A polyomavirus enhancer A-binding protein-3 site and Ets-2 protein have a major role in the 12-O-tetradecanoylphorbol-13-acetate response of the human stromelysin gene.
The expression of stromelysin, a major matrix metalloproteinase of connective tissues, is regulated by several cytokines, growth factors, protooncogenes as well as by 12-O-tetradecanoylphorbol-13-acetate (TPA). The human stromelysin gene promoter contains an activator protein-1 (Fos/Jun) binding site at -70, which is required for basal expression but is not necessary for the TPA response. In this study, using promoter deletion mutants in transient gene transfection experiments, we first identify the sequence from -220 to -202 as necessary for the TPA response of the stromelysin gene. Further, among the restriction fragments from the 1.3-kilobase long promoter, only the proximal fragment (-274 to -101) conferred a TPA response on the heterologous thymidine kinase gene promoter. The -220 to -202 sequence contains two copies of a motif similar to the polyomavirus enhancer A-binding protein-3 (PEA-3) site, which binds the Ets family of oncoproteins and transcription factors. Point mutations of either one of the two PEA-3 sites, in the 1.3-kilobase long stromelysin promoter context, reduced basal gene expression. However, only the mutation of the proximal, but not the distal PEA-3 site, significantly inhibited the TPA response. In cotransfection experiments, the Ets-2 protein transactivated the stromelysin promoter and the promoter proximal fragment containing the PEA-3 sites but not the promoters containing mutated PEA-3 sites. These data suggest that the PEA-3 site, but not the activator protein-1 site, and Ets-2 protein have a major role in the TPA induction of the human stromelysin gene transcription.
Buttice
G
G
University of Medicine and Dentistry of New Jersey-Robert Wood Johnson Medical School, Piscataway 08854.
Kurkinen
M
M
D000818:Animals; D001483:Base Sequence; D004247:DNA; D004268:DNA-Binding Proteins; D005786:Gene Expression Regulation; D006801:Humans; D019278:Matrix Metalloproteinase 3; D008666:Metalloendopeptidases / Q000235:genetics*; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic; D050787:Proto-Oncogene Protein c-ets-2; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D016755:Proto-Oncogene Proteins c-jun / Q000378:metabolism; D051381:Rats; D012097:Repressor Proteins; D015183:Restriction Mapping; D017384:Sequence Deletion; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*; D013937:Thymidine Kinase / Q000235:genetics / Q000378:metabolism; D015534:Trans-Activators; D014157:Transcription Factors / Q000378:metabolism*; D014158:Transcription, Genetic / Q000187:drug effects; D014407:Tumor Cells, Cultured
D004268:DNA-Binding Proteins; C107465:ERF protein, human; C494069:ETS2 protein, human; C494071:Ets2 protein, rat; D050787:Proto-Oncogene Protein c-ets-2; D011518:Proto-Oncogene Proteins; D016755:Proto-Oncogene Proteins c-jun; D012097:Repressor Proteins; D015534:Trans-Activators; D014157:Transcription Factors; C065788:transcription factor PEA3; D004247:DNA; D013937:Thymidine Kinase; D008666:Metalloendopeptidases; D019278:Matrix Metalloproteinase 3; D013755:Tetradecanoylphorbol Acetate