PubMed Health⌕ Search

Biomedical subjects

S Izawa

Publications and source records attributed to S Izawa.

At least 19 recordsLinked to original sources
PubMed2006

2'-,5'-Oligoadenylate synthetase response ratio predicting virological response to PEG-interferon-alpha2b plus ribavirin therapy in patients with chronic hepatitis C.

OBJECTIVE: Although all the mechanisms of elimination of hepatitis C virus (HCV) by Interferon (IFN) have not been fully elucidated, the 2'-5'-oligoadenylate (2-5A) system is one of the mechanisms of the antiviral effect of IFN. Consequently, the measurement of 2'-5'-oligoadenylate synthetase (2-5AS) activity could be useful for the evaluation of IFN treatment. This retrospective study was aimed at assessing whether 2-5AS activity functions as a clinical marker of virological response to PEG-interferon-alpha2b (PEG-IFN) plus ribavirin therapy of chronic hepatitis C. METHODS: The 32 patients included in this study had high viral loads of serum HCV-RNA of genotype 1b with chronic hepatitis C. All the patients received a regimen of PEG-IFN plus ribavirin for 48 weeks, and were then divided into two groups: one group (effective group) with undetectable serum HCV-RNA levels at 24 weeks (n = 22) of therapy, the other group (ineffective group) with persistent presence of HCV-RNA in serum at 24 weeks (n = 10). The 2-5AS activity in serum was measured 2, 8 and 12 weeks before initial administration. RESULTS: The 2-5AS response ratio (measured value/measured value of baseline 2-5AS) at 2, 8 and 12 weeks after the administration in the effective group was significantly higher than that in the ineffective group. CONCLUSIONS: These results suggest that the ratio of 2-5AS is closely related to the antiviral effect, and that the measurement of 2-5AS response ratio may be a useful clinical parameter of virological response to PEG-IFN plus ribavirin therapy of chronic hepatitis C.

2',5'-Oligoadenylate Synthetase↗
PubMed2005

Interferon-beta induction/interferon-alpha2b plus ribavirin therapy in patients with chronic hepatitis C.

Treatment of chronic hepatitis C virus (HCV) infection with interferon (IFN) and ribavirin improves the rate of eradication of the virus by less than 20% in patients with genotype 1b and a high viral load. In this study we assessed whether IFN-beta induction/IFN-alpha2b plus ribavirin enhances the efficacy of the therapy in patients with chronic hepatitis C. The efficacy of IFN-beta induction/IFN-alpha2b plus ribavirin therapy (group A, n=7) was compared with that of IFN-alpha2b plus ribavirin (group B, n=7) in 14 patients with high levels of HCV-RNA (> 100 K/U/ml). No significant differences were observed in the clearance of HCV-RNA between the two groups (A and B, respectively) 2 weeks after the start of the treatment (0% and 14.3%), at the end of the treatment (71.4% and 100%) and 6 months after the end of the treatment (28.6% and 14.3%). Recovery was complete in 28.6% and 14.3%, transient in 42.9% and 85.7% and absent in 28.6% and 0% in groups A and B, respectively. Early log changes in the viral load from the baseline after 2 weeks of treatment were 2.41 +/- 0.91 and 2.77 +/- 0.20 in groups A and B, respectively, with no significant difference between the two groups. In the present study, we were not able to demonstrate that IFN-beta induction/IFN-alpha2b plus ribavirin therapy was superior to IFN-alpha2b plus ribavirin therapy in patients with genotype 1b and high viral loads.

Adult↗
PubMed2003

A screening system for antioxidants using thioredoxin-deficient yeast: discovery of thermostable antioxidant activity from Agaricus blazei Murill.

Previously, we found that cytosolic thioredoxin is a negative regulator for an oxidative stress responsive transcription factor, Yap1p (yeast AP-1-like transcription factor), i.e., this transcription factor is constitutively concentrated in the nucleus in the thioredoxin-deficient mutant ( trx1delta trx2delta) due to an impairment of the reactive oxygen species-scavenging activity of this mutant [Izawa et al. (1999) J Biol Chem 274:28459-28465]. Based on these findings, we developed a screening method to discover substances that show antioxidant activity. With this method, antioxidant activity was evaluated by monitoring the subcellular localization of Yap1p. Since Yap1p is oxidized and accumulates in the nucleus in trx1delta trx2Delta cells, it is easy to identify antioxidant activity by observing the localization of green fluorescent protein (GFP)-tagged Yap1p. If exogenous substances taken in by trx1delta trx2Delta cells were able to function as antioxidants to reduce the oxidized form of Yap1p, GFP1-Yap1p would diffuse into the cytoplasm. We used this system to screen for antioxidant activity in mushrooms, and found that the edible mushroom Agaricus blazei Murill is an excellent source of antioxidants.

Agaricus↗
PubMed2003

Nuclear thioredoxin peroxidase Dot5 in Saccharomyces cerevisiae: roles in oxidative stress response and disruption of telomeric silencing.

The DOT5 gene was originally cloned as one of the DOT (disrupter of telomeric silencing) genes; and later it was re-discovered as a nuclear thioredoxin peroxidase in Saccharomyces cerevisiae. Here, we demonstrate that the telomeric-silencing disruption activity of Dot5 is independent of thioredoxin peroxidase activity. In addition, Dot5 cannot suppress the increased susceptibility to peroxides of mutants defected in cytosolic thioredoxin peroxidase, even when Dot5 is expressed in the cytoplasm. Furthermore, Dot5 does not affect redox regulation of the Yap1 transcription factor. These results suggest that Dot5 is less important as an antioxidant in yeast cells.

Cell Nucleus↗
PubMed2001

The Zrc1 is involved in zinc transport system between vacuole and cytosol in Saccharomyces cerevisiae.

The ZRC1 gene encodes a multicopy suppressor of zinc toxicity in Saccharomyces cerevisiae; however, previously we found that the expression of ZRC1 was induced when the intracellular zinc level was decreased. Zrc1 has six putative transmembrane domains and we determined that a Zrc1-GFP fusion protein was localized to the vacuolar membrane. The steady state level of intracellular zinc in a zrc1Delta mutant cultured in the zinc-abundant medium was lower than that in wild type. No distinct difference was observed in the basal activity of glyoxalase I, which is a cytosolic enzyme requiring zinc for catalytic function and is used here as a marker for cytosolic zinc-availability, between wild type and zrc1Delta mutant, although the activity was decreased much greater extent in the zrc1Delta mutant if the cells were exposed to the metal-limited medium. Similarly, the basal expression level of ZRC1-lacZ reporter gene in zrc1Delta mutant was the same as that in wild type; however, the fold of induction of ZRC1-lacZ expression in zrc1Delta mutant under the zinc-limited conditions was higher than that in the wild type. Based on these results, we present a tentative model for the function of Zrc1 as a mechanism to maintain the zinc homeostasis in yeast.

Base Sequence↗
PubMed2001

Regulation of the yeast Yap1p nuclear export signal is mediated by redox signal-induced reversible disulfide bond formation.

Yap1p, a crucial transcription factor in the oxidative stress response of Saccharomyces cerevisiae, is transported in and out of the nucleus under nonstress conditions. The nuclear export step is specifically inhibited by H(2)O(2) or the thiol oxidant diamide, resulting in Yap1p nuclear accumulation and induction of transcription of its target genes. Here we provide evidence for sensing of H(2)O(2) and diamide mediated by disulfide bond formation in the C-terminal cysteine-rich region (c-CRD), which contains 3 conserved cysteines and the nuclear export signal (NES). The H(2)O(2) or diamide-induced oxidation of the c-CRD in vivo correlates with induced Yap1p nuclear localization. Both were initiated within 1 min of application of oxidative stress, before the intracellular redox status of thioredoxin and glutathione was affected. The cysteine residues in the middle region of Yap1p (n-CRD) are required for prolonged nuclear localization of Yap1p in response to H(2)O(2) and are thus also required for maximum transcriptional activity. Using mass spectrometry analysis, the H(2)O(2)-induced oxidation of the c-CRD in vitro was detected as an intramolecular disulfide linkage between the first (Cys(598)) and second (Cys(620)) cysteine residues; this linkage could be reduced by thioredoxin. In contrast, diamide induced each pair of disulfide linkage in the c-CRD, but in this case the cysteine residues in the n-CRD appeared to be dispensable for the response. Our data provide evidence for molecular mechanisms of redox signal sensing through the thiol-disulfide redox cycle coupled with the thioredoxin system in the Yap1p NES.

Cell Nucleus↗
PubMed2000

Role of glutathione in heat-shock-induced cell death of Saccharomyces cerevisiae.

Previously we reported that expression of GSH1 (gamma-glutamylcysteine synthetase) and GSH2 (glutathione synthetase) of the yeast Saccharomyces cerevisiae was increased by heat-shock stress in a Yap1p-dependent fashion and consequently intracellular glutathione content was increased [Sugiyama, Izawa and Inoue (2000) J. Biol. Chem. 275, 15535-15540]. In the present study, we discuss the physiological role of glutathione in the heat-shock stress response in this yeast. Both gsh1 and gsh2 mutants could acquire thermotolerance by mild heat-shock stress and induction of Hsp104p in both mutants was normal; however, mutant cells died faster by heat shock than their parental wild-type strain. After pretreatment at a sublethal temperature, the number of respiration-deficient mutants increased in a gsh1 mutant strain in the early stages of exposure to a lethal temperature, although this increase was partially suppressed by the addition of glutathione. These results lead us to suspect that an increase of glutathione synthesis during heat-shock stress is to protect mitochondrial DNA from oxidative damage. To investigate the correlation between mitochondrial DNA damage and glutathione, mitochondrial Mn-superoxide dismutase (the SOD2 gene product) was disrupted. As a result, the rate of generation of respiration-deficient mutants of a sod2 delta strain was higher than that of the isogenic wild-type strain and treatment of the sod2 delta mutant with buthionine sulphoximine, an inhibitor of glutathione synthesis, inhibited cell growth. These results suggest that glutathione synthesis is induced by heat shock to protect the mitochondrial DNA from oxidative damage that may lead to cell death.

Blotting, Western↗
PubMed2000

Expression of ZRC1 coding for suppressor of zinc toxicity is induced by zinc-starvation stress in Zap1-dependent fashion in Saccharomyces cerevisiae.

The ZRC1 gene was cloned as a multicopy suppressor of zinc toxicity in Saccharomyces cerevisiae. Zrc1 belongs to CDF (cation diffusion facilitator) family. The transporters belonging to this family are thought to play an important role in metal detoxification. However, we found that cell growth of zrc1Delta mutant was lowered under the metal-limited conditions, which was restored by zinc. The Zap1 transcription factor is crucial for expression of several genes responsive to zinc-starvation stress. The expression of ZRC1 was induced in Zap1-dependent fashion when the intracellular zinc level was decreased and this induction was repressed by zinc. These results imply an important role of Zrc1 in the zinc-starvation stress.

Base Sequence↗
PubMed2000

Activity of neurons in the nucleus of the solitary tract of rats: effect of osmotic and mechanical stimuli.

Patch-clamp recordings were used to examine the osmosensitivity and mechanosensitivity of neurons in the caudal part of the nucleus tractus solitarius in coronal slices from rat brain. Firing rates and membrane potentials were measured as slices were exposed to perfusate which varied in its osmolality and/or sodium concentration. In all cells tested, the responses to change in the sodium concentration of perfusate were duplicated by osmolality changes of sucrose or mannitol. When nucleus tractus solitarius cells were tested with changes in pressure applied via the pipette, responses to positive or negative pressure paralleled their responses to osmotic stimulation. We suggest that a mechanosensitive receptor exists on osmosensitive neurons within the nucleus tractus solitarius, and this receptor may be responsible for changes in the firing rate and membrane potential which occur in the nucleus tractus solitarius neurons.

Action Potentials↗
PubMed2000

The Yap1p-dependent induction of glutathione synthesis in heat shock response of Saccharomyces cerevisiae.

Glutathione is synthesized in two sequential reactions catalyzed by gamma-glutamylcysteine synthetase (GSH1 gene product) and glutathione synthetase (GSH2 gene product). The expression of GSH1 in Saccharomyces cerevisiae has been known to be up-regulated by Yap1p, a critical transcription factor for the oxidative stress response in yeast. The present study demonstrates that GSH2 expression is also regulated by Yap1p under oxidative stress-induced conditions. In addition to oxidative stress, expression of GSH1 and GSH2 was induced by heat shock stress in a Yap1p-dependent manner with subsequent increases in intracellular glutathione content. Oxygen respiration rate increased when cells were exposed to higher temperatures, and as a result, intracellular oxidation levels were increased. The heat shock-induced expression of GSH1 and GSH2 did not occur under anaerobic conditions. Furthermore, even under aerobic conditions, the heat shock response of these genes was not observed when cells were pretreated with KCN to block oxygen respiration. We speculate that heat shock stress enhances oxygen respiration, which in turn results in an increase in the generation of reactive oxygen species in mitochondria. This signal may be mediated by Yap1p, resulting in the elevation of intracellular glutathione levels.

Base Sequence↗
PubMed2000

Cooperative regulation of DOG2, encoding 2-deoxyglucose-6-phosphate phosphatase, by Snf1 kinase and the high-osmolarity glycerol-mitogen-activated protein kinase cascade in stress responses of Saccharomyces cerevisiae.

We screened the genome of Saccharomyces cerevisiae for the genes responsive to oxidative stress by using the lacZ transposon-insertion library. As a result, we found that expression of the DOG2 gene coding for 2-deoxyglucose-6-phosphate phosphatase was induced by oxidative stress. The expression of DOG2 was also induced by osmotic stress. We found a putative cis element (STRE, a stress response element) in the DOG2 promoter adjacent to a consensus sequence to which the Mig1p repressor is known to bind. The basal levels of DOG2 gene expression were increased in a mig1Delta mutant, while the derepression of DOG2 was not observed in a snf1Delta mutant under glucose-deprived conditions. Induction of the DOG2 gene expression by osmotic stress was observed in any of the three disruptants pbs2Delta, hog1Delta, and snf1Delta. However, the osmotic induction was completely abolished in both the snf1Delta pbs2Delta mutant and the snf1Delta hog1Delta mutant. Additionally, these single mutants as well as double mutants failed to induce DOG2 expression by oxidative stress. These results suggest that Snf1p kinase and the high-osmolarity glycerol-mitogen-activated protein kinase cascade are likely to be involved in the signaling pathway of oxidative stress and osmotic stress in regulation of DOG2.

Base Sequence↗
PubMed1999

Thioredoxin deficiency causes the constitutive activation of Yap1, an AP-1-like transcription factor in Saccharomyces cerevisiae.

Yap1 is a transcription factor that responds to oxidative stress in Saccharomyces cerevisiae. The activity of Yap1 is regulated at the level of its intracellular localization, and a cysteine-rich domain at the C terminus of Yap1 is involved in this regulation. We investigated the effects of redox-regulatory proteins, thioredoxin and glutaredoxin, on the regulation of Yap1, using the deficient mutants of these thiol-disulfide oxidoreductases. In the thioredoxin-deficient mutant (trx1Delta/trx2Delta), Yap1 was constitutively concentrated in the nucleus and the level of expression of the Yap1 target genes was high under normal conditions, while this was not the case for the glutaredoxin-deficient mutant (grx1Delta/grx2Delta). No distinct difference was observed in the levels of Yap1 protein between the wild type and trx1Delta/trx2Delta. The constitutive activation of Yap1 in trxDelta/trx2Delta was observed under aerobic conditions but not under anaerobic conditions. These findings suggest that thioredoxin has negative effects on this regulation via the redox states. We also show the synthetic lethality between yap1Delta and trx1Delta/trx2Delta mutation, but the yap1Delta/grx1Delta/grx2Delta triple mutant was viable, suggesting a difference of the functions between thioredoxin and glutaredoxin and a genetic interaction between Yap1 and thioredoxin in vivo.

Base Sequence↗
PubMed1999

Genetic analysis of glutathione peroxidase in oxidative stress response of Saccharomyces cerevisiae.

Three glutathione peroxidase homologs (YKL026C, YBR244W, and YIR037W/HYR1) were found in the Saccharomyces Genome Database. We named them GPX1, GPX2, and GPX3, respectively, and we investigated the function of each gene product. The gpx3Delta mutant was hypersensitive to peroxides, whereas null mutants of the GPX1 and GPX2 did not show any obvious phenotypes. Glutathione peroxidase activity decreased approximately 57 and 93% in the gpx3Delta and gpx1Delta/gpx2Delta/gpx3Delta mutants, respectively, compared with that of wild type. Expression of the GPX3 gene was not induced by any stresses tested, whereas that of the GPX1 gene was induced by glucose starvation. The GPX2 gene expression was induced by oxidative stress, which was dependent upon the Yap1p. The TSA1 (thiol-specific antioxidant) gene encodes thioredoxin peroxidase that can reduce peroxides by using thioredoxin as a reducing power. Disruption of the TSA1 gene enhanced the basal expression level of the Yap1p target genes such as GSH1, GLR1, and GPX2 and that resulted in increases of total glutathione level and activities of glutathione reductase and glutathione peroxidase. However, expression of the TSA1 gene did not increase in the gpx1Delta/gpx2Delta/gpx3Delta mutant. Therefore, de novo synthesis and recycling of glutathione were increased in the tsa1Delta mutant to maintain the catalytic cycle of glutathione peroxidase reaction efficiently as a backup system for thioredoxin peroxidase.

Amino Acid Sequence↗
PubMed1998

Importance of glucose-6-phosphate dehydrogenase in the adaptive response to hydrogen peroxide in Saccharomyces cerevisiae.

Glucose-6-phosphate dehydrogenase (G6PDH)-deficient cells of Saccharomyces cerevisiae showed increased susceptibility and were unable to induce adaptation to oxidative stress. Historically, mainly in human erythrocytes, it has been suggested and accepted that decreased cellular GSH, due to loss of the NADPH-dependent activity of glutathione reductase (GR), is responsible for the increased sensitivity to oxidative stress in G6PDH-deficient cells. In the present study we investigated whether the increased susceptibility and the inability to induce adaptation to H2O2 stress of G6PDH-deficient yeast is caused by incompleteness of glutathione recycling. We constructed G6PDH- and GR-deficient mutants and analysed their adaptive response to H2O2 stress. Although G6PDH-deficient cells contained comparable amounts of GSH and GR activity to wild-type cells, GSSG was not reduced efficiently, and intracellular GSSG levels and the ratio of GSSG to total glutathione (GSSG/tGSH) were higher in G6PDH-deficient cells than in wild-type. On the other hand, GR-deficient cells showed a susceptibility identical with that of wild-type cells and induced adaptation to H2O2 stress, even though the GSSG/tGSH ratio in GR-deficient cells was higher than in G6PDH-deficient cells. These results indicate that incompleteness of glutathione recycling alone is not sufficient to account for the increased sensitivity and inability to induce adaptation to H2O2 stress of G6PDH-deficient yeast cells. In S. cerevisiae, G6PDH appears to play other important roles in the adaptive response to H2O2 stress besides supplying NADPH to the GR reaction.

Adaptation, Physiological↗
PubMed1998

Molecular identification of glutathione synthetase (GSH2) gene from Saccharomyces cerevisiae.

The hypothetical protein YOL049w on the chromosome XV was identified to be the structural gene for glutathione synthetase (GSH2) of Saccharomyces cerevisiae. Translational initiation site was identified by making the GSH2-lacZ fusion. The GSH2 gene contained an open reading frame (1473 bp) with 491 amino acids, and molecular weight of the GSH2 gene product was calculated to be 55,812. Glutathione synthetase activity in transformant carrying the GSH2 gene with multicopy plasmid increased approximately 4-fold. The GSH2 gene was not essential for growth of yeast cell, and glutathione was not detected from the gsh2 disrupter.

Amino Acid Sequence↗
PubMed1996

Importance of catalase in the adaptive response to hydrogen peroxide: analysis of acatalasaemic Saccharomyces cerevisiae.

Controversy about the importance of catalase in the detoxification of H2O2 in human erythrocytes continues. It has been suggested that catalase has no role in the clearance of H2O2 in erythrocytes. In the present study we investigated the role of catalase in the defence mechanism against oxidative stress using Saccharomyces cerevisiae. S. cerevisiae has two catalases, catalase A and catalase T. We constructed a double mutant (acatalasaemic mutant) unable to produce either catalase A or catalase T, and compared it with wild-type and single-mutant cells. The acatalasaemic mutant cells showed a similar growth rate to wild-type cells under non-oxidative stress conditions, and showed a similar susceptibility to H2O2 stress in the exponential growth phase. The acatalasaemic mutant cells at stationary phase were, however, much more sensitive to H2O2 stress than wild-type and single-mutant cells. Moreover, the ability of acatalasaemic and single-mutant cells to show adaptation to 2 mM H2O2 was distinctly inferior to that of wild-type cells. These results suggest that catalase is not essential for yeast cells under normal conditions, but plays an important role in the acquisition of tolerance to oxidative stress in the adaptive response of these cells.

Adaptation, Physiological↗
PubMed1996

Modification of Escherichia coli B glutathione synthetase with polyethylene glycol for clinical application to enzyme replacement therapy for glutathione deficiency.

Glutathione synthetase of Escherichia coli B was modified with polyethylene glycol, and the properties of the resultant modified enzyme were investigated. The thermal stability of the modified enzyme and its resistance against several proteases increased compared with those of the native enzyme. The modified enzyme was injected intravenously via the rat tail vein, and the circulating life of the enzyme in plasma was monitored. The half-life of the native enzyme was 50 min, whereas that of the modified enzyme was approximately 24 h. The systemic anaphylaxis reaction was tested by using rats intravenously injected with the native and modified enzymes. For the native enzyme, strong reactions such as dyspnea and tumble were observed; however, no symptom or only a very weak reaction, such as scratching, was observed with the modified enzyme.

Anaphylaxis↗