Biomedical subjects

Pedro Moradas-Ferreira

Publications and source records attributed to Pedro Moradas-Ferreira.

5 recordsLinked to original sources

PubMed2003

Mnsod overexpression extends the yeast chronological (G(0)) life span but acts independently of Sir2p histone deacetylase to shorten the replicative life span of dividing cells.

Studies in Drosophila and Caenorhabditis elegans have shown increased longevity with the increased free radical scavenging that accompanies overexpression of oxidant-scavenging enzymes. This study used yeast, another model for aging research, to probe the effects of overexpressing the major activity protecting against superoxide generated by the mitochondrial respiratory chain. Manganese superoxide dismutase (MnSOD) overexpression increased chronological life span (optimized survival of stationary (G(0)) yeast over time), showing this is a survival ultimately limited by oxidative stress. In contrast, the same overexpression dramatically reduced the replicative life span of dividing cells (the number of daughter buds produced by each newly born mother cell). This reduction in the generational life span by MnSOD overexpression was greater than that generated by loss of the major redox-responsive regulator of the yeast replicative life span, NAD+-dependent Sir2p histone deacetylase. It was also independent of the latter activity. Expression of a mitochondrially targeted green fluorescent protein in the MnSOD overexpressor revealed that the old mother cells of this overexpressor, which had divided for a few generations, were defective in segregation of the mitochondrion from the mother to daughter. Mitochondrial defects are, therefore, the probable reason that MnSOD overexpression shortens replicative life span.

Cell Division↗

PubMed2003

H2O2, but not menadione, provokes a decrease in the ATP and an increase in the inosine levels in Saccharomyces cerevisiae. An experimental and theoretical approach.

When Saccharomyces cerevisiae cells, grown in galactose, glucose or mannose, were treated with 1.5 mm hydrogen peroxide (H2O2) for 30 min, an important decrease in the ATP, and a less extensive decrease in the GTP, CTP, UTP and ADP-ribose levels was estimated. Concomitantly a net increase in the inosine levels was observed. Treatment with 83 mm menadione promoted the appearance of a compound similar to adenosine but no appreciable changes in the nucleotide content of yeast cells, grown either in glucose or galactose. Changes in the specific activities of the enzymes involved in the pathway from ATP to inosine, in yeast extracts from (un)treated cells, could not explain the effect of H2O2 on the levels of ATP and inosine. Application of a mathematical model of differential equations previously developed in this laboratory pointed to a potential inhibition of glycolysis as the main reason for that effect. This theoretical consideration was reinforced both by the lack of an appreciable effect of 1.5 mm (or even higher concentrations) H2O2 on yeast grown in the presence of ethanol or glycerol, and by the observed inhibition of the synthesis of ethanol promoted by H2O2. Normal values for the adenylic charge, ATP and inosine levels were reached at 5, 30 and 120 min, respectively, after removal of H2O2 from the culture medium. The strong decrease in the ATP level upon H2O2 treatment is an important factor to be considered for understanding the response of yeast, and probably other cell types, to oxidative stress.

Adenosine Triphosphate↗

PubMed2003

Response of a strict anaerobe to oxygen: survival strategies in Desulfovibrio gigas.

The biochemical response to oxygen of the strictly anaerobic sulfate-reducing bacterium Desulfovibrio gigas was studied with the goal of elucidating survival strategies in oxic environments. Cultures of D. gigas on medium containing lactate and sulfate were exposed to oxygen (concentration 5-120 micro M). Growth was fully inhibited by oxygen, but the cultures resumed growth as soon as they were shifted back to anoxic conditions. Following 24 h exposure to oxygen the growth rate was as high as 70 % of the growth rates observed before oxygenation. Catalase levels and activity were enhanced by exposure to oxygen whereas superoxide-scavenging and glutathione reductase activities were not affected. The general pattern of cellular proteins as analysed by two-dimensional electrophoresis was altered in the presence of oxygen, the levels of approximately 12 % of the detected proteins being markedly increased. Among the induced proteins, a homologue of a 60 kDa eukaryotic heat-shock protein (Hsp60) was identified by immunoassay analysis. In the absence of external substrates, the steady-state levels of nucleoside triphosphates detected by in vivo (31)P-NMR under saturating concentrations of oxygen were 20 % higher than under anoxic conditions. The higher energy levels developed under oxygen correlated with a lower rate of substrate (glycogen) mobilization, but no experimental evidence for a contribution from oxidative phosphorylation was found. The hypothesis that oxygen interferes with ATP dissipation processes is discussed.

Adenosine Triphosphate↗

PubMed2002

Hydrogen peroxide-induced carbonylation of key metabolic enzymes in Saccharomyces cerevisiae: the involvement of the oxidative stress response regulators Yap1 and Skn7.

H(2)O(2) induces a specific protein oxidation in yeast cells, and the glycolytic enzyme glyceraldehyde-3-phosphate dehydrogenase (Tdh) is a major target. Using a 2D-gel system to study protein carbonylation, it is shown in this work that both Tdh2p and Tdh3p isozymes were oxidized during exposure to H(2)O(2). In addition, we identified two other proteins carbonylated and inactivated: Cu,Zn-superoxide dismutase and phosphoglycerate mutase. The oxidative inactivation of Cu,Zn-superoxide dismutase decreases the antioxidant capacity of yeast cells and probably contributes to H(2)O(2)-induced cell death. Cyclophilin 1 was also carbonylated, but CPH1 gene disruption did not affect peroxide stress sensitivity. The correlation between H(2)O(2) sensitivity and the accumulation of oxidized proteins was evaluated by assaying protein carbonyls in mutants deficient in the stress response regulators Yap1p and Skn7p. The results show that the high sensitivity of yap1delta and skn7delta mutants to H(2)O(2) was correlated with an increased induction of protein carbonylation. In wild-type cells, the acquisition of stress resistance by pre-exposure to a sublethal H(2)O(2) stress was associated with a lower accumulation of oxidized proteins. However, pre-exposure of yap1delta and skn7delta cells to 0.4 mM H(2)O(2) decreased protein carbonylation induced by 1.5 mM H(2)O(2), indicating that the adaptive mechanism involved in the protection of proteins from carbonylation is Yap1p- and Skn7p-independent.

Carbon↗

PubMed2002

Why does SOD overexpression sometimes enhance, sometimes decrease, hydrogen peroxide production? A minimalist explanation.

Toxic effects of superoxide dismutase (SOD) overexpression are commonly attributed to increased hydrogen peroxide (H(2)O(2)) production. Still, published experiments yield contradictory evidence on whether SOD overexpression increases or decreases H(2)O(2) production. We analyzed this issue using a minimal mathematical model. The most relevant mechanisms of superoxide consumption are treated as pseudo first-order processes, and both superoxide production and the activity of enzymes other than SOD were considered constant. Even within this simple framework, SOD overexpression may increase, hold constant, or decrease H(2)O(2) production. At normal SOD levels, the outcome depends on the ratio between the rate of processes that consume superoxide without forming H(2)O(2) and the rate of processes that consume superoxide with high (>/= 1) H(2)O(2) yield. In cells or cellular compartments where this ratio is exceptionally low (< 1), a modest decrease in H(2)O(2) production upon SOD overexpression is expected. Where the ratio is higher than unity, H(2)O(2) production should increase, but at most linearly, with SOD activity. The results are consistent with the available experimental observations. According to the minimal model, only where most superoxide is eliminated through H(2)O(2)-free processes does SOD activity have the moderately large influence on H(2)O(2) production observed in some experiments.

Hydrogen Peroxide↗