title: Modulation of focal adhesion constituents and their down-stream events by EGF: On the cross-talk of integrins and growth factor receptors.
abstract: Within the concept of integrin growth factor receptor (GFR) cross-talk, little is known about the effects of GFRs on focal adhesions (FAs). Therefore, we tested the hypothesis whether EGF can modulate constituents of FAs and subsequent down-stream events. To this end, EGF-treated keratinocytes were subjected to combined fluorescence imaging and western blotting, to quantify expression and/or activation of molecules, involved in integrin GFR cross-talk, and receptor proximal and distal signaling events. Generally, EGF response revealed an amplified redistribution or activation of molecules under study, which will be explained in detail from the plasma membrane to the cell interior. In addition to significant activation of EGF receptor (EGFR) at tyrosine Tyr845, a remarkable redistribution was detectable for the focal adhesion constituents, integrin ß1 and ß3, and zyxin. Increased activation also applied to focal adhesion kinase (FAK) by phosphorylation at Tyr397, Tyr576, and Src at Tyr418, while total FAK remained unchanged. Risen activity was seen as well for the analyzed distal down-stream events, p190RhoGAP and MAP kinases p42/44. Intriguingly, Src-specific inhibitor Herbimycin A abrogated the entire EGF response except FAK Tyr397 phosphorylation, independent of EGF presence. Mechanistically, our results show that EGF modulates adhesion in a dual fashion, by firstly redistributing focal adhesion constituents to adhesion sites, but also by amplifying levels of activated RhoA antagonist p190RhoGAP, important for cell motility. Further, the findings suggest that the observed EGF response underlies an EGFR integrin cross-talk under recruitment of receptor proximal FAK and Src, and MAP kinase and p190RhoGAP as receptor distal events.
authors.affiliation: University Eye Hospital Freiburg, Department for Ophthalmology, Germany.
authors.forename: Philipp
authors.initials: P
authors.lastname: Eberwein
authors.affiliation: University Medical Center Freiburg, Department of Oral Biotechnology, Germany.
authors.forename: Dougal
authors.initials: D
authors.lastname: Laird
authors.affiliation: University Medical Center Freiburg, Department of Oral Biotechnology, Germany.
authors.forename: Simon
authors.initials: S
authors.lastname: Schulz
authors.affiliation: University Eye Hospital Freiburg, Department for Ophthalmology, Germany.
authors.forename: Thomas
authors.initials: T
authors.lastname: Reinhard
authors.affiliation: University Medical Center Freiburg, Department of Oral Biotechnology, Germany.
authors.forename: Thorsten
authors.initials: T
authors.lastname: Steinberg
authors.affiliation: University Medical Center Freiburg, Department of Oral Biotechnology, Germany.
authors.forename: Pascal
authors.initials: P
authors.lastname: Tomakidi
mesh_terms: D002461:Cell Line, Transformed; D004815:Epidermal Growth Factor / Q000235:genetics / Q000378:metabolism*; D066246:ErbB Receptors / Q000235:genetics / Q000378:metabolism*; D051417:Focal Adhesion Kinase 1 / Q000235:genetics / Q000378:metabolism; D022001:Focal Adhesions / Q000235:genetics / Q000378:metabolism*; D020662:Guanine Nucleotide Exchange Factors / Q000235:genetics / Q000378:metabolism; D006801:Humans; D016023:Integrins / Q000235:genetics / Q000378:metabolism*; D015603:Keratinocytes / Q000166:cytology / Q000378:metabolism*; D020935:MAP Kinase Signaling System / Q000502:physiology*; D019950:Mitogen-Activated Protein Kinase 1 / Q000235:genetics / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000235:genetics / Q000378:metabolism; D020663:ras Guanine Nucleotide Exchange Factors
chemical_list: D020662:Guanine Nucleotide Exchange Factors; D016023:Integrins; C417242:RAPGEF5 protein, human; D020663:ras Guanine Nucleotide Exchange Factors; D004815:Epidermal Growth Factor; C512478:EGFR protein, human; D066246:ErbB Receptors; D051417:Focal Adhesion Kinase 1; C496050:PTK2 protein, human; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3
keywords: Cell motility; Epidermal growth factor; Focal adhesions; Growth factor receptors; Integrins; Mechanotransduction; Zyxin