Epidermal growth factor and thrombin induced proliferation of immortalized human keratinocytes is coupled to the synthesis of Egr-1, a zinc finger transcriptional regulator.
The epidermal growth factor (EGF) receptor is highly expressed in HaCaT keratinocytes as shown by Western blotting. Stimulation of HaCaT cells with EGF, and also with the serine protease thrombin, induced DNA synthesis, measured by incorporation of 5-bromo-2'-deoxyuridine into the DNA of proliferating cells. Using antibodies directed against the active form of the EGF receptor, we show that in HaCaT cells EGF and thrombin triggered a rapid activation of the EGF receptor, followed by the phosphorylation and activation of the extracellular signal-regulated protein kinase (ERK). Moreover, EGF and thrombin induced a transient synthesis of the zinc finger transcriptional regulator Egr-1. Proliferation, activation of ERK, and biosynthesis of Egr-1 was completely inhibited in EGF or thrombin-treated HaCaT cells by the MAP kinase kinase inhibitor PD98059 and by AG1487, an EGF receptor-specific tyrosine kinase inhibitor. These data indicate that phosphorylation and activation of both the EGF receptor and ERK are essential for mitogenic signaling via EGF and thrombin. The synthesis of Egr-1 in HaCaT cells as a result of EGF or thrombin stimulation suggests that Egr-1 may be an important "late" part of the EGF and thrombin-initiated signaling cascades. We postulate that Egr-1 may function as a "third messenger" in keratinocytes connecting mitogenic stimulation with changes in gene transcription.
Kaufmann
Katrin
K
Department of Medical Biochemistry and Molecular Biology, University of Saarland Medical Center, D-66421 Homburg, Germany.
Thiel
Gerald
G
D001973:Bromodeoxyuridine; D002455:Cell Division / Q000187:drug effects; D004268:DNA-Binding Proteins / Q000096:biosynthesis*; D051766:Early Growth Response Protein 1; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004815:Epidermal Growth Factor / Q000494:pharmacology*; D066246:ErbB Receptors / Q000378:metabolism; D006490:Hemostatics / Q000494:pharmacology*; D006801:Humans; D017874:Immediate-Early Proteins; D015603:Keratinocytes / Q000187:drug effects* / Q000378:metabolism; D020935:MAP Kinase Signaling System; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D010766:Phosphorylation; D013917:Thrombin / Q000494:pharmacology*; D014157:Transcription Factors / Q000096:biosynthesis*; D014407:Tumor Cells, Cultured / Q000187:drug effects / Q000378:metabolism; D016335:Zinc Fingers
D004268:DNA-Binding Proteins; C497325:EGR1 protein, human; D051766:Early Growth Response Protein 1; D004791:Enzyme Inhibitors; D006490:Hemostatics; D017874:Immediate-Early Proteins; D014157:Transcription Factors; D004815:Epidermal Growth Factor; D066246:ErbB Receptors; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases; D013917:Thrombin; D001973:Bromodeoxyuridine