Role of protein kinase Czeta in Ras-mediated transcriptional activation of vascular permeability factor/vascular endothelial growth factor expression.
Vascular permeability factor/vascular endothelial growth factor (VPF/VEGF), a multifunctional cytokine, is regulated by different factors including degree of cell differentiation, hypoxia, and certain oncogenes namely, ras and src. The up-regulation of VPF/VEGF expression by Ras has been found to be through both transcription and mRNA stability. The present study investigates a novel pathway whereby Ras promotes the transcription of VPF/VEGF by activating protein kinase Czeta (PKCzeta). The Ras-mediated overexpression of VPF/VEGF was also found to be inhibited by using the antisense or the dominant-negative mutant of PKCzeta. In co-transfection assays, by overexpressing oncogenic Ha-Ras (12 V) and PKCzeta, there was an additive effect up to 4-fold in activation of Sp1-mediated VPF/VEGF transcription. It has been shown through electrophoretic mobility shift assay that Ras promoted the PKCzeta-induced binding of Sp1 to the VPF/VEGF promoter. In the presence of PDK-1, a major activating kinase for PKC, the Ras-mediated activation of VPF/VEGF promoter through PKCzeta was further increased, suggesting that PKCzeta can serve as an effector for both Ras and PDK-1. In other experiments, with the use of a dominant-negative mutant of phosphatidylinositol 3-kinase, the activation of VPF/VEGF promoter through Ras, PDK-1, and PKCzeta was completely repressed, indicating phosphatidylinositol 3-kinase as an important component of this pathway. Taken together, these data elucidate the signaling mechanism of Ras-mediated VPF/VEGF transcriptional activation through PKCzeta and also provide insight into PKCzeta and Sp1-dependent transcriptional regulation of VPF/VEGF.
Pal
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Department of Pathology, Beth Israel Deaconess Medical Center and Harvard Medical School, Boston, Massachusetts 02215, USA.
Datta
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Khosravi-Far
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R
Mukhopadhyay
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D064413:3-Phosphoinositide-Dependent Protein Kinases; D016228:Endothelial Growth Factors / Q000235:genetics*; D004789:Enzyme Activation; D008222:Lymphokines / Q000235:genetics*; D009389:Neovascularization, Pathologic; D016376:Oligonucleotides, Antisense; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D011493:Protein Kinase C / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D019908:Proto-Oncogene Proteins c-raf / Q000378:metabolism; D015398:Signal Transduction; D016329:Sp1 Transcription Factor / Q000378:metabolism; D015533:Transcriptional Activation; D014407:Tumor Cells, Cultured; D042461:Vascular Endothelial Growth Factor A; D042442:Vascular Endothelial Growth Factors; D018631:ras Proteins / Q000378:metabolism*; D000099221:Protein Kinase C zeta
D064413:3-Phosphoinositide-Dependent Protein Kinases; D016228:Endothelial Growth Factors; D008222:Lymphokines; D016376:Oligonucleotides, Antisense; D019869:Phosphatidylinositol 3-Kinases; D011493:Protein Kinase C; D017346:Protein Serine-Threonine Kinases; D019908:Proto-Oncogene Proteins c-raf; D016329:Sp1 Transcription Factor; D042461:Vascular Endothelial Growth Factor A; D042442:Vascular Endothelial Growth Factors; D018631:ras Proteins; D000099221:Protein Kinase C zeta