The TATA-less promoter of mouse ribonucleotide reductase R1 gene contains a TFII-I binding initiator element essential for cell cycle-regulated transcription.
Mammalian ribonucleotide reductase shows S-phase specific expression and consists of two non-identical subunits, proteins R1 (large subunit) and R2 (small subunit). A comparison between the human and mouse TATA-less R1 gene promoters revealed four highly conserved DNA regions, while the remaining sequence showed a low degree of conservation. Two regions, alpha and beta, were earlier identified as protein binding regions in the mouse R1 promoter by using DNase footprinting technique. The two new regions are located to the transcription start and to a DNA sequence about 40 base pairs downstream from the start. Gel shift assays using TFII-I antibodies and competition with an oligonucleotide representing the terminal deoxynucleotidyl transferase inhibitor element identified the start region as a TFII-I binding initiator element. The conserved downstream region, called gamma, also formed specific DNA-protein complexes in gel shift assays. Functional studies, using synchronized cells stably transformed by R1 promoter-luciferase reporter gene constructs, indicated that the initiator and the gamma elements together were necessary for cell cycle-regulated R1 promoter activity. Earlier published data, indicating Sp1 binding to the R1 alpha/beta regions, could not be confirmed, suggesting that the R1 initiator element may function independent of Sp1.
Johansson
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Department of Medical Biochemistry and Biophysics, Umeå University, Sweden.
Skogman
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Thelander
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D016475:3T3 Cells; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D002453:Cell Cycle; D017124:Conserved Sequence; D018983:DNA Footprinting; D017931:DNA Primers; D004268:DNA-Binding Proteins / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D008156:Luciferases / Q000096:biosynthesis; D046911:Macromolecular Substances; D051379:Mice; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic; D011994:Recombinant Proteins / Q000096:biosynthesis; D012264:Ribonucleotide Reductases / Q000235:genetics*; D012689:Sequence Homology, Nucleic Acid; D016385:TATA Box; D014157:Transcription Factors / Q000378:metabolism*; D014162:Transfection
D017931:DNA Primers; D004268:DNA-Binding Proteins; D046911:Macromolecular Substances; D011994:Recombinant Proteins; D014157:Transcription Factors; D008156:Luciferases; D012264:Ribonucleotide Reductases