Activated Ha-Ras but not TPA induces transcription through binding sites for activating transcription factor 3/Jun and a novel nuclear factor.
We report the identification of a 20-base pair sequence mediating induced transcription in response to an activated Ha-ras gene and epidermal growth factor (EGF) but not 12-O-tetradecanoylphorbol-13-acetate stimulation. This signal-specific nuclear target is present in the long terminal repeat of a mouse VL30 retrotransposon expressed in epidermis. Functional studies and in vitro binding analyses using cultured keratinocytes (Balb/MK) reveal that the response element is composed of two cooperating sequence motifs in juxtaposed position, both of which are targets for induced binding activity 1-2 h after EGF stimulation. Of many different activating transcription factor/cAMP-responsive element binding protein/activating protein 1 factors tested, one part of the sequence selectively binds endogenous proteins immunologically related to activating transcription factor 3 (ATF3) and Jun isotypes. The other sequence is a target for a nuclear factor showing binding specificity unrelated to factors known to mediate EGF- or ras-induced transcription as determined by its sequence specificity and by antibody experiments. This component has been characterized and partially purified by gel filtration chromatography and velocity centrifugation revealing a Stokes radius of 43.6 A and a sedimentation coefficient of 9.7 S in solution. Based on these parameters, a molecular mass of 178,000 Da was calculated. The results indicate that the specific binding of ATF3/Jun and a previously uncharacterized factor account for signal-specific transcription in response to EGF or an activated Ha-ras gene in a cell type in which the cooperative action of an activated Ha-ras gene and 12-O-tetradecanoylphorbol-13-acetate cause tumor growth.
Nilsson
M
M
Center for Nutrition and Toxicology, Karolinska Institute, NOVUM, Huddinge, Sweden.
Toftgård
R
R
Bohm
S
S
D016475:3T3 Cells; D051700:Activating Transcription Factor 3; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D002478:Cells, Cultured; D005786:Gene Expression Regulation; D011905:Genes, ras; D015603:Keratinocytes / Q000378:metabolism*; D051379:Mice; D008969:Molecular Sequence Data; D016755:Proto-Oncogene Proteins c-jun / Q000378:metabolism*; D016283:Proto-Oncogene Proteins p21(ras) / Q000235:genetics / Q000494:pharmacology*; D012091:Repetitive Sequences, Nucleic Acid; D018626:Retroelements / Q000187:drug effects* / Q000235:genetics; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*; D014157:Transcription Factors / Q000378:metabolism*
D051700:Activating Transcription Factor 3; D016755:Proto-Oncogene Proteins c-jun; D018626:Retroelements; D014157:Transcription Factors; D016283:Proto-Oncogene Proteins p21(ras); D013755:Tetradecanoylphorbol Acetate