title: Research resource: New and diverse substrates for the insulin receptor isoform A revealed by quantitative proteomics after stimulation with IGF-II or insulin.
abstract: The isoform A of the insulin receptor (IR) (IR-A) is a bifunctional receptor, because it binds both insulin and IGF-II. IR-A activation by IGF-II plays a role in development, but its physiological role in adults is unknown. IGF-II signaling through IR-A is deregulated in cancer and favors tumor progression. We hypothesized that IGF-II binding to the IR-A elicits a unique signaling pathway. In order to obtain an unbiased evaluation of IR-A substrates differentially involved after IGF-II and insulin stimulation, we performed quantitative proteomics of IR-A substrates recruited to tyrosine-phosphorylated protein complexes using stable isotope labeling with amino acids in cell culture in combination with antiphosphotyrosine antibody pull down and mass spectrometry. Using cells expressing only the human IR-A and lacking the IGF-I receptor, we identified 38 IR-A substrates. Only 10 were known IR mediators, whereas 28 substrates were not previously related to IR signaling. Eleven substrates were recruited by stimulation with both ligands: two equally recruited by IGF-II and insulin, three more strongly recruited by IGF-II, and six more strongly recruited by insulin. Moreover, 14 substrates were recruited solely by IGF-II and 13 solely by insulin stimulation. Interestingly, discoidin domain receptors, involved in cell migration and tumor metastasis, and ephrin receptor B4, involved in bidirectional signaling upon cell-cell contact, were predominantly activated by IGF-II. These findings indicate that IR-A activation by IGF-II elicits a unique signaling pathway that may play a distinct role in physiology and in disease.
authors.affiliation: Department of Clinical and Experimental Medicine, University Magna Graecia of Catanzaro, Catanzaro, Italy.
authors.forename: Alaide
authors.initials: A
authors.lastname: Morcavallo
authors.forename: Marco
authors.initials: M
authors.lastname: Gaspari
authors.forename: Giuseppe
authors.initials: G
authors.lastname: Pandini
authors.forename: Angela
authors.initials: A
authors.lastname: Palummo
authors.forename: Giovanni
authors.initials: G
authors.lastname: Cuda
authors.forename: Martin R
authors.initials: MR
authors.lastname: Larsen
authors.forename: Riccardo
authors.initials: R
authors.lastname: Vigneri
authors.forename: Antonino
authors.initials: A
authors.lastname: Belfiore
mesh_terms: D000818:Animals; D015153:Blotting, Western; D002460:Cell Line; D006801:Humans; D007328:Insulin / Q000494:pharmacology*; D007335:Insulin-Like Growth Factor II / Q000494:pharmacology*; D042541:Intracellular Space / Q000187:drug effects / Q000378:metabolism; D007553:Isotope Labeling; D051379:Mice; D008954:Models, Biological; D020033:Protein Isoforms / Q000378:metabolism; D040901:Proteomics / Q000379:methods*; D011972:Receptor, Insulin / Q000378:metabolism*; D015203:Reproducibility of Results; D015398:Signal Transduction / Q000187:drug effects; D013379:Substrate Specificity / Q000187:drug effects; D013997:Time Factors
chemical_list: D007328:Insulin; D020033:Protein Isoforms; D007335:Insulin-Like Growth Factor II; D011972:Receptor, Insulin