Identification of the DNA damage-responsive element of RNR2 and evidence that four distinct cellular factors bind it.
The RNR2 gene encodes the small subunit of ribonucleotide reductase, the enzyme that catalyzes the first step in the pathway for the production of the deoxyribonucleotides needed for DNA synthesis. Transcription of this gene is induced approximately 20-fold in response to environmental stimuli that damage DNA or block DNA replication. Deletion and subcloning analysis identified two, and possibly three, upstream activating sequences (UAS) and one repressing (URS) element in the RNR2 regulatory region. A 42-base-pair (bp) fragment from this region was found to be necessary for proper regulation of RNR2 and to be capable of conferring DNA damage inducibility upon a heterologous promoter. This fragment contained both positively and negatively acting sequences. Four DNA-binding factors interacted with the RNR2 regulatory region. One factor was identified as the GRF1 protein, the product of the RAP1 gene. GRF1 bound to the UAS2 element of RNR2, which was found to be directly adjacent to the 42-bp fragment. UAS2 activity was repressed by the 42-bp fragment. Three other factors bound to the 42-bp fragment; one of these factors, RRF3, had a second binding site in the RNR2 promoter. These factors are likely to mediate the response of RNR2 to DNA damage.
Elledge
S J
SJ
Department of Biochemistry, Baylor College of Medicine, Houston, Texas 77030.
Davis
R W
RW
D001483:Base Sequence; D002467:Cell Nucleus / Q000378:metabolism; D002872:Chromosome Deletion; D003001:Cloning, Molecular; D004249:DNA Damage; D004261:DNA Replication; D004271:DNA, Fungal / Q000235:genetics* / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004794:Enzyme Repression; D005656:Fungal Proteins / Q000378:metabolism; D015971:Gene Expression Regulation, Enzymologic; D015966:Gene Expression Regulation, Fungal; D005800:Genes, Fungal; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D010957:Plasmids; D011401:Promoter Regions, Genetic; D012045:Regulatory Sequences, Nucleic Acid; D015183:Restriction Mapping; D012264:Ribonucleotide Reductases / Q000096:biosynthesis / Q000235:genetics*; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics*; D014157:Transcription Factors; D014158:Transcription, Genetic
D004271:DNA, Fungal; D004268:DNA-Binding Proteins; D005656:Fungal Proteins; D046911:Macromolecular Substances; D014157:Transcription Factors; D012264:Ribonucleotide Reductases