title: Interplay of Sp1 and Egr-1 in the proximal platelet-derived growth factor A-chain promoter in cultured vascular endothelial cells.
abstract: The platelet-derived growth factor (PDGF) A-chain has been implicated in the initiation and progression of vascular occlusive lesions. The elements in the human PDGF-A promoter that mediate increased expression of the gene in vascular endothelial cells have not been identified. A potent inducer of PDGF-A expression in endothelial cells is phorbol 12-myristate 13-acetate (PMA). 5'-Deletion and transfection analysis revealed that a G+C-rich region in the proximal PDGF-A promoter is required for PMA-inducible gene expression. This region bears overlapping consensus recognition sequences for Sp1 and Egr-1. PMA induces Egr-1 mRNA expression within 1 h, whereas PDGF-A transcript levels increase after 2-4 h. Constitutive levels of Sp1 are not altered over 24 h. A specific nucleoprotein complex is formed when an oligonucleotide bearing the G+C-rich element is incubated with nuclear extracts from PMA-treated cells. The temporal appearance of this complex is consistent with the transient increase in Egr-1 transcripts. Antibodies to Egr-1 completely supershift the PMA-induced complex. Interestingly, increased nuclear levels of Egr-1 attenuate the ability of Sp1 to interact with the oligonucleotide, implicating competition between Egr-1 and Sp1 for the G+C-rich element. Binding studies with recombinant proteins demonstrate that Egr-1 can displace Sp1 from this region. Insertion of the G+C-rich element into a hybrid promoter-reporter construct confers PMA inducibility on the construct. Mutations that abolish Egr-1 binding also abrogate expression induced by PMA or overexpressed Egr-1. These findings demonstrate that PMA-induced Egr-1 displaces Sp1 from the G+C-rich element and activates expression driven by the PDGF-A proximal promoter in endothelial cells. The Sp1/Egr-1 displacement mechanism may be an important regulatory circuit in the control of inducible gene expression in vascular endothelial cells.
mesh_terms: D000818:Animals; D001011:Aorta; D001483:Base Sequence; D001665:Binding Sites; D002417:Cattle; D002467:Cell Nucleus / Q000378:metabolism; D002478:Cells, Cultured; D016384:Consensus Sequence; D004268:DNA-Binding Proteins / Q000096:biosynthesis / Q000378:metabolism*; D051766:Early Growth Response Protein 1; D004730:Endothelium, Vascular / Q000187:drug effects / Q000378:metabolism*; D015870:Gene Expression / Q000187:drug effects*; D006801:Humans; D017874:Immediate-Early Proteins; D008954:Models, Biological; D008969:Molecular Sequence Data; D010982:Platelet-Derived Growth Factor / Q000096:biosynthesis* / Q000235:genetics*; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000096:biosynthesis; D016329:Sp1 Transcription Factor / Q000378:metabolism*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014157:Transcription Factors / Q000096:biosynthesis / Q000378:metabolism*; D014158:Transcription, Genetic / Q000187:drug effects; D016335:Zinc Fingers
chemical_list: D004268:DNA-Binding Proteins; C497325:EGR1 protein, human; D051766:Early Growth Response Protein 1; D017874:Immediate-Early Proteins; D010982:Platelet-Derived Growth Factor; D012333:RNA, Messenger; D016329:Sp1 Transcription Factor; D014157:Transcription Factors; D013755:Tetradecanoylphorbol Acetate