The helix-loop-helix protein rE12 and the C/EBP-related factor rNFIL-6 bind to neighboring sites within the c-fos serum response element.
We show that members of two major families of transcription factors, the helix-loop-helix and C/EBP families, interact with the c-fos serum response element (SRE). Two cDNA clones encoding SRE binding factors (clones 9 and 21) were isolated by the direct screening of a PC12 lambda gt11 cDNA library using SRE oligonucleotide sequences as probes. Clone 9 encodes the rat homolog of the human HLH transcription factor, E12 (called here rE12). Clone 21 encodes a b-zip domain polypeptide that is related to the liver transcription factor C/EBP, and is homologous to the human NFIL-6 transcription factor (called here rNFIL-6). Using in vitro-translated products we show that rNFIL-6 recognizes a 'CCAATT' motif which overlaps the c-fos dyad symmetry element (DSE), the binding site for serum regulatory factor (SRF). Factor rE12 binds to an E-box enhancer sequence, 'CATCTG', immediately adjacent to the rNFIL-6 site, within the SRE. Antibodies specific to rE12 and rNFIL-6 disrupt nucleoprotein complexes with these DNA-binding sites, confirming the interaction of native in vivo factors. We present evidence that rNFIL-6 and SRF binding are mutually exclusive, consistent with the overlap of their binding sites. The demonstration that rE12 and rNFIL-6 bind to the SRE at sites adjacent to the major c-fos regulatory element, the DSE, raises the possibility that helix-loop-helix and C/EBP families regulate the SRE and provide a new basis for the multifunctional properties of the SRE, including possible tissue specificity of expression.
Metz
R
R
Howard Hughes Medical Institute, Department of Biochemistry, New York, New York.
Ziff
E
E
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D022762:CCAAT-Enhancer-Binding Proteins; D003001:Cloning, Molecular; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D015723:Gene Library; D016762:Genes, fos; D006801:Humans; D016350:Leucine Zippers / Q000235:genetics; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism*; D009838:Oligodeoxyribonucleotides; D016716:PC12 Cells; D010957:Plasmids; D014176:Protein Biosynthesis; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012689:Sequence Homology, Nucleic Acid; D051820:TCF Transcription Factors; D058125:Transcription Factor 7-Like 1 Protein; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism; D001616:beta-Galactosidase / Q000235:genetics / Q000378:metabolism
D022762:CCAAT-Enhancer-Binding Proteins; D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D009838:Oligodeoxyribonucleotides; D011993:Recombinant Fusion Proteins; D051820:TCF Transcription Factors; C544912:TCF7L1 protein, human; C553060:Tcf7l1 protein, rat; D058125:Transcription Factor 7-Like 1 Protein; D014157:Transcription Factors; D001616:beta-Galactosidase