title: Identification of a novel proline-arginine motif involved in CIN85-dependent clustering of Cbl and down-regulation of epidermal growth factor receptors.
abstract: CIN85 is a multidomain adaptor protein implicated in Cbl-mediated down-regulation of receptor tyrosine kinases. CIN85 binding to Cbl is increased after growth factor stimulation and is critical for targeting receptor tyrosine kinases to clathrin-mediated endocytosis. Here we report the identification of a novel polyproline-arginine motif (PXXXPR), specifically recognized by the SH3 domains of CIN85 and its homologue CMS/CD2AP. This motif was indispensable for CIN85 binding to Cbl/Cbl-b, to other CIN85 SH3 domains' effectors, and for mediating an intramolecular interaction between the SH3-A domain and the proline-rich region of CIN85. Individual SH3 domains of CIN85 bound to PXXXPR peptides of Cbl/Cbl-b with micromolar affinities, whereas an extended structure of two or three SH3 domains bound with higher stoichiometry and increased affinity to the same peptides. This enabled full size CIN85 to simultaneously interact with multiple Cbl molecules, promoting their clustering in mammalian cells. The ability of CIN85 to cluster Cbl was important for ligand-induced stabilization of CIN85.Cbl.epidermal growth factor receptor complexes, as well as for epidermal growth factor receptor degradation in the lysosome. Thus, specific interactions of CIN85 SH3 domains with the PXXXPR motif in Cbl play multiple roles in down-regulation of receptor tyrosine kinases.
authors.affiliation: Institute of Biochemistry II, University of Frankfurt Medical School, Theodor-Stern-Kai 7, Frankfurt 60590, Germany.
authors.forename: Katarzyna
authors.initials: K
authors.lastname: Kowanetz
authors.forename: Iwona
authors.initials: I
authors.lastname: Szymkiewicz
authors.forename: Kaisa
authors.initials: K
authors.lastname: Haglund
authors.forename: Marcin
authors.initials: M
authors.lastname: Kowanetz
authors.forename: Koraljka
authors.initials: K
authors.lastname: Husnjak
authors.forename: Jonathan D
authors.initials: JD
authors.lastname: Taylor
authors.forename: Philippe
authors.initials: P
authors.lastname: Soubeyran
authors.forename: Ulla
authors.initials: U
authors.lastname: Engstrom
authors.forename: John E
authors.initials: JE
authors.lastname: Ladbury
authors.forename: Ivan
authors.initials: I
authors.lastname: Dikic
mesh_terms: D048868:Adaptor Proteins, Signal Transducing; D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D001120:Arginine / Q000737:chemistry; D001665:Binding Sites; D016466:CHO Cells; D002352:Carrier Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D016384:Consensus Sequence; D006224:Cricetinae; D015536:Down-Regulation; D004705:Endocytosis; D066246:ErbB Receptors / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D066298:In Vitro Techniques; D008969:Molecular Sequence Data; D050722:Oncogene Protein v-cbl; D010750:Phosphoproteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D011392:Proline / Q000737:chemistry; D050721:Proto-Oncogene Proteins c-cbl; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D015682:Retroviridae Proteins, Oncogenic / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D044767:Ubiquitin-Protein Ligases; D018909:src Homology Domains
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D002352:Carrier Proteins; D050722:Oncogene Protein v-cbl; D010750:Phosphoproteins; D011994:Recombinant Proteins; D015682:Retroviridae Proteins, Oncogenic; C433810:SH3KBP1 protein, human; D001120:Arginine; D011392:Proline; C492211:CBLB protein, human; D050721:Proto-Oncogene Proteins c-cbl; D044767:Ubiquitin-Protein Ligases; D066246:ErbB Receptors