Redox-mediated interactions of VHb (Vitreoscilla haemoglobin) with OxyR: novel regulation of VHb biosynthesis under oxidative stress

2010 ◽  
Vol 426 (3) ◽  
pp. 271-280 ◽  
Author(s):  
Arvind Anand ◽  
Brian T. Duk ◽  
Sandeep Singh ◽  
Meltem Y. Akbas ◽  
Dale A. Webster ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Transcriptional Control ◽  
High Sensitivity ◽  
Reduced Form ◽  
Gene Encoding ◽  
Antioxidant Genes ◽  
E Coli ◽  
Hypoxic Conditions ◽  
Catalase Peroxidase

The bacterial haemoglobin from Vitreoscilla, VHb, displays several unusual properties that are unique among the globin family. When the gene encoding VHb, vgb, is expressed from its natural promoter in either Vitreoscilla or Escherichia coli, the level of VHb increases more than 50-fold under hypoxic conditions and decreases significantly during oxidative stress, suggesting similar functioning of the vgb promoter in both organisms. In the present study we show that expression of VHb in E. coli induced the antioxidant genes katG (catalase–peroxidase G) and sodA (superoxide dismutase A) and conferred significant protection from oxidative stress. In contrast, when vgb was expressed in an oxyR mutant of E. coli, VHb levels increased and the strain showed high sensitivity to oxidative stress without induction of antioxidant genes; this indicates the involvement of the oxidative stress regulator OxyR in mediating the protective effect of VHb under oxidative stress. A putative OxyR-binding site was identified within the vgb promoter and a gel-shift assay confirmed its interaction with oxidized OxyR, an interaction which was disrupted by the reduced form of the transcriptional activator Fnr (fumurate and nitrate reductase). This suggested that the redox state of OxyR and Fnr modulates their interaction with the vgb promoter. VHb associated with reduced OxyR in two-hybrid screen experiments and in vitro, converting it into an oxidized state in the presence of NADH, a condition where VHb is known to generate H2O2. These observations unveil a novel mechanism by which VHb may transmit signals to OxyR to autoregulate its own biosynthesis, simultaneously activating oxidative stress functions. The activation of OxyR via VHb, reported in the present paper for the first time, suggests the involvement of VHb in transcriptional control of many other genes as well.

Hydrogen peroxide-induced response in E. coli and S. cerevisiae: different stages of the flow of the genetic information

2009 ◽  
Vol 4 (2) ◽  
pp. 142-153 ◽  
Author(s):  
Halyna Semchyshyn
Keyword(s):  
Oxidative Stress ◽  
Hydrogen Peroxide ◽  
Genetic Information ◽  
Transcriptional Control ◽  
Induced Response ◽  
Global Response ◽  
Antioxidant Genes ◽  
E Coli ◽  
Basic And Applied Research ◽  
Different Levels

AbstractAdaptation to oxidative stress is a major topic in basic and applied research. Cell response to stressful changes is realized through coordinated reorganization of gene expression. E. coli and S. cerevisiae are extremely amenable to genetic or molecular biological and biochemical approaches, which make these microorganisms suitable models to study stress response at a molecular level in prokaryotes and eukaryotes, respectively. The main focus of this review is (i) to discuss transcriptional control of global response to hydrogen peroxide in E. coli and S. cerevisiae, (ii) to summarize recent literature data on E. coli and S. cerevisiae adaptive response to oxidative stress at different stages of the flow of the genetic information: from transcription and translation to functionally active proteins and (iii) to discuss possible reasons for a lack of correlation between the expression of certain antioxidant genes at different levels of cellular organization.

scholarly journals Phosphorylation of Nucleosides by the Mutated Acid Phosphatase from Morganella morganii

2000 ◽  
Vol 66 (7) ◽  
pp. 2811-2816 ◽  
Author(s):  
Yasuhiro Mihara ◽  
Takashi Utagawa ◽  
Hideaki Yamada ◽  
Yasuhisa Asano
Keyword(s):  
Acid Phosphatase ◽  
Random Mutagenesis ◽  
Reaction Yield ◽  
Mutant Library ◽  
Morganella Morganii ◽  
Gene Encoding ◽  
E Coli ◽  
Acid Phosphatase Gene ◽  
Phosphate Source

ABSTRACT A novel nucleoside phosphorylation process using the food additive pyrophosphate as the phosphate source was investigated. TheMorganella morganii gene encoding a selective nucleoside pyrophosphate phosphotransferase was cloned. It was identical to theM. morganii PhoC acid phosphatase gene. Sequential in vitro random mutagenesis was performed on the gene by error-prone PCR to construct a mutant library. The mutant library was introduced intoEscherichia coli, and the transformants were screened for the production of 5′-IMP. One mutated acid phosphatase with an increased phosphotransferase reaction yield was obtained. With E. coli overproducing the mutated acid phosphatase, 101 g of 5′-IMP per liter (192 mM) was synthesized from inosine in an 88% molar yield. This improvement was achieved with two mutations, Gly to Asp at position 92 and Ile to Thr at position 171. A decreasedKm value for inosine was responsible for the increased productivity.

scholarly journals WNK4 Kinase: from structure to physiology

2021 ◽  
Author(s):  
Adrian Rafael Murillo-de-Ozores ◽  
Alejandro Rodriguez-Gama ◽  
Hector Carbajal-Contreras ◽  
Gerardo Gamba ◽  
Maria Castaneda-Bueno
Keyword(s):  
Oxidative Stress ◽  
Mouse Models ◽  
Serine Threonine Kinase ◽  
Gene Encoding ◽  
Threonine Kinase ◽  
Physiological Conditions ◽  
Different Levels ◽  
Familial Hyperkalemic Hypertension

With No Lysine (K) kinase 4 (WNK4) belongs to a serine-threonine kinase family characterized by the atypical positioning of its catalytic lysine. Despite the fact that WNK4 has been found in many tissues, the majority of its study has revolved around its function in the kidney, specifically as a positive regulator of the thiazide-sensitive NaCl cotransporter (NCC) in the distal convoluted tubule (DCT) of the nephron. This is explained by the description of gain-of-function mutations in the gene encoding WNK4 that cause Familial Hyperkalemic Hypertension (FHHt). This disease is mainly driven by increased downstream activation of the Ste20-related Proline Alanine Rich Kinase (SPAK)/Oxidative Stress Responsive Kinase 1 (OSR1)-NCC pathway, which increases salt reabsorption in the DCT and indirectly impairs renal K+ secretion. Here, we review the large volume of information that has accumulated about different aspects of WNK4 function. We first review the knowledge on WNK4 structure and enumerate the functional domains and motifs that have been characterized. Then, we discuss WNK4 physiological functions based on the information obtained from in vitro studies and from a diverse set of genetically modified mouse models with altered WNK4 function. We then review in vitro and in vivo evidence on the different levels of regulation of WNK4. Finally, we go through the evidence that has suggested how different physiological conditions act through WNK4 to modulate NCC activity.

scholarly journals Oxidative stress promotes liver cancer metastasis via a PKA-activated RNF25/ECAD/YAP circuit

2022 ◽  
Author(s):  
Zhao Huang ◽  
Li Zhou ◽  
Jiufei Duan ◽  
Siyuan Qin ◽  
Yu Wang ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Cancer Metastasis ◽  
Anoikis Resistance ◽  
Antioxidant Genes ◽  
Redox Sensor ◽  
Stress Signals ◽  
Hcc Cells

Abstract Loss of E-cadherin (ECAD), often caused by epigenetic inactivation, is closely associated with tumor metastasis. However, how ECAD is regulated in response to oxidative stress during tumorigenesis is largely unknown. Here we identify RNF25 as a new E3 ligase of ECAD, whose activation by oxidative stress leads to ECAD protein degradation in hepatocellular carcinoma (HCC). Loss of ECAD activates YAP, which in turn promotes the transcription of RNF25, thus forming a positive feedback loop to sustain the ECAD downregulation. YAP activation mitigates oxidative stress in detached HCC cells by upregulating antioxidant genes, protecting detached HCC cells from ferroptosis, resulting in anoikis resistance. Mechanistically, we found that protein kinase A (PKA) senses oxidative stress by redox modification in its β catalytic subunit (PRKACB) at Cys200 and Cys344, which increases its kinase activity towards RNF25 phosphorylation at Ser450, facilitating RNF25-mediated degradation of ECAD. Moreover, RNF25 expression is associated with HCC metastasis and depletion of RNF25 is sufficient to diminish HCC invasion and metastasis in vitro and in vivo. Together, these results identify a dual role of RNF25 as a critical regulator of ECAD protein turnover, promoting both anoikis resistance and metastasis, and PKA is a necessary redox sensor to enable this process. Our study provides mechanistic insight into how tumor cells sense oxidative stress signals to spread while escaping cell death.

scholarly journals Cystine/glutamate antiporter xCT (SLC7A11) facilitates oncogenic RAS transformation by preserving intracellular redox balance

2019 ◽  
Vol 116 (19) ◽  
pp. 9433-9442 ◽  
Author(s):  
Jonathan K. M. Lim ◽  
Alberto Delaidelli ◽  
Sean W. Minaker ◽  
Hai-Feng Zhang ◽  
Milena Colovic ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Transcription Factor ◽  
Redox Balance ◽  
Gene Encoding ◽  
Oncogenic Ras ◽  
Opposing Effects ◽  
Whole Transcriptome ◽  
Intracellular Redox

The RAS family of proto-oncogenes are among the most commonly mutated genes in human cancers and predict poor clinical outcome. Several mechanisms underlying oncogenic RAS transformation are well documented, including constitutive signaling through the RAF-MEK-ERK proproliferative pathway as well as the PI3K-AKT prosurvival pathway. Notably, control of redox balance has also been proposed to contribute to RAS transformation. However, how homeostasis between reactive oxygen species (ROS) and antioxidants, which have opposing effects in the cell, ultimately influence RAS-mediated transformation and tumor progression is still a matter of debate and the mechanisms involved have not been fully elucidated. Here, we show that oncogenic KRAS protects fibroblasts from oxidative stress by enhancing intracellular GSH levels. Using a whole transcriptome approach, we discovered that this is attributable to transcriptional up-regulation of xCT, the gene encoding the cystine/glutamate antiporter. This is in line with the function of xCT, which mediates the uptake of cystine, a precursor for GSH biosynthesis. Moreover, our results reveal that the ETS-1 transcription factor downstream of the RAS-RAF-MEK-ERK signaling cascade directly transactivates the xCT promoter in synergy with the ATF4 endoplasmic reticulum stress-associated transcription factor. Strikingly, xCT was found to be essential for oncogenic KRAS-mediated transformation in vitro and in vivo by mitigating oxidative stress, as knockdown of xCT strongly impaired growth of tumor xenografts established from KRAS-transformed cells. Overall, this study uncovers a mechanism by which oncogenic RAS preserves intracellular redox balance and identifies an unexpected role for xCT in supporting RAS-induced transformation and tumorigenicity.

scholarly journals Differential Involvement of Histidine Kinase Receptors in Pseudohyphal Development, Stress Adaptation, and Drug Sensitivity of the Opportunistic Yeast Candida lusitaniae

2007 ◽  
Vol 6 (10) ◽  
pp. 1782-1794 ◽  
Author(s):  
Florence Chapeland-Leclerc ◽  
Paméla Paccallet ◽  
Gwenaël Ruprich-Robert ◽  
David Reboutier ◽  
Christiane Chastin ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Histidine Kinase ◽  
High Sensitivity ◽  
Triple Mutant ◽  
Mating Ability ◽  
Reverse Transcription Pcr ◽  
Moderate Sensitivity ◽  
Candida Lusitaniae ◽  
High Level

ABSTRACT Fungal histidine kinase receptors (HKRs) sense and transduce many extracellular signals. We investigated the role of HKRs in morphogenetic transition, osmotolerance, oxidative stress response, and mating ability in the opportunistic yeast Candida lusitaniae. We isolated three genes, SLN1, NIK1, and CHK1, potentially encoding HKRs of classes VI, III, and X, respectively. These genes were disrupted by a transformation system based upon the “URA3 blaster” strategy. Functional analysis of disruptants was undertaken, except for the sln1 nik1 double mutant and the sln1 nik1 chk1 triple mutant, which are not viable in C. lusitaniae. The sln1 mutant revealed a high sensitivity to oxidative stress, whereas both the nik1 and chk1 mutants exhibited a more moderate sensitivity to peroxide. We also showed that the NIK1 gene was implicated in phenylpyrrole and dicarboximide compound susceptibility while HKRs seem not to be involved in resistance toward antifungals of clinical relevance. Concerning mating ability, all disruptants were still able to reproduce sexually in vitro in unilateral or bilateral crosses. The most important result of this study was that the sln1 mutant displayed a global defect of pseudohyphal differentiation, especially in high-osmolarity and oxidative-stress conditions. Thus, the SLN1 gene could be crucial for the C. lusitaniae yeast-to-pseudohypha morphogenetic transition. This implication is strengthened by a high level of SLN1 mRNAs revealed by semiquantitative reverse transcription-PCR when the yeast develops pseudohyphae. Our findings highlight a differential contribution of the three HKRs in osmotic and oxidant adaptation during the morphological transition in C. lusitaniae.

scholarly journals A Kayvirus Distant Homolog of Staphylococcal Virulence Determinants and VISA Biomarker Is a Phage Lytic Enzyme

Viruses ◽  
2020 ◽  
Vol 12 (3) ◽  
pp. 292
Author(s):  
Aleksandra Głowacka-Rutkowska ◽  
Magdalena Ulatowska ◽  
Joanna Empel ◽  
Magdalena Kowalczyk ◽  
Jakub Boreczek ◽  
...  
Keyword(s):  
Cell Walls ◽  
Bacterial Transport ◽  
Lytic Enzyme ◽  
Virulence Determinants ◽  
Gene Encoding ◽  
Phage Gene ◽  
E Coli ◽  
Lytic Transglycosylase ◽  
Distant Homolog

Staphylococcal bacteriophages of the Kayvirus genus are candidates for therapeutic applications. One of their proteins, Tgl, is slightly similar to two staphylococcal virulence factors, secreted autolysins of lytic transglycosylase motifs IsaA and SceD. We show that Tgl is a lytic enzyme secreted by the bacterial transport system and localizes to cell peripheries like IsaA and SceD. It causes lysis of E. coli cells expressing the cloned tgl gene, but could be overproduced when depleted of signal peptide. S. aureus cells producing Tgl lysed in the presence of nisin, which mimics the action of phage holin. In vitro, Tgl protein was able to destroy S. aureus cell walls. The production of Tgl decreased S. aureus tolerance to vancomycin, unlike the production of SceD, which is associated with decreased sensitivity to vancomycin. In the genomes of kayviruses, the tgl gene is located a few genes away from the lysK gene, encoding the major endolysin. While lysK is a late phage gene, tgl can be transcribed by a host RNA polymerase, like phage early genes. Taken together, our data indicate that tgl belongs to the kayvirus lytic module and encodes an additional endolysin that can act in concert with LysK in cell lysis.

scholarly journals Phase Variation of Ag43 Is Independent of the Oxidation State of OxyR

2003 ◽  
Vol 185 (7) ◽  
pp. 2203-2209 ◽  
Author(s):  
Anu Wallecha ◽  
Jason Correnti ◽  
Vincent Munster ◽  
Marjan van der Woude
Keyword(s):  
Oxidative Stress ◽  
Hydrogen Peroxide ◽  
Oxidation State ◽  
Regulatory Region ◽  
Phase Variation ◽  
In Vitro Transcription ◽  
Reduced Form ◽  
Target Sequences

ABSTRACT OxyR is a DNA binding protein that differentially regulates a cell's response to hydrogen peroxide-mediated oxidative stress. We previously reported that the reduced form of OxyR is sufficient for repression of transcription of agn43 from unmethylated template DNA, which is essential for deoxyadenosine methylase (Dam)- and OxyR-dependent phase variation of agn43. Here we provide evidence that the oxidized form of OxyR [OxyR(ox)] also represses agn43 transcription. In vivo, we found that exogenous addition of hydrogen peroxide, sufficient to oxidize OxyR, did not affect the expression of agn43. OxyR(ox) repressed in vitro transcription but only from an unmethylated agn43 template. The −10 sequence of the promoter and three Dam target sequences were protected in an in vitro DNase I footprint assay by OxyR(ox). Furthermore, OxyR(ox) bound to the agn43 regulatory region DNA with an affinity similar to that for the regulatory regions of katG and oxyS, which are activated by OxyR(ox), indicating that binding at agn43 can occur at biologically relevant concentrations. OxyR-dependent regulation of Ag43 expression is therefore unusual in firstly that OxyR binding at agn43 is dependent on the methylation state of Dam target sequences in its binding site and secondly that OxyR-dependent repression appears to be independent of hydrogen-peroxide mediated oxidative stress and the oxidation state of OxyR.

scholarly journals DDB2, an Essential Mediator of Premature Senescence

2010 ◽  
Vol 30 (11) ◽  
pp. 2681-2692 ◽  
Author(s):  
Nilotpal Roy ◽  
Tanya Stoyanova ◽  
Carmen Dominguez-Brauer ◽  
Hyun Jung Park ◽  
Srilata Bagchi ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Dna Damage ◽  
Excision Repair ◽  
Culture Shock ◽  
Premature Senescence ◽  
Antioxidant Genes ◽  
Repair Protein ◽  
High Level

ABSTRACT Reactive oxygen species (ROS) is critical for premature senescence, a process significant in tumor suppression and cancer therapy. Here, we reveal a novel function of the nucleotide excision repair protein DDB2 in the accumulation of ROS in a manner that is essential for premature senescence. DDB2-deficient cells fail to undergo premature senescence induced by culture shock, exogenous oxidative stress, oncogenic stress, or DNA damage. These cells do not accumulate ROS following DNA damage. The lack of ROS accumulation in DDB2 deficiency results from high-level expression of the antioxidant genes in vitro and in vivo. DDB2 represses antioxidant genes by recruiting Cul4A and Suv39h and by increasing histone-H3K9 trimethylation. Moreover, expression of DDB2 also is induced by ROS. Together, our results show that, upon oxidative stress, DDB2 functions in a positive feedback loop by repressing the antioxidant genes to cause persistent accumulation of ROS and induce premature senescence.

Role of the Vitamin D Upregulated Protein 1 (VDUP1)-Controlled Stress Pathway in Acute Myeloid Leukemia.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 3386-3386
Author(s):  
Stefan J. Erkeland ◽  
Marijke Valkhof ◽  
Astrid Danen-van Oorschot ◽  
Ivo P. Touw
Keyword(s):  
Oxidative Stress ◽  
Vitamin D ◽  
Stress Responses ◽  
Ectopic Expression ◽  
Gene Encoding ◽  
Protein Levels ◽  
Lipid Phosphatases ◽  
Multiple Virus ◽  
Dual Specific Phosphatase

Abstract Graffi-1.4 (Gr-1.4) and CasBrM murine leukemia viruses induce myeloid leukemias due to deregulation of genes by proviral integration. Some of these genes, such as peroxiredoxin (PRDX-2), vitamin D upregulated protein 1 (VDUP1), and dual specific phosphatase 10 (DUSP10), are involved in the regulation of reactive oxygen species (ROS) induced stress pathways. ROS are known to influence signal transduction by modulating the activity of protein and lipid phosphatases and cell differentiation at relatively low levels, whereas high ROS levels induce apoptosis. Perturbations of the cellular redox state have a high impact on these processes. VDUP1 is a recently identified oxidative stress-responsive gene that suppresses thioredoxin activity in cardiomyocytes, thereby controlling cell survival. Integrations in the gene encoding VDUP1 all occurred at the 5′ and 3′ region with a frequency of 100% of the Gr-1.4 and 62% of the CasBrM induced leukemias. All cases of CasBrM leukemias with 3′ integrations (33% of leukemias) have additional integrations at the 5′end of VDUP1, suggesting that multiple virus integration sites cooperate in gene deregulation. A hotspot of integrations was found around 900 base pairs upstream of the ATG, near two newly identified heat shock elements. The presence of Gr-1.4 LTR sequences in the VDUP1 promoter and in the 3′ untranslated region results in a 2–2.5 times enhanced luciferase signal when compared to normal promoter activity. This effect was even greater (up to 6-fold) under stress conditions, suggesting that normal VDUP1 regulation is disrupted by viral integration. In human AML, we found that expression of VDUP1 transcripts is different in distinct patient clusters recently identified by gene expression profiling (Valk et al NEJM 2004, 350:1617-28), suggesting a specific involvement of this gene in certain subgroups of AML. For instance, AML samples exhibiting t(8;21) have significantly higher VDUP1 transcript levels compared to other AML patients. Furthermore, high VDUP1 protein levels significantly correlated with FAB classifications M4 and M5 and with younger age (<35 yrs), whereas low VDUP1 expression were found in FAB-M1 and M2 and in patients older than 50 years. To study the consequences of VDUP1 overexpression in normal myelopoiesis, we overexpressed the gene in murine hematopoietic progenitors by retroviral gene transfer and performed in vitro colony assays with G-CSF and liquid culture assay with different cytokine cocktails. Irrespective of the cytokine used, ectopic expression of VDUP1 resulted in accelerated apoptosis and inhibited proliferation, indicating that deregulation of VDUP1 as a single event does not confer a growth advantage and implying that additional events are needed for full leukemic transformation of myeloid precursors. DUSP10 or other members of the DUSP family might be candidates, as we observed that overexpression of DUSP10 in myeloid 32D cells decreased oxidative stress-induced activation of JNK and p38MAPK and inhibited apoptosis. We are currently generating 32D models in which VDUP1 and DUSP10 can be inducibly expressed to further address this hypothesis. In conclusion, we found that VDUP1 expression is frequently enhanced in mouse leukemia models due to viral integrations and also in distinct subgroups of human AML. Our data thus identify disruption of VDUP1-controlled oxidative stress responses as a novel mechanism involved in the pathogenesis of AML.

Sign in / Sign up

Export Citation Format

Share Document