No abstract

ABSTRACT. The resistant to glucose repression mutants of Schizosaccharomyces pombe (ird5, ird13, and ird14) have a high tolerance to oxidative stress induced by H 2 O 2 . In all ird mutants, the increased expression level of the fbp1 gene can be interpreted as a lack of glucose repression in these mutants. To investigate the mechanisms of the oxidative stress response in ird mutants, we analyzed the transcription of stress response-related genes, sod1, ctt1, atf1, pap1, and sty1, under stressed and non-stressed conditions. We then analyzed the phosphorylation state of the Sty1-MAP kinase in ird mutants. Our findings support the concept of an adaptive response to oxidative stress in these mutants. In addition, these results imply that either glucose signaling mechanisms leading to glucose repression and glucose utilization as an energy source are regulated apart from each other or, like Saccharomyces cerevisiae, S. pombe might have additional glucose detection systems.

Background/Aims: MiRNAs transpire as promising elements in molecular medicine for the identification of new diagnostic, prognostic and targeting therapeutic biomarkers. This study consisted of four steps: First, to investigate one or a group of specific diagnostic miRNAs for Systemic Lupus Erythematosus (SLE) disease in patients with and without renal involvement, second, to identify cytokines genes’ expression profiling, third, comparing the profiles with related amounts in the serum and finally, to study target-gene-mediated functional roles of miRNAs, which have been correlated to disease development and progression. Methods: In order to use in microarray assays total RNA and miRNAs were isolated from blood and serum samples that were obtained from 16 SLE patients (9 with renal involvement and 7 without renal involvement). Taking coexistence of factors such as hypocomplementemia, positive ANA and anti-DNA into account, obtained data were processed. For each differentially expressed miRNA, potential target genes were predicted by microRNAorg, TargetScan and PITA prediction tools. Obtained mRNA profiling data were interrogated for the target genes. MiRNA and mRNA microarray results were confirmed by QRT-PCR. Finally, the amounts of cytokines were measured by multiplex ELISA method. Results: The results of study showed that among differentially expressed miRNAs in SLE patients with renal involvement compared to those without renal involvement, hsa-miR-766-3p, may play pivotal roles in PI3K-AKT-mTOR pathway. In addition according to the obtained data it is suggested that blood-borne proinflammatory cytokines such as IL-4, IL-6 and TNF-α alongside with disease stage and severity may contribute to this differential expression of these miRNA which may be leading to insulin resistance. Finally, hsa-miR-621, which was differentially expressed in hypertensive SLE patients without renal involvement and a positive ANA test with its predicted target gene “Kallikrein-related peptidase 9” may play a role in the pathophysiology of hypertension in SLE. Conclusions: We reported some human miRNAs which were differentially expressed in SLE patients according to disease activity and renal involvement. Larger studies are necessary to confirm our findings and detect further biomarkers.

ABSTRACT.The relationship between glucose repression and the oxidative stress response was investigated in Schizosaccharomyces pombe wild type cells (972h -) and glucose repression resistant mutant type cells (ird11). We aimed to reveal the mechanism of simultaneous resistance to glucose repression and oxidative stress in ird11 mutants. Compared to the wild type, the expression of the sty1 gene was not altered in the ird11 mutant under normal growth conditions, but decreased after exposure to H 2 O 2 . This effect was clearly explained by the immunoblotting results, which showed elevated levels of a much more stable phosphorylated form of Sty1 mitogen-activated protein kinase in the ird11 mutant. Increased ght3 gene expression levels were also found, which may play a role in protecting the ird11 mutant from the deleterious effects of oxidative stress. In addition, decreased expression levels of glycolytic enzyme enolase-and thiamine synthesis/ transport-related genes were detected. This might have resulted from the flux redirection toward mitochondrial respiration, which would enhance NADPH generation to prevent the high reactive oxygen species accumulation that is generated by respiration. Some evidence supported a flux shift toward fermentation as well as respiration. We conclude that a defect in the glucose-sensing signaling pathway in Effects of thiamine in oxidative stress ird11 mutants likely causes erroneous low glucose-sensing signaling and high ATP production. This most likely occurs because high glucose availability in the medium induces an impairment in the respiratory chain and fermentation balance in these cells, which might explain the glucose repression and oxidative stress resistance in ird11 compared to the wild type.

Previously we reported that the glucose sensing/signaling pathway affected the oxidative stress response of Schizosaccharomyces pombe, based on studies of glucose-repression and oxidative-stress-resistant mutants including ird5. sty1, encoding a protein kinase, and aft1 and pap1, encoding transcription factors, are important components of the oxidative stress response in S. pombe. To analyze the relationship between the glucose sensing/signaling pathway and the oxidative stress response, we generated sty1, aft1, and pap1 knockout mutants in ird5 and wild-type backgrounds. We evaluated the survival rates of the ird5 double mutants (5A, 5P, and 5S) and wild-type single mutants (9A, 9P, and 9S) under mild oxidative stress. In addition, we analyzed the transcript levels of genes related to oxidative stress (sod1, encoding superoxide dismutase; ctt1, encoding catalase) and glucose metabolism (fbp1, encoding fructose-1, 6-bisphosphatase; hxk2, encoding hexokinase). All deletion mutants showed very low survival rates under oxidative stress (<7%). It was shown that transcript levels of sod1 drastically increased in single mutants, while those of ctt1 slightly increased in double mutants. Our results indicate that these mutants are highly sensitive to oxidative stress, and that atf1, pap1, and sty1, as well as sod1 and ctt1, are very important in the oxidative stress response of S. pombe.