Intersectin regulates epidermal growth factor receptor endocytosis, ubiquitylation, and signaling.
Receptor tyrosine kinases (RTKs) are critical for normal cell growth, differentiation, and development, but they contribute to various pathological conditions when disrupted. Activation of RTKs stimulates a plethora of pathways, including the ubiquitylation and endocytosis of the receptor itself. Although endocytosis terminates RTK signaling, it has emerged as a requisite step in RTK activation of signaling pathways. We have discovered that the endocytic scaffolding protein intersectin (ITSN) cooperated with epidermal growth factor receptor (EGFR) in the regulation of cell growth and signaling. However, a biochemical link between ITSN and EGFR was not defined. In this study, we demonstrate that ITSN is a scaffold for the E3 ubiquitin ligase Cbl. ITSN forms a complex with Cbl in vivo mediated by the Src homology (SH) 3 domains binding to the Pro-rich COOH terminus of Cbl. This interaction stimulates the ubiquitylation and degradation of the activated EGFR. Furthermore, silencing ITSN by RNA interference attenuated EGFR internalization as well as activation of the extracellular signal-regulated kinasemitogen-activated protein kinase pathway, thereby demonstrating the importance of ITSN in EGFR function. Given the cooperativity between ITSN and additional RTKs, these results point to an important evolutionarily conserved, regulatory role for ITSN in RTK function that is necessary for both signaling from receptors as well as the ultimate termination of receptor signaling.
Martin
Negin P
NP
Laboratory of Signal Transduction, National Institute of Environmental Health Sciences, National Institutes of Health, Research Triangle Park, North Carolina, USA.
Mohney
Robert P
RP
Dunn
Sara
S
Das
Margaret
M
Scappini
Erica
E
O'Bryan
John P
JP
D033942:Adaptor Proteins, Vesicular Transport / Q000378:metabolism*; D000818:Animals; D019556:COS Cells; D002522:Chlorocebus aethiops; D022162:Cytoplasmic Vesicles / Q000187:drug effects; D004705:Endocytosis / Q000187:drug effects / Q000502:physiology*; D004815:Epidermal Growth Factor / Q000494:pharmacology; D066246:ErbB Receptors / Q000378:metabolism*; D020868:Gene Silencing; D006801:Humans; D047468:Immunoprecipitation; D008954:Models, Biological; D011485:Protein Binding / Q000187:drug effects; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D021381:Protein Transport / Q000187:drug effects; D050721:Proto-Oncogene Proteins c-cbl / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D025801:Ubiquitin / Q000378:metabolism*; D014981:Xenopus
D033942:Adaptor Proteins, Vesicular Transport; C485568:ITSN1 protein, human; D025801:Ubiquitin; D004815:Epidermal Growth Factor; D050721:Proto-Oncogene Proteins c-cbl; D066246:ErbB Receptors