scholarly journals The Saccharomyces cerevisiae Homologues of Endonuclease III from Escherichia coli, Ntg1 and Ntg2, Are Both Required for Efficient Repair of Spontaneous and Induced Oxidative DNA Damage in Yeast

1999 ◽  
Vol 19 (5) ◽  
pp. 3779-3787 ◽  
Author(s):  
Ingrun Alseth ◽  
Lars Eide ◽  
Manuela Pirovano ◽  
Torbjørn Rognes ◽  
Erling Seeberg ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Saccharomyces Cerevisiae ◽  
Dna Damage ◽  
Oxidative Dna Damage ◽  
High Efficiency ◽  
Intracellular Localization ◽  
Genome Project ◽  
Yeast Genome ◽  
Enzyme Analysis ◽  
Endonuclease Iii

ABSTRACT Endonuclease III from Escherichia coli is the prototype of a ubiquitous DNA repair enzyme essential for the removal of oxidized pyrimidine base damage. The yeast genome project has revealed the presence of two genes in Saccharomyces cerevisiae,NTG1 and NTG2, encoding proteins with similarity to endonuclease III. Both contain the highly conserved helix-hairpin-helix motif, whereas only one (Ntg2) harbors the characteristic iron-sulfur cluster of the endonuclease III family. We have characterized these gene functions by mutant and enzyme analysis as well as by gene expression and intracellular localization studies. Targeted gene disruption of NTG1 and NTG2produced mutants with greatly increased spontaneous and hydrogen peroxide-induced mutation frequency relative to the wild type, and the mutation response was further increased in the double mutant. Both enzymes were found to remove thymine glycol and 2,6-diamino-4-hydroxy-5-N-methylformamidopyrimidine (faPy) residues from DNA with high efficiency. However, on UV-irradiated DNA, saturating concentrations of Ntg2 removed only half of the cytosine photoproducts released by Ntg1. Conversely, 5-hydroxycytosine was removed efficiently only by Ntg2. The enzymes appear to have different reaction modes, as judged from much higher affinity of Ntg2 for damaged DNA and more efficient borhydride trapping of Ntg1 to abasic sites in DNA despite limited DNA binding. Northern blot and promoter fusion analysis showed that NTG1 is inducible by cell exposure to DNA-damaging agents, whereas NTG2 is constitutively expressed. Ntg2 appears to be a nuclear enzyme, whereas Ntg1 was sorted both to the nucleus and to the mitochondria. We conclude that functions of both NTG1 and NTG2 are important for removal of oxidative DNA damage in yeast.

scholarly journals A highly conserved endonuclease activity present in Escherichia coli, bovine, and human cells recognizes oxidative DNA damage at sites of pyrimidines.

1987 ◽  
Vol 7 (1) ◽  
pp. 26-32 ◽  
Author(s):  
P W Doetsch ◽  
W D Henner ◽  
R P Cunningham ◽  
J H Toney ◽  
D E Helland
Keyword(s):  
Escherichia Coli ◽  
Dna Damage ◽  
Oxidative Dna Damage ◽  
Uv Light ◽  
Human Fibroblasts ◽  
Human Cells ◽  
Sequence Specificity ◽  
Endonuclease Activity ◽  
E Coli ◽  
Endonuclease Iii

We have compared the sites of nucleotide incision on DNA damaged by oxidizing agents when cleavage is mediated by either Escherichia coli endonuclease III or an endonuclease present in bovine and human cells. E. coli endonuclease III, the bovine endonuclease isolated from calf thymus, and the human endonuclease partially purified from HeLa and CEM-C1 lymphoblastoid cells incised DNA damaged with osmium tetroxide, ionizing radiation, or high doses of UV light at sites of pyrimidines. For each damaging agent studied, regardless of whether the E. coli, bovine, or human endonuclease was used, the same sequence specificity of cleavage was observed. We detected this endonuclease activity in a variety of human fibroblasts derived from normal individuals as well as individuals with the DNA repair deficiency diseases ataxia telangiectasia and xeroderma pigmentosum. The highly conserved nature of such a DNA damage-specific endonuclease suggests that a common pathway exists in bacteria, humans, and other mammals for the reversal of certain types of oxidative DNA damage.

A highly conserved endonuclease activity present in Escherichia coli, bovine, and human cells recognizes oxidative DNA damage at sites of pyrimidines

1987 ◽  
Vol 7 (1) ◽  
pp. 26-32
Author(s):  
P W Doetsch ◽  
W D Henner ◽  
R P Cunningham ◽  
J H Toney ◽  
D E Helland
Keyword(s):  
Escherichia Coli ◽  
Dna Damage ◽  
Oxidative Dna Damage ◽  
Uv Light ◽  
Human Fibroblasts ◽  
Human Cells ◽  
Sequence Specificity ◽  
Endonuclease Activity ◽  
E Coli ◽  
Endonuclease Iii

We have compared the sites of nucleotide incision on DNA damaged by oxidizing agents when cleavage is mediated by either Escherichia coli endonuclease III or an endonuclease present in bovine and human cells. E. coli endonuclease III, the bovine endonuclease isolated from calf thymus, and the human endonuclease partially purified from HeLa and CEM-C1 lymphoblastoid cells incised DNA damaged with osmium tetroxide, ionizing radiation, or high doses of UV light at sites of pyrimidines. For each damaging agent studied, regardless of whether the E. coli, bovine, or human endonuclease was used, the same sequence specificity of cleavage was observed. We detected this endonuclease activity in a variety of human fibroblasts derived from normal individuals as well as individuals with the DNA repair deficiency diseases ataxia telangiectasia and xeroderma pigmentosum. The highly conserved nature of such a DNA damage-specific endonuclease suggests that a common pathway exists in bacteria, humans, and other mammals for the reversal of certain types of oxidative DNA damage.

scholarly journals ZDS1 and ZDS2, genes whose products may regulate Cdc42p in Saccharomyces cerevisiae.

1996 ◽  
Vol 16 (10) ◽  
pp. 5264-5275 ◽  
Author(s):  
E Bi ◽  
J R Pringle
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Amino Acids ◽  
Intracellular Signaling ◽  
Double Mutant ◽  
Genome Project ◽  
Yeast Genome ◽  
Glutathione S Transferase ◽  
Gene Encoding ◽  
Biological Phenomena ◽  
Wide Range

A genetic screen for GTPase-activating proteins (GAPs) or other negative regulators of the Rac/Rho family GTPase Cdc42p in Saccharomyces cerevisiae identified ZDS1, a gene encoding a protein of 915 amino acids. Sequence from the yeast genome project identified a homolog, ZDS2, whose predicted product of 942 amino acids is 38% identical in sequence to Zds1p. Zds1p and Zds2p have no detectable homology to known Rho-GAPs or to other known proteins. However, by several assays, it appears that overexpression of either Zds1p or Zds2p decreases the level of Cdc42p activity. Deletion analysis also suggests that Zds1p and Zds2p are at least partially overlapping in function. Deletion of ZDS2 produced no obvious phenotype, and deletion of ZDS1 produced no obvious phenotype other than a mild effect on cell shape. However, the zds1 zds2 double mutant grew slowly with an apparent mitotic delay and produced elongated cells and buds with other evidence of abnormal morphogenesis. A glutathione S-transferase-Zds1p fusion protein that fully complemented the double mutant localized to presumptive bud sites and the tips of small buds. The similarity of this localization to that of Cdc42p suggests that Zds1p may interact directly with Cdc42p. As ZDS1 and ZDS2 have recently been identified also by numerous other groups studying a wide range of biological phenomena, the roles of Cdc42p in intracellular signaling may be more diverse than has previously been appreciated.

scholarly journals Dynamic Compartmentalization of Base Excision Repair Proteins in Response to Nuclear and Mitochondrial Oxidative Stress

2008 ◽  
Vol 29 (3) ◽  
pp. 794-807 ◽  
Author(s):  
Lyra M. Griffiths ◽  
Dan Swartzlander ◽  
Kellen L. Meadows ◽  
Keith D. Wilkinson ◽  
Anita H. Corbett ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Dna Damage ◽  
Dna Repair ◽  
Base Excision Repair ◽  
Oxidative Dna Damage ◽  
Excision Repair ◽  
Intracellular Localization ◽  
Genotoxic Stress ◽  
Mitochondrial Oxidative Stress ◽  
Base Excision

ABSTRACT DNAs harbored in both nuclei and mitochondria of eukaryotic cells are subject to continuous oxidative damage resulting from normal metabolic activities or environmental insults. Oxidative DNA damage is primarily reversed by the base excision repair (BER) pathway, initiated by N-glycosylase apurinic/apyrimidinic (AP) lyase proteins. To execute an appropriate repair response, BER components must be distributed to accommodate levels of genotoxic stress that may vary considerably between nuclei and mitochondria, depending on the growth state and stress environment of the cell. Numerous examples exist where cells respond to signals, resulting in relocalization of proteins involved in key biological transactions. To address whether such dynamic localization contributes to efficient organelle-specific DNA repair, we determined the intracellular localization of the Saccharomyces cerevisiae N-glycosylase/AP lyases, Ntg1 and Ntg2, in response to nuclear and mitochondrial oxidative stress. Fluorescence microscopy revealed that Ntg1 is differentially localized to nuclei and mitochondria, likely in response to the oxidative DNA damage status of the organelle. Sumoylation is associated with targeting of Ntg1 to nuclei containing oxidative DNA damage. These studies demonstrate that trafficking of DNA repair proteins to organelles containing high levels of oxidative DNA damage may be a central point for regulating BER in response to oxidative stress.

scholarly journals Oxidative DNA damage is instrumental in hyperreplication stress-induced inviability of Escherichia coli

2014 ◽  
Vol 42 (21) ◽  
pp. 13228-13241 ◽  
Author(s):  
Godefroid Charbon ◽  
Louise Bjørn ◽  
Belén Mendoza-Chamizo ◽  
Jakob Frimodt-Møller ◽  
Anders Løbner-Olesen
Keyword(s):  
Escherichia Coli ◽  
Dna Damage ◽  
Oxidative Dna Damage

scholarly journals Brewer’s Spent Grains Protects against Oxidative DNA Damage in Saccharomyces cerevisiae

2017 ◽  
Vol 9 (7) ◽  
pp. 12
Author(s):  
Manuela M. Moreira ◽  
Daniel O. Carvalho ◽  
Rui Oliveira ◽  
Björn Johansson ◽  
Luís F. Guido
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Flow Cytometry ◽  
Dna Damage ◽  
Oxidative Dna Damage ◽  
Yeast Cells ◽  
Protective Effects ◽  
Genotoxic Effects ◽  
Barley Malt ◽  
Fold Reduction ◽  
Spent Grain

Brewer’s spent grain (BSG), obtained from barley malt during brewing, contains high amounts of phenolic acids, predominantly ferulic and p-coumaric acids. The protective effects of BSG extracts against oxidative DNA damage induced by H2O2 in Saccharomyces cerevisiae cells were investigated using an optimized yeast comet assay and flow cytometry. The results indicated that BSG extracts from black malt exhibited a 5-fold reduction in the genotoxic effects of H2O2, compared to the 2-fold decrease by the BSG extracts from pilsen malts. Flow cytometry analysis with dichlorofluorescein diacetate demonstrated that the intracellular oxidation of S. cerevisiae is also reduced to approximately 50% in the presence of 20-fold diluted BSG extracts. BSG extracts obtained from pilsen and black malt types exert dose-dependent protective properties against the genotoxic effects induced by ROS and decrease intracellular oxidation of yeast cells.

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