Common target for 12-O-tetradecanoylphorbol-13-acetate and ras in the transcriptional enhancer of the growth factor-inducible JE gene.
JE gene expression in the mouse osteoblast cell line MC3T3-E1 is activated transiently by the tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA). In ras-transformed MC3T3-E1 cells the JE gene is constitutively expressed at a high level, whereas in their raf-transformed counterparts it is not constitutively expressed or inducible by TPA. Using these cells, we investigated a specific sequence recognized by nuclear factors in the 5'-upstream region of the rat JE gene. By gel-mobility shift assays, we determined that the amount of nuclear factors that bind to the region -130 to -96 bp upstream from the cap site of the rat JE gene (JE-1 probe) increased after TPA treatment of MC3T3-E1 cells. However, in the ras transformants it was elevated constitutively, and in the raf transformants it was not detectable. Both unlabeled JE-1 probe and a probe containing a TPA-responsive element (TGACTCA) competed with the binding of these nuclear factors. Preincubation of the nuclear extracts with fos- or jun-specific antibodies interfered with the binding of the factors to the JE-1 probe. The essential sequence in the JE-1 element for the binding of nuclear factors was found to be TTACTCA. c-fos and c-jun proteins synthesized in vitro could bind to the DNA fragment containing this sequence, but the binding was weaker than to the TPA-responsive element (TGACTCA). These results suggest that the sequence TTACTCA in the JE-1 element is a common target of TPA and ras and that protein complexes containing fos- and jun-related proteins recognize the sequence.
Department of Cancer Cell Research, University of Tokyo, Japan.
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D000818:Animals; D001483:Base Sequence; D001667:Binding, Competitive; D002460:Cell Line; D002461:Cell Line, Transformed; D018932:Chemokine CCL2; D002630:Chemotactic Factors / Q000235:genetics* / Q000378:metabolism; D016207:Cytokines / Q000235:genetics* / Q000378:metabolism; D015342:DNA Probes; D004742:Enhancer Elements, Genetic / Q000187:drug effects; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects / Q000235:genetics; D011905:Genes, ras / Q000502:physiology*; D051379:Mice; D008969:Molecular Sequence Data; D017354:Point Mutation; D011401:Promoter Regions, Genetic; D016760:Proto-Oncogene Proteins c-fos / Q000378:metabolism; D016755:Proto-Oncogene Proteins c-jun / Q000378:metabolism; D012333:RNA, Messenger / Q000096:biosynthesis; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*; D014158:Transcription, Genetic / Q000187:drug effects / Q000502:physiology
D018932:Chemokine CCL2; D002630:Chemotactic Factors; D016207:Cytokines; D015342:DNA Probes; D016760:Proto-Oncogene Proteins c-fos; D016755:Proto-Oncogene Proteins c-jun; D012333:RNA, Messenger; D013755:Tetradecanoylphorbol Acetate