Project description:Cbf1 is a basic helix-loop-helix transcription factor which regulates the expression of many genes of the sulphur assimilation pathway in yeast. Gamma-glutamylcysteine synthetase (GSH1), encoding the rate-limiting enzyme in glutathione biosynthesis, is constitutively elevated in cbf1 mutants indicating that Cbf1 normally represses GSH1 expression. We used transcriptional profiling to show that a number of antioxidant and stress genes are similarly repressed by Cbf1, consistent with the high oxidant tolerance of cbf1 mutants. Our data indicate that Cbf1 plays a key role in the regulation of gene expression during oxidative stress conditions induced by exposure to hydrogen peroxide. The Yap1 transcription factor induces GSH1 expression in response to hydrogen peroxide in a mechanism that does not require the Met4 transcription factor to overcome the inhibitory action of Cbf1. We show that hydrogen peroxide does not affect Cbf1 occupancy on the GSH1 promoter, but results in specific modification of Cbf1 by phosphorylation. Our data indicate that casein kinase (CK2) can directly phosphorylate Cbf1 in vitro. Furthermore, CK2 activity is required to phosphorylate Cbf1 and induce GSH1 expression in yeast cells in response to hydrogen peroxide. CK2 mutants are sensitive to hydrogen peroxide consistent with a role for CK2 in regulating the cellular response to oxidative stress. CK2 is a ubiquitous serine/threonine protein kinase and the finding that it is regulated by oxidative stress is particularly interesting, since there is increasing evidence that it is involved in a number of pathological conditions including cancer, neurodegenerative and cardiovascular diseases.
Project description:To gain insight into the basic mechanism of Hydrogen peroxide detoxification in the methylotrophic yeast, H. polymorpha, we analyzed changes in transcriptional profiles in response to hydrogen peroxide exposure.
Project description:To gain insight into the basic mechanism of Hydrogen peroxide detoxification in the methylotrophic yeast, H. polymorpha, we analyzed changes in transcriptional profiles in response to hydrogen peroxide exposure. Total RNA samples were collected from H. polymorpha cells after 30 min incubation with 0.5mM hydrogen peroxide. Using the RNA sample obtained prior to hydrogen peroxide addition as a reference, the differential fluorescence intensities of each RNA sample prepared at the indicated time was measured after labeling with Cy3 or Cy5 fluorochromes. For all analyses, we performed dye swapping experiments to avoid dye bias.
Project description:Aims: Vascular wall resident stem cells (SCs) hold great promise for cardiovascular regenerative therapy. This study evaluated the impact of oxidative stress on the viability and functionality of saphenous vein'derived pericytes (SVPs), SVPs'deriv ed cells and terminally differentiated vascular cells. We also investigated the molecular mechanisms underlying the SVP resistance to oxidative stress. Results: SVPs exhibit a significantly higher resistance to oxidative stress resulting in reduced apoptosis upon exposure to hydrogen peroxide in comparison to endothelial cells (ECs). This was attributed to upregulation of genes encoding for anti'oxidant enzymes, especially superoxide dismutases (SODs) and catalase. Pharmacological inhibition of SODs increased ROS levels in SVPs and impaired their survival. Furthermore, we confirm that, when exposed to differentiation stimuli, SVPs are able to generate mesodermic lineages. Interestingly, differentiation of SVPs resulted in SOD down'regulation and increased apoptosis upon exposure to hydrogen peroxide. Oxidative stress caused an incremental increase on SVPs migration, whilst being inhibitory for the ECs. Additionally, oxidative stress did not impair SVPs capacity to promote angiogenesis in vitro. Innovations: This study for the first time demonstrates that SCs resident in veins of cardiovascular patients are endowed with enhanced detoxifier and antioxidant system. Conclusions: SVPs expanded from vein remnants of coronary artery bypass graft surgery express antioxidant defense mechanisms which allow them resist to high levels of ROS. These properties highlight the potential of SVPs in cardiovascular regenerative medicine. 2 groups 3 replicates
Project description:Transcriptional profiling of Escherichia coli K-12 comparing luxS mutant LW12 with wild type W3110 exposure to 10mM or 30mM hydrogen peroxide. Two-condition experiment, luxS mutant LW12 vs. wild type W3110, treatment with 10mM hydrogen peroxide for 30min or treatment with 30mM hydrogen peroxide for 30min. Two biological replicates.
Project description:We report 293 Neisseria gonorrhoeae genes that show differential transcript abundance in response to 15 mM hydrogen peroxide treatment by RNA-Seq. We analyze the major physiological functional groups of genes affected by hydrogen peroxide exposure. In addition, we analyze which genes in our hydrogen peroxide-responsive set of genes belong to major known transcriptional regulatory circuits like iron homeostasis, anaerobiosis and others. We annotate which of the 293 hydrogen peroxide-responsive genes belong to operons. We annotate global transcriptional start sites and identify transcriptional start sites that are only present in hydrogen peroxide-treated bacteria. We validate the RNA-Seq data for a subset of representative genes by RT-qPCR and whether transcript abundance in this same subset of genes differs upon treatement with other reactive oxygen species encountered during infection, like organic peroxide, super oxide anion, and bleach.
Project description:MnSOD is an essential primary antioxidant enzyme that converts superoxide radicals and protons to hydrogen peroxide (H2O2) within the mitochondrial matrix, generated by respiratory chain activity We used microarrays of cells knocked down for MnSOD and a mock transfected cells as their control (siScramble) to reveal changes in gene expression profile
Project description:Aims: Vascular wall resident stem cells (SCs) hold great promise for cardiovascular regenerative therapy. This study evaluated the impact of oxidative stress on the viability and functionality of saphenous vein'derived pericytes (SVPs), SVPs'deriv ed cells and terminally differentiated vascular cells. We also investigated the molecular mechanisms underlying the SVP resistance to oxidative stress. Results: SVPs exhibit a significantly higher resistance to oxidative stress resulting in reduced apoptosis upon exposure to hydrogen peroxide in comparison to endothelial cells (ECs). This was attributed to upregulation of genes encoding for anti'oxidant enzymes, especially superoxide dismutases (SODs) and catalase. Pharmacological inhibition of SODs increased ROS levels in SVPs and impaired their survival. Furthermore, we confirm that, when exposed to differentiation stimuli, SVPs are able to generate mesodermic lineages. Interestingly, differentiation of SVPs resulted in SOD down'regulation and increased apoptosis upon exposure to hydrogen peroxide. Oxidative stress caused an incremental increase on SVPs migration, whilst being inhibitory for the ECs. Additionally, oxidative stress did not impair SVPs capacity to promote angiogenesis in vitro. Innovations: This study for the first time demonstrates that SCs resident in veins of cardiovascular patients are endowed with enhanced detoxifier and antioxidant system. Conclusions: SVPs expanded from vein remnants of coronary artery bypass graft surgery express antioxidant defense mechanisms which allow them resist to high levels of ROS. These properties highlight the potential of SVPs in cardiovascular regenerative medicine.
Project description:Acinetobacter baumannii is a Gram-negative opportunistic pathogen that causes multiple infections, including pneumonia, bacteremia, and wound infections. Due to multiple intrinsic and acquired drug-resistance mechanisms, A. baumannii isolates are commonly multi-drug resistant and infections are notoriously difficult to treat. Therefore, it is important to identify mechanisms used by A. baumannii to survive stresses encountered during infection as a means of identifying new drug targets. In this study, we determined the transcriptional response of A. baumannii to hydrogen peroxide stress using RNASequencing. Upon exposure to hydrogen peroxide, A. baumannii differentially transcribes several hundred genes. In this study, we also determined the transcriptional profile of A. baumannii strains with the transcriptional regulators mumR or oxyR genetically inactivated and identified transcriptional differences between these strains and wild-type A. baumannii in response to hydrogen peroxide stress. In doing this, the function of A. baumannii OxyR in hydrogen peroxide stress resistance and regulation of genes required for hydrogen peroxide detoxification was defined. Moreover, the contribution of the uncharacterized regulator MumR to hydrogen peroxide stress resistance was also explored. This work reveals the transcriptome of an important human pathogen in the presence of hydrogen peroxide stress.