title: Intersectin 1 enhances Cbl ubiquitylation of epidermal growth factor receptor through regulation of Sprouty2-Cbl interaction.
abstract: Ubiquitylation of receptor tyrosine kinases plays a critical role in regulating the trafficking and lysosomal degradation of these important signaling molecules. We identified the multidomain scaffolding protein intersectin 1 (ITSN1) as an important regulator of this process (N. P. Martin et al., Mol. Pharmacol. 70:1463-1653, 2006) ITSN1 stimulates ubiquitylation of the epidermal growth factor receptor (EGFR) through enhancing the activity of the Cbl E3 ubiquitin ligase. However, the precise mechanism through which ITSN1 enhances Cbl activity was unclear. In this study, we found that ITSN1 enhances Cbl activity through disrupting the interaction of Cbl with the Sprouty2 (Spry2) inhibitory protein. We demonstrate that ITSN1 binds Pro-rich regions in both Cbl and Spry2 and that interaction of ITSN1 with Spry2 disrupts Spry2-Cbl interaction, resulting in enhanced ubiquitylation of the EGFR. Disruption of ITSN1 binding to Spry2 through point mutation of the Pro-rich ITSN1 binding site in Spry2 results in enhanced Cbl-Spry2 interaction and inhibition of receptor ubiquitylation. This study demonstrates that ITSN1 enhances Cbl activity by modulating the interaction of Cbl with Spry2. In addition, our results reveal a new level of complexity in the regulation of Cbl through the interaction with ITSN1 and Spry2.
authors.affiliation: Departments of Pharmacology, University of Illinois at Chicago, Chicago, Illinois, USA.
authors.forename: Mustafa Nazir
authors.initials: MN
authors.lastname: Okur
authors.forename: Jolene
authors.initials: J
authors.lastname: Ooi
authors.forename: Chee Wai
authors.initials: CW
authors.lastname: Fong
authors.forename: Natalia
authors.initials: N
authors.lastname: Martinez
authors.forename: Carlota
authors.initials: C
authors.lastname: Garcia-Dominguez
authors.forename: Jose M
authors.initials: JM
authors.lastname: Rojas
authors.forename: Graeme
authors.initials: G
authors.lastname: Guy
authors.forename: John P
authors.initials: JP
authors.lastname: O'Bryan
mesh_terms: D048868:Adaptor Proteins, Signal Transducing; D033942:Adaptor Proteins, Vesicular Transport / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D019943:Amino Acid Substitution; D000818:Animals; D001483:Base Sequence; D019556:COS Cells; D002522:Chlorocebus aethiops; D017931:DNA Primers / Q000235:genetics; D066246:ErbB Receptors / Q000378:metabolism*; D057809:HEK293 Cells; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D016297:Mutagenesis, Site-Directed; D054730:Protein Interaction Domains and Motifs; D017346:Protein Serine-Threonine Kinases; D050721:Proto-Oncogene Proteins c-cbl / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D020798:Two-Hybrid System Techniques; D054875:Ubiquitination
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D033942:Adaptor Proteins, Vesicular Transport; D017931:DNA Primers; D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins; D011994:Recombinant Proteins; C485561:SPRY2 protein, human; C115919:intersectin 1; D050721:Proto-Oncogene Proteins c-cbl; D066246:ErbB Receptors; D017346:Protein Serine-Threonine Kinases; C118389:Spry2 protein, mouse; C067419:CBL protein, human