scholarly journals Mutations in PMR1 suppress oxidative damage in yeast cells lacking superoxide dismutase.

1995 ◽  
Vol 15 (3) ◽  
pp. 1382-1388 ◽  
Author(s):  
P J Lapinskas ◽  
K W Cunningham ◽  
X F Liu ◽  
G R Fink ◽  
V C Culotta
Keyword(s):  
Superoxide Dismutase ◽  
Oxidative Damage ◽  
Eukaryotic Cell ◽  
Yeast Cells ◽  
Manganese Ions ◽  
Wild Type ◽  
Sod1 Gene ◽  
Gene Encoding ◽  
Manganese Ion ◽  
P Type

Mutants of Saccharomyces cerevisiae lacking a functional SOD1 gene encoding Cu/Zn superoxide dismutase (SOD) are sensitive to atmospheric levels of oxygen and are auxotrophic for lysine and methionine when grown in air. We have previously shown that these defects of SOD-deficient yeast cells can be overcome through mutations in either the BSD1 or BSD2 (bypass SOD defects) gene. In this study, the wild-type allele of BSD1 was cloned by functional complementation and was physically mapped to the left arm of chromosome VII. BSD1 is identical to PMR1, encoding a member of the P-type ATPase family that localizes to the Golgi apparatus. PMR1 is thought to function in calcium metabolism, and we provide evidence that PMR1 also participates in the homeostasis of manganese ions. Cells lacking a functional PMR1 gene accumulate elevated levels of intracellular manganese and are also extremely sensitive to manganese ion toxicity. We demonstrate that mutations in PMR1 bypass SOD deficiency through a mechanism that depends on extracellular manganese. Collectively, these findings indicate that oxidative damage in a eukaryotic cell can be prevented through alterations in manganese homeostasis.

scholarly journals Cell fusion during yeast mating requires high levels of a-factor mating pheromone.

1996 ◽  
Vol 135 (6) ◽  
pp. 1727-1739 ◽  
Author(s):  
V Brizzio ◽  
A E Gammie ◽  
G Nijbroek ◽  
S Michaelis ◽  
M D Rose
Keyword(s):  
Cell Wall ◽  
Cell Fusion ◽  
Plasma Membranes ◽  
Yeast Cells ◽  
Wild Type ◽  
Mating Pheromone ◽  
Gene Encoding ◽  
Fluorescence And Electron Microscopy ◽  
Identical Cell ◽  
In Trans

During conjugation, two yeast cells fuse to form a single zygote. Cell fusion requires extensive remodeling of the cell wall, both to form a seal between the two cells and to remove the intervening material. The two plasma membranes then fuse to produce a continuous cytoplasm. We report the characterization of two cell fusion defective (Fus-) mutants, fus5 and fus8, isolated previously in our laboratory. Fluorescence and electron microscopy demonstrated that the fus5 and fus8 mutant zygotes were defective for cell wall remodeling/removal but not plasma membrane fusion. Strikingly, fus5 and fus8 were a specific; both mutations caused the mutant phenotype when present in the MATa parent but not in the MAT alpha parent. Consistent with an a-specific defect, the fus5 and fus8 mutants produced less a-factor than the isogenic wild-type strain. FUS5 and FUS8 were determined to be allelic to AXL1 and RAM1, respectively, two genes known to be required for biogenesis of a-factor. Several experiments demonstrated that the partial defect in a-factor production resulted in the Fus- phenotype. First, overexpression of a-factor in the fus mutants suppressed the Fus- defect. Second, matings to an MAT alpha partner supersensitive to mating pheromone (sst2 delta) suppressed the Fus- defect in trans. Finally, the gene encoding a-factor, MFA1, was placed under the control of a repressible promoter; reduced levels of wild-type a-factor caused an identical cell fusion defect during mating. We conclude that high levels of pheromone are required as one component of the signal for prezygotes to initiate cell fusion.

scholarly journals Regulation of Salt Tolerance by Torulaspora delbrueckii Calcineurin Target Crz1p

2006 ◽  
Vol 5 (3) ◽  
pp. 469-479 ◽  
Author(s):  
Maria Jose Hernandez-Lopez ◽  
Joaquin Panadero ◽  
Jose Antonio Prieto ◽  
Francisca Randez-Gil
Keyword(s):  
Salt Tolerance ◽  
Transcriptional Activation ◽  
Molecular Mechanisms ◽  
Divalent Cations ◽  
Immunosuppressive Agents ◽  
Wild Type ◽  
Gene Encoding ◽  
Torulaspora Delbrueckii ◽  
Regulatory Circuits ◽  
P Type

ABSTRACT Recently, the academic interest in the yeast Torulaspora delbrueckii has increased notably due to its high resistance to several types of stress, including salt and osmotic imbalance. However, the molecular mechanisms underlying these unusual properties are poorly understood. In Saccharomyces cerevisiae, the high-salt response is mediated by calcineurin, a conserved Ca2+/calmodulin-modulated protein phosphatase that regulates the transcriptional factor Crz1p. Here, we cloned the T. delbrueckii TdCRZ1 gene, which encodes a putative zinc finger transcription factor homologue to Crz1p. Consistent with this, overexpression of TdCRZ1 enhanced the salt tolerance of S. cerevisiae wild-type cells and suppressed the sensitivity phenotype of cnb1Δ and crz1Δ mutants to monovalent and divalent cations. However, T. delbrueckii cells lacking TdCrz1p showed phenotypes distinct from those previously observed in S. cerevisiae crz1Δ mutants. Quite remarkably, Tdcrz1-null cells were insensitive to high Na+ and were more Li+ tolerant than wild-type cells. Clearly, TdCrz1p was not required for the salt-induced transcriptional activation of the TdENA1 gene, encoding a putative P-type ATPase homologue to the main S. cerevisiae Na+ pump ENA1. Furthermore, T. delbrueckii cells were insensitive to the immunosuppressive agents FK506 and cyclosporine A, both in the presence and in the absence of NaCl. Signaling through the calcineurin/Crz1 pathway appeared to be essential only on high-Ca2+/Mn2+ media. Hence, T. delbrueckii and S. cerevisiae differ in the regulatory circuits and mechanisms that drive the adaptive response to salt stress.

scholarly journals l-Proline Accumulation and Freeze Tolerance in Saccharomyces cerevisiae Are Caused by a Mutation in the PRO1 Gene Encoding γ-Glutamyl Kinase

2003 ◽  
Vol 69 (1) ◽  
pp. 212-219 ◽  
Author(s):  
Yuko Morita ◽  
Shigeru Nakamori ◽  
Hiroshi Takagi
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Genomic Library ◽  
Reductase Activity ◽  
Freeze Tolerance ◽  
Proline Accumulation ◽  
Yeast Cells ◽  
Single Amino Acid ◽  
Wild Type ◽  
Gene Encoding

ABSTRACT We previously isolated a mutant which showed a high tolerance to freezing that correlated with higher levels of intracellular l-proline derived from l-proline analogue-resistant mutants. The mutation responsible for the analogue resistance and l-proline accumulation was a single nuclear dominant mutation. By introducing the mutant-derived genomic library into a non-l-proline-utilizing strain, the mutant was found to carry an allele of the wild-type PRO1 gene encoding γ-glutamyl kinase, which resulted in a single amino acid replacement; Asp (GAC) at position 154 was replaced by Asn (AAC). Interestingly, the allele of PRO1 was shown to enhance the activities of γ-glutamyl kinase and γ-glutamyl phosphate reductase, both of which catalyze the first two steps of l-proline synthesis from l-glutamate and which together may form a complex in vivo. When cultured in liquid minimal medium, yeast cells expressing the mutated γ-glutamyl kinase were found to accumulate intracellular l-proline and showed a prominent increase in cell viability after freezing at −20°C compared to the viability of cells harboring the wild-type PRO1 gene. These results suggest that the altered γ-glutamyl kinase results in stabilization of the complex or has an indirect effect on γ-glutamyl phosphate reductase activity, which leads to an increase in l-proline production in Saccharomyces cerevisiae. The approach described in this paper could be a practical method for breeding novel freeze-tolerant yeast strains.

scholarly journals Role of Manganese-Containing Superoxide Dismutase in Oxidative Stress and Virulence of Streptococcus pneumoniae

2000 ◽  
Vol 68 (5) ◽  
pp. 2819-2826 ◽  
Author(s):  
Hasan Yesilkaya ◽  
Aras Kadioglu ◽  
Neill Gingles ◽  
Janet E. Alexander ◽  
Tim J. Mitchell ◽  
...  
Keyword(s):  
Superoxide Dismutase ◽  
Streptococcus Pneumoniae ◽  
Anaerobic Growth ◽  
Wild Type ◽  
Gene Encoding ◽  
Lower Growth ◽  
Redox Active ◽  
Lower Growth Rate

ABSTRACT Streptococcus pneumoniae was shown to contain two types of superoxide dismutase, MnSOD and FeSOD. Levels of MnSOD increased during growth in an aerobic environment. The sodA gene, encoding MnSOD, of virulent S. pneumoniae type 2 strain D39 was inactivated to give mutant D39HY1. Aerobically, D39HY1 had a lower growth rate than the wild type and exhibited susceptibility to the redox-active compound paraquat, but anaerobic growth of D39HY1 was identical to that of the wild type. Virulence studies showed that the median survival time of mice infected intranasally with D39HY1 was significantly longer than that of mice infected with the wild-type pneumococcus. In contrast to the wild type, D39HY1 did not multiply in lungs during the first 24 h but thereafter grew at the same rate as the wild type. Appearance in the bloodstream was also delayed, but growth in the blood was unimpaired by the sodA mutation. The pattern of inflammation in lungs infected with D39HY1 differed from that in wild-type-infected mice. After infection with D39HY1, neutrophils were densely packed around bronchioles, in contrast to the wild-type infection, where neutrophils were more diffusely localized.

scholarly journals Requirement of the self-glucosylating initiator proteins Glg1p and Glg2p for glycogen accumulation in Saccharomyces cerevisiae.

1995 ◽  
Vol 15 (12) ◽  
pp. 6632-6640 ◽  
Author(s):  
C Cheng ◽  
J Mu ◽  
I Farkas ◽  
D Huang ◽  
M G Goebl ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Glycogen Synthase ◽  
Eukaryotic Cell ◽  
The Self ◽  
Yeast Cells ◽  
Wild Type ◽  
Glycogen Accumulation ◽  
Glycogen Biosynthesis ◽  
Mammalian Protein

Glycogen, a branched polymer of glucose, is a storage molecule whose accumulation is under rigorous nutritional control in many cells. We report the identification of two Saccharomyces cerevisiae genes, GLG1 and GLG2, whose products are implicated in the biogenesis of glycogen. These genes encode self-glucosylating proteins that in vitro can act as primers for the elongation reaction catalyzed by glycogen synthase. Over a region of 258 residues, the Glg proteins have 55% sequence identify to each other and approximately 33% identity to glycogenin, a mammalian protein postulated to have a role in the initiation of glycogen biosynthesis. Yeast cells defective in either GLG1 or GLG2 are similar to the wild type in their ability to accumulate glycogen. Disruption of both genes results in the inability of the cells to synthesize glycogen despite normal levels of glycogen synthase. These results suggest that a self-glucosylating protein is required for glycogen biosynthesis in a eukaryotic cell. The activation state of glycogen synthase in glg1 glg2 cells is suppressed, suggesting that the Glg proteins may additionally influence the phosphorylation state of glycogen synthase.

scholarly journals Copper/Zinc-Superoxide Dismutase Is Required for Oxytetracycline Resistance of Saccharomyces cerevisiae

2000 ◽  
Vol 182 (1) ◽  
pp. 76-80 ◽  
Author(s):  
Simon V. Avery ◽  
Srividya Malkapuram ◽  
Carolina Mateus ◽  
Kimberly S. Babb
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Lipid Peroxidation ◽  
Superoxide Dismutase ◽  
Oxidative Damage ◽  
Protein Oxidation ◽  
Wild Type ◽  
Copper Zinc Superoxide Dismutase ◽  
Zinc Superoxide Dismutase ◽  
Copper Zinc

ABSTRACT Saccharomyces cerevisiae, along with other eukaryotes, is resistant to tetracyclines. We found that deletion ofSOD1 (encoding Cu/Zn superoxide dismutase) renderedS. cerevisiae hypersensitive to oxytetracycline (OTC): asod1Δ mutant exhibited a >95% reduction in colony-forming ability at an OTC concentration of 20 μg ml−1, whereas concentrations of up to 1,000 μg ml−1 had no effect on the growth of the wild type. OTC resistance was restored in the sod1Δ mutant by complementation with wild-type SOD1. The effect of OTC appeared to be cytotoxic and was not evident in a ctt1Δ (cytosolic catalase) mutant or in the presence of tetracycline.SOD1 transcription was not induced by OTC, suggesting that constitutive SOD1 expression is sufficient for wild-type OTC resistance. OTC uptake levels in wild-type and sod1Δ strains were similar. However, lipid peroxidation and protein oxidation were both enhanced during exposure of the sod1Δ mutant, but not the wild type, to OTC. We propose that Sod1p protects S. cerevisiae against a mode of OTC action that is dependent on oxidative damage.

scholarly journals Suppression of oxidative damage by Saccharomyces cerevisiae ATX2, which encodes a manganese-trafficking protein that localizes to Golgi-like vesicles.

1996 ◽  
Vol 16 (11) ◽  
pp. 6303-6312 ◽  
Author(s):  
S J Lin ◽  
V C Culotta
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Golgi Apparatus ◽  
Oxidative Damage ◽  
Oxygen Toxicity ◽  
Subcellular Fractionation ◽  
Yeast Cells ◽  
Manganese Ions ◽  
Copper Zinc Superoxide Dismutase ◽  
Punctate Pattern ◽  
Vesicular Compartment

Oxygen toxicity in Saccharomyces cerevisiae lacking the copper/zinc superoxide dismutase (SOD1) can be suppressed by overexpression of the S. cerevisiae ATX2 gene. Multiple copies of ATX2 were found to reverse the aerobic auxotrophies of sod1(delta) mutants for lysine and methionine and also to enhance the resistance of these yeast strains to paraquat and atmospheric levels of oxygen. ATX2 encodes a novel 34.4-kDa polypeptide with a number of potential membrane-spanning domains. Our studies indicate that Atx2p localizes to the membrane of a vesicular compartment in yeast cells reminiscent of the Golgi apparatus. With indirect immunofluorescence microscopy, Atx2p exhibited a punctate pattern of staining typical of the Golgi apparatus, and upon subcellular fractionation, Atx2p colocalized with a biochemical marker for the yeast Golgi apparatus. We demonstrate here that this vesicle protein normally functions in the homeostasis of manganese ions and that this role in metal metabolism is necessary for the ATX1 suppression of SOD1 deficiency. First, overexpression of ATX2 caused cells to accumulate increased levels of manganese. Second, a deletion in ATX2 caused a decrease in the apparent available level of intracellular manganese and caused sod1(delta) mutants to become dependent upon exogenous manganese for aerobic growth. Third, ATX2 was incapable of suppressing oxidative damage in cells depleted of manganese ions or lacking the plasma membrane transporter for manganese. The effect of ATX2 overexpression on manganese accumulation and oxygen resistance is similar to what we have previously reported for mutations in PMR1, which encodes a manganese-trafficking protein that also resides in a vesicular compartment. Our studies are consistent with a model in which Atx2p and Pmr1p work in opposite directions to control manganese homeostasis.

scholarly journals Elevated Mutation Frequency in Surviving Populations of Carbon-Starved rpoS-Deficient Pseudomonas putida Is Caused by Reduced Expression of Superoxide Dismutase and Catalase

2009 ◽  
Vol 191 (11) ◽  
pp. 3604-3614 ◽  
Author(s):  
Kairi Tarassova ◽  
Radi Tegova ◽  
Andres Tover ◽  
Riho Teras ◽  
Mariliis Tark ◽  
...  
Keyword(s):  
Superoxide Dismutase ◽  
Oxidative Damage ◽  
Pseudomonas Putida ◽  
Mutation Frequency ◽  
Test System ◽  
Base Substitution ◽  
Repair System ◽  
Wild Type ◽  
Large Deletions ◽  
In The Wild

ABSTRACTRpoS is a bacterial sigma factor of RNA polymerase which is involved in the expression of a large number of genes to facilitate survival under starvation conditions and other stresses. The results of our study demonstrate that the frequency of emergence of base substitution mutants is significantly increased in long-term-starved populations ofrpoS-deficientPseudomonas putidacells. The increasing effect of the lack of RpoS on the mutation frequency became apparent in both a plasmid-based test system measuring Phe+reversion and a chromosomalrpoBsystem detecting rifampin-resistant mutants. The elevated mutation frequency coincided with the death of about 95% of the cells in a population ofrpoS-deficientP.putida. Artificial overexpression of superoxide dismutase or catalase in therpoS-deficient strain restored the survival of cells and resulted in a decline in the mutation frequency. This indicated that, compared to wild-type bacteria,rpoS-deficient cells are less protected against damage caused by reactive oxygen species. 7,8-Dihydro-8-oxoguanine (GO) is known to be one of the most stable and frequent base modifications caused by oxygen radical attack on DNA. However, the spectrum of base substitution mutations characterized inrpoS-deficientP.putidawas different from that in bacteria lacking the GO repair system: it was broader and more similar to that identified in the wild-type strain. Interestingly, the formation of large deletions was also accompanied by a lack of RpoS. Thus, the accumulation of DNA damage other than GO elevates the frequency of mutation in these bacteria. It is known that oxidative damage of proteins and membrane components, but not that of DNA, is a major reason for the death of cells. Since the increased mutation frequency was associated with a decline in the viability of bacteria, we suppose that the elevation of the mutation frequency in the surviving population of carbon-starvedrpoS-deficientP.putidamay be caused both by oxidative damage of DNA and enzymes involved in DNA replication and repair fidelity.

Catalase expression impairs oxidative stress-mediated signalling inTrypanosoma cruzi

Parasitology ◽  
2017 ◽  
Vol 144 (11) ◽  
pp. 1498-1510 ◽  
Author(s):  
ANNA CLÁUDIA GUIMARÃES FREIRE ◽  
CERES LUCIANA ALVES ◽  
GRAZIELLE RIBEIRO GOES ◽  
BRUNO CARVALHO RESENDE ◽  
NILMAR SILVIO MORETTI ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Hydrogen Peroxide ◽  
Superoxide Dismutase ◽  
Life Cycle ◽  
Trypanothione Reductase ◽  
Wild Type ◽  
Gene Encoding ◽  
Oxidative Stresses ◽  
Low Concentrations ◽  
Shed Light

SUMMARYTrypanosoma cruziis exposed to oxidative stresses during its life cycle, and amongst the strategies employed by this parasite to deal with these situations sits a peculiar trypanothione-dependent antioxidant system. Remarkably,T. cruzi’s antioxidant repertoire does not include catalase. In an attempt to shed light on what are the reasons by which this parasite lacks this enzyme, aT. cruzicell line stably expressing catalase showed an increased resistance to hydrogen peroxide (H2O2) when compared with wild-type cells. Interestingly, preconditioning carried out with low concentrations of H2O2led untransfected parasites to be as much resistant to this oxidant as cells expressing catalase, but did not induce the same level of increased resistance in the latter ones. Also, presence of catalase decreased trypanothione reductase and increased superoxide dismutase levels inT. cruzi, resulting in higher levels of residual H2O2after challenge with this oxidant. Although expression of catalase contributed to elevated proliferation rates ofT. cruziinRhodnius prolixus, it failed to induce a significant increase of parasite virulence in mice. Altogether, these results indicate that the absence of a gene encoding catalase inT. cruzihas played an important role in allowing this parasite to develop a shrill capacity to sense and overcome oxidative stress.

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