scholarly journals DNA damage induction of ribonucleotide reductase.

1989 ◽  
Vol 9 (11) ◽  
pp. 4932-4940 ◽  
Author(s):  
S J Elledge ◽  
R W Davis
Keyword(s):  
Dna Damage ◽  
Stress Response ◽  
Ribonucleotide Reductase ◽  
Uv Light ◽  
Small Subunit ◽  
Lacz Fusion ◽  
Dna Damaging Agents ◽  
Yeast Strains ◽  
Increased Sensitivity ◽  
Beta Galactosidase

RNR2 encodes the small subunit of ribonucleotide reductase, the enzyme that catalyzes the first step in the pathway for the production of deoxyribonucleotides needed for DNA synthesis. RNR2 is a member of a group of genes whose activities are cell cycle regulated and that are transcriptionally induced in response to the stress of DNA damage. An RNR2-lacZ fusion was used to further characterize the regulation of RNR2 and the pathway responsible for its response to DNA damage. beta-Galactosidase activity in yeast strains containing the RNR2-lacZ fusion was inducible in response to DNA-damaging agents (UV light, 4-nitroquinoline-1-oxide [4-NQO], and methyl methanesulfonate [MMS]) and agents that block DNA replication (hydroxyurea [HU] and methotrexate) but not heat shock. When MATa cells were arrested in G1 by alpha-factor, RNR2 mRNA was still inducible by DNA damage, indicating that the observed induction can occur outside of S phase. In addition, RNR2 induction was not blocked by the presence of cycloheximide and is therefore likely to be independent of protein synthesis. A mutation, rnr2-314, was found to confer hypersensitivity to HU and increased sensitivity to MMS. In rnr2-314 mutant strains, the DNA damage stress response was found to be partially constitutive as well as hypersensitive to induction by HU but not MMS. The induction properties of RNR2 were examined in a rad4-2 mutant background; in this genetic background, RNR2 was hypersensitive to induction by 4-NQO but not MMS. Induction of the RNR2-lacZ fusion in a RAD(+) strain in response to 4-NQO was not enhanced by the presence of an equal number of rad4-2 cells that lacked the fusion, implying that the DNA damage stress response in cell autonomous.

DNA damage induction of ribonucleotide reductase

1989 ◽  
Vol 9 (11) ◽  
pp. 4932-4940
Author(s):  
S J Elledge ◽  
R W Davis
Keyword(s):  
Dna Damage ◽  
Stress Response ◽  
Ribonucleotide Reductase ◽  
Uv Light ◽  
Small Subunit ◽  
Lacz Fusion ◽  
Dna Damaging Agents ◽  
Yeast Strains ◽  
Increased Sensitivity ◽  
Beta Galactosidase

RNR2 encodes the small subunit of ribonucleotide reductase, the enzyme that catalyzes the first step in the pathway for the production of deoxyribonucleotides needed for DNA synthesis. RNR2 is a member of a group of genes whose activities are cell cycle regulated and that are transcriptionally induced in response to the stress of DNA damage. An RNR2-lacZ fusion was used to further characterize the regulation of RNR2 and the pathway responsible for its response to DNA damage. beta-Galactosidase activity in yeast strains containing the RNR2-lacZ fusion was inducible in response to DNA-damaging agents (UV light, 4-nitroquinoline-1-oxide [4-NQO], and methyl methanesulfonate [MMS]) and agents that block DNA replication (hydroxyurea [HU] and methotrexate) but not heat shock. When MATa cells were arrested in G1 by alpha-factor, RNR2 mRNA was still inducible by DNA damage, indicating that the observed induction can occur outside of S phase. In addition, RNR2 induction was not blocked by the presence of cycloheximide and is therefore likely to be independent of protein synthesis. A mutation, rnr2-314, was found to confer hypersensitivity to HU and increased sensitivity to MMS. In rnr2-314 mutant strains, the DNA damage stress response was found to be partially constitutive as well as hypersensitive to induction by HU but not MMS. The induction properties of RNR2 were examined in a rad4-2 mutant background; in this genetic background, RNR2 was hypersensitive to induction by 4-NQO but not MMS. Induction of the RNR2-lacZ fusion in a RAD(+) strain in response to 4-NQO was not enhanced by the presence of an equal number of rad4-2 cells that lacked the fusion, implying that the DNA damage stress response in cell autonomous.

Identification and isolation of the gene encoding the small subunit of ribonucleotide reductase from Saccharomyces cerevisiae: DNA damage-inducible gene required for mitotic viability

1987 ◽  
Vol 7 (8) ◽  
pp. 2783-2793
Author(s):  
S J Elledge ◽  
R W Davis
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Amino Acids ◽  
Dna Damage ◽  
Ribonucleotide Reductase ◽  
Small Subunit ◽  
Cross Reaction ◽  
Expression Library ◽  
Gene Encoding ◽  
Ribonucleotide Reductases ◽  
Colony Color

Ribonucleotide reductase catalyzes the first step in the pathway for the production of deoxyribonucleotides needed for DNA synthesis. The gene encoding the small subunit of ribonucleotide reductase was isolated from a Saccharomyces cerevisiae genomic DNA expression library in lambda gt11 by a fortuitous cross-reaction with anti-RecA antibodies. The cross-reaction was due to an identity between the last four amino acids of each protein. The gene has been named RNR2 and is centromere linked on chromosome X. The nucleotide sequence was determined, and the deduced amino acid sequence, 399 amino acids, shows extensive homology with other eucaryotic ribonucleotide reductases. Transplason mutagenesis was used to disrupt the RNR2 gene. A novel assay using colony color sectoring was developed to demonstrate visually that RNR2 is essential for mitotic viability. RNR2 encodes a 1.5-kilobase mRNA whose levels increase 18-fold after treatment with the DNA-damaging agent 4-nitroquinoline 1-oxide. CDC8 was also found to be inducible by DNA damage, but POL1 and URA3 were not inducible by 4-nitroquinoline 1-oxide. The expression of these genes defines a new mode of regulation for enzymes involved in DNA biosynthesis and sharpens our picture of the events leading to DNA repair in eucaryotic cells.

Upstream regulatory sequences of the yeast RNR2 gene include a repression sequence and an activation site that binds the RAP1 protein

1989 ◽  
Vol 9 (12) ◽  
pp. 5359-5372
Author(s):  
H K Hurd ◽  
J W Roberts
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Ribonucleotide Reductase ◽  
Small Subunit ◽  
Methyl Methanesulfonate ◽  
Regulatory Sequences ◽  
Start Site ◽  
Transcription Start ◽  
Dna Damaging Agents ◽  
Number Of Genes ◽  
Upstream Regulatory Sequences

The small subunit of ribonucleotide reductase in Saccharomyces cerevisiae (RNR2) was induced 3- to 20-fold by a variety of DNA-damaging agents. Induction of the RNR2 transcript by at least one of these agents, methyl methanesulfonate, did not require protein synthesis. To identify sequences involved in the regulation of RNR2, we introduced deletions upstream of the transcription start site. Sequences required for induction were contained within a 200-base-pair region that could confer methyl methanesulfonate inducibility on the heterologous CYC1 promoter. This region contained a repression sequence and at least two positive activation sites. One of these activation sites bound RAP1, a protein known to associate with mating-type silencers and the upstream activation sequences of a number of genes. The behavior of deletions of the repression sequence suggests that induction of RNR2 may occur, at least in part, through relief of repression.

scholarly journals Topoisomerase III Acts Upstream of Rad53p in the S-Phase DNA Damage Checkpoint

2001 ◽  
Vol 21 (21) ◽  
pp. 7150-7162 ◽  
Author(s):  
Ronjon K. Chakraverty ◽  
Jonathan M. Kearsey ◽  
Thomas J. Oakley ◽  
Muriel Grenon ◽  
Maria-Angeles de la Torre Ruiz ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Dna Damage ◽  
Cellular Response ◽  
Cell Cycle Progression ◽  
Uv Light ◽  
S Phase ◽  
Dna Damaging Agents ◽  
Topoisomerase Iii ◽  
Rad53 Kinase ◽  
S Phase Checkpoint

ABSTRACT Deletion of the Saccharomyces cerevisiae TOP3gene, encoding Top3p, leads to a slow-growth phenotype characterized by an accumulation of cells with a late S/G2content of DNA (S. Gangloff, J. P. McDonald, C. Bendixen, L. Arthur, and R. Rothstein, Mol. Cell. Biol. 14:8391–8398, 1994). We have investigated the function of TOP3 during cell cycle progression and the molecular basis for the cell cycle delay seen in top3Δ strains. We show that top3Δ mutants exhibit a RAD24-dependent delay in the G2 phase, suggesting a possible role for Top3p in the resolution of abnormal DNA structures or DNA damage arising during S phase. Consistent with this notion,top3Δ strains are sensitive to killing by a variety of DNA-damaging agents, including UV light and the alkylating agent methyl methanesulfonate, and are partially defective in the intra-S-phase checkpoint that slows the rate of S-phase progression following exposure to DNA-damaging agents. This S-phase checkpoint defect is associated with a defect in phosphorylation of Rad53p, indicating that, in the absence of Top3p, the efficiency of sensing the existence of DNA damage or signaling to the Rad53 kinase is impaired. Consistent with a role for Top3p specifically during S phase, top3Δ mutants are sensitive to the replication inhibitor hydroxyurea, expression of the TOP3 mRNA is activated in late G1 phase, and DNA damage checkpoints operating outside of S phase are unaffected by deletion of TOP3. All of these phenotypic consequences of loss of Top3p function are at least partially suppressed by deletion of SGS1, the yeast homologue of the human Bloom's and Werner's syndrome genes. These data implicate Top3p and, by inference, Sgs1p in an S-phase-specific role in the cellular response to DNA damage. A model proposing a role for these proteins in S phase is presented.

scholarly journals Induction of the Mouse Ribonucleotide Reductase R1 and R2 Genes in Response to DNA Damage by UV Light

1996 ◽  
Vol 271 (39) ◽  
pp. 23698-23704 ◽  
Author(s):  
Dmitri Filatov ◽  
Stefan Björklund ◽  
Erik Johansson ◽  
Lars Thelander
Keyword(s):  
Dna Damage ◽  
Ribonucleotide Reductase ◽  
Uv Light

scholarly journals Upstream regulatory sequences of the yeast RNR2 gene include a repression sequence and an activation site that binds the RAP1 protein.

1989 ◽  
Vol 9 (12) ◽  
pp. 5359-5372 ◽  
Author(s):  
H K Hurd ◽  
J W Roberts
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Ribonucleotide Reductase ◽  
Small Subunit ◽  
Methyl Methanesulfonate ◽  
Regulatory Sequences ◽  
Start Site ◽  
Transcription Start ◽  
Dna Damaging Agents ◽  
Number Of Genes ◽  
Upstream Regulatory Sequences

The small subunit of ribonucleotide reductase in Saccharomyces cerevisiae (RNR2) was induced 3- to 20-fold by a variety of DNA-damaging agents. Induction of the RNR2 transcript by at least one of these agents, methyl methanesulfonate, did not require protein synthesis. To identify sequences involved in the regulation of RNR2, we introduced deletions upstream of the transcription start site. Sequences required for induction were contained within a 200-base-pair region that could confer methyl methanesulfonate inducibility on the heterologous CYC1 promoter. This region contained a repression sequence and at least two positive activation sites. One of these activation sites bound RAP1, a protein known to associate with mating-type silencers and the upstream activation sequences of a number of genes. The behavior of deletions of the repression sequence suggests that induction of RNR2 may occur, at least in part, through relief of repression.

Identification of a Topoisomerase I Mutant, scsA1, as an Extragenic Suppressor of a Mutation in scaANBS1, the Apparent Homolog of Human Nibrin in Aspergillus nidulans

Genetics ◽  
2003 ◽  
Vol 164 (3) ◽  
pp. 935-945 ◽  
Author(s):  
Marcia R Z Kress Fagundes ◽  
Larissa Fernandes ◽  
Marcela Savoldi ◽  
Steven D Harris ◽  
Maria H S Goldman ◽  
...  
Keyword(s):  
Dna Damage ◽  
Aspergillus Nidulans ◽  
Topoisomerase I ◽  
Cell Cycle Checkpoint ◽  
Dna Double Strand Breaks ◽  
Dna Damaging Agents ◽  
Cycle Checkpoint ◽  
Increased Sensitivity ◽  
Cellular Dna ◽  
Extragenic Suppressors

Abstract The Mre11-Rad50-Nbs1 protein complex has emerged as a central player in the human cellular DNA damage response, and recent observations suggest that these proteins are at least partially responsible for the linking of DNA damage detection to DNA repair and cell cycle checkpoint functions. Mutations in scaANBS1, which encodes the apparent homolog of human nibrin in Aspergillus nidulans, inhibit growth in the presence of the antitopoisomerase I drug camptothecin. This article describes the selection and characterization of extragenic suppressors of the scaA1 mutation, with the aim of identifying other proteins that interfere with the pathway or complex in which the ScaA would normally be involved. Fifteen extragenic suppressors of the scaA1 mutation were isolated. The topoisomerase I gene can complement one of these suppressors. Synergistic interaction between the scaANBS1 and scsATOP1 genes in the presence of DNA-damaging agents was observed. Overexpression of topoisomerase I in the scaA1 mutant causes increased sensitivity to DNA-damaging agents. The scsATOP1 and the scaANBS1 gene products could functionally interact in pathways that either monitor or repair DNA double-strand breaks.

Identification of the gene for the yeast ribonucleotide reductase small subunit and its inducibility by methyl methanesulfonate

1987 ◽  
Vol 7 (10) ◽  
pp. 3673-3677
Author(s):  
H K Hurd ◽  
C W Roberts ◽  
J W Roberts
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Dna Damage ◽  
Ribonucleotide Reductase ◽  
Alkylating Agent ◽  
Small Subunit ◽  
Methyl Methanesulfonate

We have identified, cloned, and sequenced the gene for the small subunit of ribonucleotide diphosphate reductase of Saccharomyces cerevisiae. The protein and its transcript are induced about 10-fold by the alkylating agent methyl methanesulfonate, a result which suggests that the gene is induced by DNA damage.

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