Insulin-like growth factor-1 regulates endogenous RUNX2 activity in endothelial cells through a phosphatidylinositol 3-kinase/ERK-dependent and Akt-independent signaling pathway.
Insulin-like growth factor-1 (IGF-1) is an angiogenic and oncogenic factor that activates signal transduction pathways involved in the expression of transcriptional regulators of tumorigenesis. RUNX2, a member of the Ig-loop family of transcription factors is expressed in vascular endothelial cells (EC) and regulates EC migration, invasion, and proliferation. Here we show that IGF-1 and its receptor regulate post-translational changes in RUNX2 to activate DNA binding in proliferating EC. The phosphatidylinositol 3-kinase (PI3K) inhibitor, LY294002, reduced both basal and IGF-1-stimulated RUNX2 DNA binding activity in the absence of changes in RUNX2 protein as did the overexpression of the phosphatidylinositol 3-phosphate phosphatase, confirming that PI3K signaling mediates RUNX2 activation. IGF-1 increased ERK1/2 activation, which was abrogated by the inhibition of PI3K, thus linking these two pathways in EC. Treatment with U0126, which inhibits ERK1/2 activation, reduced IGF-1-stimulated RUNX2 DNA binding without affecting RUNX2 protein levels. Overexpression of constitutively active MKK1 increased RUNX2 DNA binding and phosphorylation. No additive effects of PI3K or ERK inhibitors on DNA binding were evident. Surprisingly, these IGF-1-mediated effects on RUNX2 were not regulated by Akt phosphorylation, a common downstream target of PI3K, as determined by pharmacological or genetic inhibition. However, an inhibitor of the p21-activated protein kinase-1, glutathione S-transferase-Pak1-(83-149), inhibited both basal and IGF-1-stimulated RUNX2 DNA binding, suggesting that Pak1 mediates IGF-1 signaling to increase RUNX2 activity. These results indicate that the angiogenic growth factor, IGF-1, can regulate RUNX2 DNA binding through sequential activation of the PI3K/Pak1 and ERK1/2 signaling cascade.
Department of Pharmaceutical Sciences, University of Maryland School of Pharmacy, Baltimore, Maryland 21201, USA.
Meng
M
Qiao
Paul
P
Shapiro
Rakesh
R
Kumar
Antonino
A
Passaniti
D015153:Blotting, Western; D002070:Butadienes / Q000494:pharmacology; D002455:Cell Division; D002460:Cell Line; D002465:Cell Movement; D002467:Cell Nucleus / Q000378:metabolism; D002867:Chromones / Q000494:pharmacology; D050659:Core Binding Factor Alpha 1 Subunit; D004247:DNA / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D042783:Endothelial Cells / Q000378:metabolism; D004730:Endothelium, Vascular / Q000378:metabolism*; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005786:Gene Expression Regulation; D006801:Humans; D007334:Insulin-Like Growth Factor I / Q000378:metabolism / Q000502:physiology*; D048369:MAP Kinase Kinase 1 / Q000378:metabolism; D048370:MAP Kinase Kinase 2 / Q000378:metabolism; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D008954:Models, Biological; D009025:Morpholines / Q000494:pharmacology; D009363:Neoplasm Proteins / Q000378:metabolism*; D009570:Nitriles / Q000494:pharmacology; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D010766:Phosphorylation; D011485:Protein Binding; D011499:Protein Processing, Post-Translational; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry; D015398:Signal Transduction; D013997:Time Factors; D014157:Transcription Factors / Q000378:metabolism*; D014162:Transfection; D054462:p21-Activated Kinases
D002070:Butadienes; D002867:Chromones; D050659:Core Binding Factor Alpha 1 Subunit; D004247:DNA; D004791:Enzyme Inhibitors; D007334:Insulin-Like Growth Factor I; D048369:MAP Kinase Kinase 1; D048370:MAP Kinase Kinase 2; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases; D009025:Morpholines; D009363:Neoplasm Proteins; D009570:Nitriles; D019869:Phosphatidylinositol 3-Kinases; D017346:Protein Serine-Threonine Kinases; D011994:Recombinant Proteins; D014157:Transcription Factors; D054462:p21-Activated Kinases; C113580:U 0126; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; C482128:MAP2K2 protein, human; C515903:PAK1 protein, human; C482125:MAP2K1 protein, human