1. Furthermore, when microinjected into cells, the fluorescently labeled Hrs also showed endosomal accumulation; however, ESCRT-0 complexes formed prior to the microinjection did not. Analysis of the state of the complex in HeLa cells using blue-native PAGE revealed that the membrane-associated Hrs exists partly as a monomer and not only in the STAM1-bound form. Thus, our data suggest that the membrane binding and dissociation cycle of the ESCRT-0 proteins on the endosomal membrane is a critical step during the cargo sorting process.
authors.affiliation: From the Department of Microbiology and Immunology, Shinshu University School of Medicine, 3-1-1 Asahi, Matsumoto, Nagano 390-8621 and.
authors.forename: Katsuhiko
authors.initials: K
authors.lastname: Kojima
authors.affiliation: From the Department of Microbiology and Immunology, Shinshu University School of Medicine, 3-1-1 Asahi, Matsumoto, Nagano 390-8621 and.
authors.forename: Yuji
authors.initials: Y
authors.lastname: Amano
authors.affiliation: From the Department of Microbiology and Immunology, Shinshu University School of Medicine, 3-1-1 Asahi, Matsumoto, Nagano 390-8621 and.
authors.forename: Kazuhisa
authors.initials: K
authors.lastname: Yoshino
authors.affiliation: the Divisions of Immunology and.
authors.forename: Nobuyuki
authors.initials: N
authors.lastname: Tanaka
authors.affiliation: Molecular and Cellular Oncology, Miyagi Cancer Center Research Institute, 47-1 Nodayama, Medeshima-Shiode, Natori 981-1293, Japan.
authors.forename: Kazuo
authors.initials: K
authors.lastname: Sugamura
authors.affiliation: From the Department of Microbiology and Immunology, Shinshu University School of Medicine, 3-1-1 Asahi, Matsumoto, Nagano 390-8621 and takesit@shinshu-u.ac.jp.
authors.forename: Toshikazu
authors.initials: T
authors.lastname: Takeshita
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D056827:Endosomal Sorting Complexes Required for Transport / Q000235:genetics / Q000378:metabolism*; D011992:Endosomes / Q000235:genetics / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D051379:Mice; D018345:Mice, Knockout; D010750:Phosphoproteins / Q000235:genetics / Q000378:metabolism*; D021381:Protein Transport / Q000502:physiology
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D056827:Endosomal Sorting Complexes Required for Transport; D010750:Phosphoproteins; C109775:STAM protein, human; C491648:Stam protein, mouse; C096164:hepatocyte growth factor-regulated tyrosine kinase substrate
keywords: ESCRT-0; Endosome; Hrs; Membrane Trafficking; Protein Sorting; Protein Targeting; STAM; Ubiquitylation (Ubiquitination)