title: Distinct activation of epidermal growth factor receptor by UTP contributes to epithelial cell wound repair.
abstract: The release of nucleotides after injury activates purinergic receptors, leading to phosphorylation of site-specific residues on epidermal growth factor receptor (EGFR). To elucidate the differences between the injury-induced response and that induced by exogenous EGF, we examined recruitment of docking proteins, internalization of EGFR, and migration after injury. Injury induced by scratch wounds or stimulation by addition of UTP caused a brief internalization of EGFR, which paralleled the lesser association with growth factor receptor-bound protein 2 (Grb2) and phosphorylation of EGFR. The internalization caused by EGF was sustained and detected for longer than 60 minutes and correlated with phosphorylation of the receptor. The EGF caused recruitment of Grb2, phospholipase C-γ-1 (PLCγ1), Shc, and Src to EGFR. Glutathione S-transferase pull downs were performed, and glutathione S-transferase-PLCγ1 showed binding of Grb2 when stimulated with EGF but not with UTP or injury. Furthermore, UTP did not induce PLCγ1 phosphorylation, and the phosphorylation induced by EGF was attenuated by costimulation with UTP. The response to heparin-binding EGF was equivalent to that of EGF. Site-directed mutagenesis showed that phosphorylation of Y1068 and Y1086 of EGFR is required for repair. Together, our results show that injury and activation of purinergic receptors and direct activation of EGFR via EGF induce distinct downstream pathways.
authors.affiliation: Department of Biochemistry, Boston University School of Medicine, Boston, Massachusetts 02118, USA.
authors.forename: Ilene
authors.initials: I
authors.lastname: Boucher
authors.forename: Amanuel
authors.initials: A
authors.lastname: Kehasse
authors.forename: Meredith
authors.initials: M
authors.lastname: Marcincin
authors.forename: Celeste
authors.initials: C
authors.lastname: Rich
authors.forename: Nader
authors.initials: N
authors.lastname: Rahimi
authors.forename: Vickery
authors.initials: V
authors.lastname: Trinkaus-Randall
mesh_terms: D000818:Animals; D002465:Cell Movement / Q000187:drug effects; D003593:Cytoplasm / Q000187:drug effects / Q000378:metabolism; D004705:Endocytosis / Q000187:drug effects; D004789:Enzyme Activation / Q000187:drug effects; D004815:Epidermal Growth Factor / Q000494:pharmacology; D004847:Epithelial Cells / Q000201:enzymology* / Q000473:pathology*; D066246:ErbB Receptors / Q000378:metabolism*; D051380:GRB2 Adaptor Protein / Q000378:metabolism; D066257:Heparin-binding EGF-like Growth Factor; D006801:Humans; D036341:Intercellular Signaling Peptides and Proteins / Q000494:pharmacology; D050505:Mutant Proteins / Q000378:metabolism; D051966:Phospholipase C gamma / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D021381:Protein Transport / Q000187:drug effects; D011817:Rabbits; D034421:Sus scrofa; D014544:Uridine Triphosphate / Q000494:pharmacology*; D014945:Wound Healing / Q000187:drug effects*
chemical_list: D051380:GRB2 Adaptor Protein; C495948:GRB2 protein, human; C587883:HBEGF protein, human; D066257:Heparin-binding EGF-like Growth Factor; D036341:Intercellular Signaling Peptides and Proteins; D050505:Mutant Proteins; D004815:Epidermal Growth Factor; D066246:ErbB Receptors; D051966:Phospholipase C gamma; D014544:Uridine Triphosphate