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Specific transcripts are elevated in Saccharomyces cerevisiae in response to DNA damage
Abstract:
Differential hybridization has been used to identify genes in Saccharomyces cerevisiae displaying increased transcript levels after treatment of cells with UV irradiation or with the mutagen/carcinogen 4-nitroquinoline-1-oxide (NQO). We describe the isolation and characterization of four DNA damage responsive genes obtained from screening ca. 9,000 yeast genomic clones. Two of these clones, lambda 78A and pBR178C, contain repetitive elements in the yeast genome as shown by Southern hybridization analysis. Although the genomic hybridization pattern is distinct for each of these two clones, both of these sequences hybridize to large polyadenylated transcripts ca. 5 kilobases in length. Two other DNA damage responsive sequences, pBRA2 and pBR3016B, are single-copy genes and hybridize to 0.5- and 3.2-kilobase transcripts, respectively. Kinetic analysis of the 0.5-kilobase transcript homologous to pBRA2 indicates that the level of this RNA increases more than 15-fold within 20 min after exposure to 4-nitroquinoline-1-oxide. Moreover, the level of this transcript is significantly elevated in cells containing the rad52-1 mutation which are deficient in DNA strand break repair and gene conversion. These results provide some of the first evidence that DNA damage stimulates transcription of specific genes in eucaryotic cells.
Insights
Researchers identified four DNA damage-responsive genes in yeast using differential hybridization. One gene
Area of Science:
- Molecular Biology
- Genetics
- Biochemistry
Background:
- DNA damage response pathways are crucial for maintaining genomic integrity in eukaryotic cells.
- Understanding gene regulation under DNA-damaging conditions is essential for comprehending cellular repair mechanisms.
Purpose of the Study:
- To identify and characterize genes in Saccharomyces cerevisiae that are upregulated in response to DNA damage.
- To investigate the transcriptional regulation of these damage-inducible genes.
Main Methods:
- Differential hybridization was employed to screen approximately 9,000 yeast genomic clones.
- Southern hybridization analysis was used to characterize repetitive elements and transcript sizes.
- Kinetic analysis and Northern hybridization were performed to quantify transcript levels.
Main Results:
- Four DNA damage-responsive genes were isolated and characterized.
- Two genes (lambda 78A and pBR178C) contained repetitive elements and hybridized to 5-kilobase transcripts.
- Two other genes (pBRA2 and pBR3016B) were single-copy and hybridized to 0.5- and 3.2-kilobase transcripts, respectively.
- The transcript for pBRA2 showed a >15-fold increase within 20 minutes after 4-nitroquinoline-1-oxide (NQO) exposure and was elevated in rad52-1 mutant cells.
Conclusions:
- Specific genes in eukaryotic cells are transcriptionally activated by DNA damage.
- The rad52-1 mutation, involved in DNA repair, affects the induction of specific damage-responsive transcripts.
- This study provides foundational evidence for DNA damage-induced gene transcription in eukaryotes.