Fibroblast growth factor-2 induction of platelet-derived growth factor-C chain transcription in vascular smooth muscle cells is ERK-dependent but not JNK-dependent and mediated by Egr-1.
Platelet-derived growth factors (PDGFs) play an integral role in normal tissue growth and maintenance as well as many human pathological states including atherosclerosis, fibrosis, and tumorigenesis. The PDGF family of ligands is comprised of A, B, C, and D chains. Here, we provide the first functional characterization of the PDGF-C promoter. We examined 797 bp of the human PDGF-C promoter and identified several putative recognition elements for Sp1, Ets Egr-1, and Smad. The proximal region of the PDGF-C promoter bears a remarkable resemblance to a comparable region of the PDGF-A promoter (1). Binding and transient transfection analysis in primary vascular smooth muscle cells revealed that PDGF-C, like PDGF-A, is under the transcriptional control of the zinc finger nuclear protein Egr-1 (early growth response-1). Electrophoretic mobility shift analysis using both smooth muscle cell nuclear extracts and recombinant protein revealed that Egr-1 and Sp1 bind this region of the PDGF-C promoter (Oligo C, -35 to -1). Egr-1 competes with Sp1 for overlapping binding sites even when the former is at a stoichiometric disadvantage. Reverse transcriptase PCR and supershift analysis demonstrate that fibroblast growth factor-2 (FGF-2) stimulates both Egr-1 and PDGF-C mRNA expression in a time-dependent and transient manner and that FGF-2-inducible Egr-1 binds the proximal PDGF-C promoter. FGF-2-inducible PDGF-C expression was completely abrogated using catalytic DNA (DNAzymes) targeting Egr-1 but not by its scrambled counterpart. Moreover, using pharmacological inhibitors we demonstrate the critical role of ERK but not JNK in FGF-2-inducible PDGF-C expression. These findings thus demonstrate that PDGF-C transcription, activated by FGF-2, is mediated by Egr-1 and its upstream kinase ERK.
Midgley
Valerie C
VC
Centre for Vascular Research, The University of New South Wales, Department of Haematology, The Prince of Wales Hospital, Sydney, New South Wales 2052, Australia.
Khachigian
Levon M
LM
D000818:Animals; D001011:Aorta / Q000166:cytology*; D001483:Base Sequence; D001665:Binding Sites; D002384:Catalysis; D002467:Cell Nucleus / Q000378:metabolism; D002478:Cells, Cultured; D004247:DNA / Q000737:chemistry; D004268:DNA-Binding Proteins / Q000378:metabolism*; D051766:Early Growth Response Protein 1; D016222:Fibroblast Growth Factor 2 / Q000378:metabolism*; D006801:Humans; D017874:Immediate-Early Proteins / Q000378:metabolism*; D048031:JNK Mitogen-Activated Protein Kinases; D008222:Lymphokines; D048670:MAP Kinase Kinase 4; D020929:Mitogen-Activated Protein Kinase Kinases / Q000378:metabolism*; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D008969:Molecular Sequence Data; D009131:Muscle, Smooth, Vascular / Q000378:metabolism*; D032389:Myocytes, Smooth Muscle / Q000378:metabolism; D009841:Oligonucleotides / Q000737:chemistry; D010957:Plasmids / Q000378:metabolism; D010982:Platelet-Derived Growth Factor / Q000235:genetics* / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D011994:Recombinant Proteins / Q000737:chemistry; D015398:Signal Transduction; D016329:Sp1 Transcription Factor / Q000378:metabolism; D013997:Time Factors; D014157:Transcription Factors / Q000378:metabolism*; D014158:Transcription, Genetic; D014162:Transfection
D004268:DNA-Binding Proteins; C497325:EGR1 protein, human; D051766:Early Growth Response Protein 1; C486110:Egr1 protein, rat; D017874:Immediate-Early Proteins; D008222:Lymphokines; D009841:Oligonucleotides; D010982:Platelet-Derived Growth Factor; D012333:RNA, Messenger; D011994:Recombinant Proteins; D016329:Sp1 Transcription Factor; D014157:Transcription Factors; C411423:platelet-derived growth factor C; D016222:Fibroblast Growth Factor 2; D004247:DNA; D048031:JNK Mitogen-Activated Protein Kinases; D020928:Mitogen-Activated Protein Kinases; D048670:MAP Kinase Kinase 4; D020929:Mitogen-Activated Protein Kinase Kinases