CIN85 participates in Cbl-b-mediated down-regulation of receptor tyrosine kinases.
The Cbl family of ubiquitin ligases in mammals contains three members, Cbl, Cbl-b, and Cbl-3, that are involved in down-regulation of receptor tyrosine kinases (RTKs) by mediating receptor ubiquitination and degradation. More recently, a novel pathway has been identified whereby Cbl promotes internalization of EGF receptor via a CIN85/endophilin pathway that is functionally separable from the ubiquitin ligase activity of Cbl (1). Here we show that Cbl-b, but not Cbl-3, utilize the same mechanism to down-regulate multiple RTKs. CIN85 was shown to bind to the minimal binding domain identified in the carboxyl terminus of Cbl-b. Ligand-induced phosphorylation of Cbl-b further increased their interactions and led to a rapid and sustained recruitment of CIN85 in the complex with EGF or PDGF receptors. Inhibition of binding between CIN85 and Cbl-b was sufficient to impair Cbl-b-mediated internalization of EGF receptors, while being dispensable for Cbl-b-directed polyubiquitination of EGF receptors. Moreover, CIN85 and Cbl/Cbl-b were constitutively associated with activated PDGF, EGF, or c-Kit receptors in several tumor cell lines. Our data reveal a common pathway utilized by Cbl and Cbl-b that may have an important and redundant function in negative regulation of ligand-activated as well as oncogenically activated RTKs in vivo.
Szymkiewicz
Iwona
I
Ludwig Institute for Cancer Research, Box 595, Husargatan 3, Uppsala, S-75124, Sweden.
Kowanetz
Katarzyna
K
Soubeyran
Philippe
P
Dinarina
Ana
A
Lipkowitz
Stanley
S
Dikic
Ivan
I
D016475:3T3 Cells; D048868:Adaptor Proteins, Signal Transducing; D000818:Animals; D016466:CHO Cells; D002352:Carrier Proteins / Q000378:metabolism*; D002460:Cell Line; D002461:Cell Line, Transformed; D003001:Cloning, Molecular; D006224:Cricetinae; D015536:Down-Regulation; D004705:Endocytosis; D004815:Epidermal Growth Factor / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D008024:Ligands; D051379:Mice; D008856:Microscopy, Fluorescence; D008954:Models, Biological; D010750:Phosphoproteins / Q000378:metabolism*; D010766:Phosphorylation; D010982:Platelet-Derived Growth Factor / Q000378:metabolism; D011233:Precipitin Tests; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D050721:Proto-Oncogene Proteins c-cbl; D019009:Proto-Oncogene Proteins c-kit / Q000378:metabolism; D020794:Receptor Protein-Tyrosine Kinases / Q000378:metabolism*; D013997:Time Factors; D014162:Transfection; D020798:Two-Hybrid System Techniques; D044767:Ubiquitin-Protein Ligases
D048868:Adaptor Proteins, Signal Transducing; D002352:Carrier Proteins; C118387:Cblb protein, mouse; D008024:Ligands; D010750:Phosphoproteins; D010982:Platelet-Derived Growth Factor; C433810:SH3KBP1 protein, human; D004815:Epidermal Growth Factor; C492211:CBLB protein, human; D050721:Proto-Oncogene Proteins c-cbl; D044767:Ubiquitin-Protein Ligases; D019009:Proto-Oncogene Proteins c-kit; D020794:Receptor Protein-Tyrosine Kinases