Reactive Oxygen Species Sensor Research Articles

Crosstalk Between Vascular Redox and Calcium Signaling in Hypertension Involves TRPM2 (Transient Receptor Potential Melastatin 2) Cation Channel.

Increased generation of reactive oxygen species (ROS) and altered Ca2+ handling cause vascular damage in hypertension. Mechanisms linking these systems are unclear, but TRPM2 (transient receptor potential melastatin 2) could be important because TRPM2 is a ROS sensor and a regulator of Ca2+ and Na+ transport. We hypothesized that TRPM2 is a point of cross-talk between redox and Ca2+ signaling in vascular smooth muscle cells (VSMC) and that in hypertension ROS mediated-TRPM2 activation increases [Ca2+]i through processes involving NCX (Na+/Ca2+ exchanger). VSMCs from hypertensive and normotensive individuals and isolated arteries from wild type and hypertensive mice (LinA3) were studied. Generation of superoxide anion and hydrogen peroxide (H2O2) was increased in hypertensive VSMCs, effects associated with activation of redox-sensitive PARP1 (poly [ADP-ribose] polymerase 1), a TRPM2 regulator. Ang II (angiotensin II) increased Ca2+ and Na+ influx with exaggerated responses in hypertension. These effects were attenuated by catalase-polyethylene glycol -catalase and TRPM2 inhibitors (2-APB, 8-Br-cADPR olaparib). TRPM2 siRNA decreased Ca2+ in hypertensive VSMCs. NCX inhibitors (Benzamil, KB-R7943, YM244769) normalized Ca2+ hyper-responsiveness and MLC20 phosphorylation in hypertensive VSMCs. In arteries from LinA3 mice, exaggerated agonist (U46619, Ang II, phenylephrine)-induced vasoconstriction was decreased by TRPM2 and NCX inhibitors. In conclusion, activation of ROS-dependent PARP1-regulated TRPM2 contributes to vascular Ca2+ and Na+ influx in part through NCX. We identify a novel pathway linking ROS to Ca2+ signaling through TRPM2/NCX in human VSMCs and suggest that oxidative stress-induced upregulation of this pathway may be a new player in hypertension-associated vascular dysfunction.

Regulation of apoptosis in the ischemic penumbra in the first day post-stroke.

Regulation of apoptosis in the ischemic penumbra in the first day post-stroke.

Manganese‐Coordinated Tyrosine Bio Materials for the Sensing of Reactive Oxygen Species

AbstractDesign of biosensor materials for the recognition of reactive oxygen species (ROS) inside living cells is necessary for the early diagnosis of cancer and other diseases induced by ROS. Here, the B3LYP−D3 dispersion‐corrected and Mo6 hybrid density functional theoretical methods and a polarized continuum model to account for the aqueous medium are used to propose models of manganese (Mn+2)‐coordinated tyrosine (Tyr) nano rods and porous bio materials to detect various ROS, such as .OH, .OOH, H2O2, .NO, ONOO−, and O2.−. It is found that the nano rods and porous polymers may sense the presence of different ROS by utilizing their edges and pores respectively. It is further revealed that due to stable binding and adsorption energies, entrapment of different ROS by the porous polymers would be sustained for longer time compared to the nano rods. However, the edges of nano rods are reactive and may initiate oxidation reactions by converting Mn+2 to Mn+3. It is thus expected that these bio materials would act as excellent ROS sensors.

Pejvakin-mediated pexophagy protects auditory hair cells against noise-induced damage

Noise overexposure causes oxidative stress, leading to auditory hair cell damage. Adaptive peroxisome proliferation involving pejvakin, a peroxisome-associated protein from the gasdermin family, has been shown to protect against this harmful oxidative stress. However, the role of pejvakin in peroxisome dynamics and homeostasis remains unclear. Here we show that sound overstimulation induces an early and rapid selective autophagic degradation of peroxisomes (pexophagy) in auditory hair cells from wild-type, but not pejvakin-deficient (Pjvk -/-), mice. Noise overexposure triggers recruitment of the autophagosome-associated protein MAP1LC3B (LC3B; microtubule-associated protein 1 light chain 3β) to peroxisomes in wild-type, but not Pjvk -/-, mice. We also show that pejvakin-LC3B binding involves an LC3-interacting region within the predicted chaperone domain of pejvakin. In transfected cells and in vivo transduced auditory hair cells, cysteine mutagenesis experiments demonstrated the requirement for both C328 and C343, the two cysteine residues closest to the C terminus of pejvakin, for reactive oxygen species-induced pejvakin-LC3B interaction and pexophagy. The viral transduction of auditory hair cells from Pjvk -/- mice in vivo with both Pjvk and Lc3b cDNAs completely restored sound-induced pexophagy, fully prevented the development of oxidative stress, and resulted in normal levels of peroxisome proliferation, whereas Pjvk cDNA alone yielded only a partial correction of the defects. Overall, our results demonstrate that pexophagy plays a key role in noise-induced peroxisome proliferation and identify defective pexophagy as a cause of noise-induced hearing loss. They suggest that pejvakin acts as a redox-activated pexophagy receptor/adaptor, thereby identifying a previously unknown function of gasdermin family proteins.

Hydrogen Peroxide and Redox Regulation of Developments.

Reactive oxygen species (ROS), which were originally classified as exclusively deleterious compounds, have gained increasing interest in the recent years given their action as bona fide signalling molecules. The main target of ROS action is the reversible oxidation of cysteines, leading to the formation of disulfide bonds, which modulate protein conformation and activity. ROS, endowed with signalling properties, are mainly produced by NADPH oxidases (NOXs) at the plasma membrane, but their action also involves a complex machinery of multiple redox-sensitive protein families that differ in their subcellular localization and their activity. Given that the levels and distribution of ROS are highly dynamic, in part due to their limited stability, the development of various fluorescent ROS sensors, some of which are quantitative (ratiometric), represents a clear breakthrough in the field and have been adapted to both ex vivo and in vivo applications. The physiological implication of ROS signalling will be presented mainly in the frame of morphogenetic processes, embryogenesis, regeneration, and stem cell differentiation. Gain and loss of function, as well as pharmacological strategies, have demonstrated the wide but specific requirement of ROS signalling at multiple stages of these processes and its intricate relationship with other well-known signalling pathways.

Activation of the intracellular temperature and ROS sensor membrane protein STIM1 as a mechanism underpinning biological effects of low-level low frequency magnetic fields

The biological effects of low frequency magnetic fields (LF MF) at high flux densities are well known and the underlying mechanism is established. In contrast, health associated effects at lower flux densities, which can be found in the human environment, are controversial, and no accepted interaction mechanism has been presented.Here we present a hypothesis regarding the biological aspect of interaction between LF MF and cells. We suggest that the endoplasmic reticulum (ER) membrane protein STIM1, which functions as a sensor for several cellular conditions (low Ca2+ levels, temperature increase, increased levels of oxygen radicals, hypoxia), is a candidate LF MF sensor. Such a sensor function can be either direct (via local temperature increase caused by intracellularly induced electric fields), or indirect due to responses to increased reactive oxygen species (ROS) levels. Activated STIM1 leads to downstream effects by activation of signal transduction processes and changes in gene expression leading to secondary events. The nature of these changes would be dependent on both cell type and the particular physiological state the cell displays at the time of STIM1 activation.Results from testing of this hypothesis, as suggested in this paper, would greatly assist in understanding of the possible health-related effects of low-level LF MF. This would benefit both safety assessments regarding MF exposure as well as possible use of MF in medicine.A better understanding of the biological mechanisms underpinning MF exposure effects of living matter allows the targeted use of the fields in medical applications. There are several examples already in use based on empiric and not on mechanistic knowledge. Knowledge generated from our hypothesis testing makes it possible for MF based medical applications to be optimized.

A gold nanoparticle system for the enhancement of radiotherapy and simultaneous monitoring of reactive-oxygen-species formation

Gold nanoparticles (AuNPs) are known to sensitize cancer cells to radiation therapy (RT) by increasing the deposition of ionizing energy in their immediate vicinity. However, this process of dose enhancement is challenging to monitor because it is heterogeneous at the sub-cellular scale. Furthermore, radiation damage is primarily mediated by reactive oxygen species (ROS) that are produced following water radiolysis. Here, radiation-responsive PEGylated gold nanoparticles (RPAuNPs) were synthesized for the enhanced generation and concurrent detection of ROS in cancer cells and tumors. PEGylated gold particles (20 nm diameter) were functionalized with dihydrorhodamine 123 (DHR-123), a known ROS sensor, to monitor ROS generation in their immediate vicinity. These NPs were able to effectively radiosensitize cells, as measured by increased cell apoptosis following RT. Furthermore, the fluorescence of these RPAuNPs was 7-fold higher after 6 Gy RT due to the local production of ROS near the surface of the NP. Finally, multispectral fluorescence imaging was used to monitor NP-induced ROS in vivo, following conformal RT, in a xenograft model of breast cancer. This theranostic NP system provides a novel approach for monitoring the nanoscale enhancement of RT by high-Z metal NPs.

Mitochondrial ROS Drive Sudden Cardiac Death and Chronic Proteome Remodeling in Heart Failure.

Despite increasing prevalence and incidence of heart failure (HF), therapeutic options remain limited. In early stages of HF, sudden cardiac death (SCD) from ventricular arrhythmias claims many lives. Reactive oxygen species (ROS) have been implicated in both arrhythmias and contractile dysfunction. However, little is known about how ROS in specific subcellular compartments contribute to HF or SCD pathophysiology. The role of ROS in chronic proteome remodeling has not been explored. We will test the hypothesis that elevated mitochondrial ROS (mROS) is a principal source of oxidative stress in HF and in vivo reduction of mROS mitigates SCD. Using a unique guinea pig model of nonischemic HF that recapitulates important features of human HF, including prolonged QT interval and high incidence of spontaneous arrhythmic SCD, compartment-specific ROS sensors revealed increased mROS in resting and contracting left ventricular myocytes in failing hearts. Importantly, the mitochondrially targeted antioxidant (MitoTEMPO) normalized global cellular ROS. Further, in vivo MitoTEMPO treatment of HF animals prevented and reversed HF, eliminated SCD by decreasing dispersion of repolarization and ventricular arrhythmias, suppressed chronic HF-induced remodeling of the expression proteome, and prevented specific phosphoproteome alterations. Pathway analysis of mROS-sensitive networks indicated that increased mROS in HF disrupts the normal coupling between cytosolic signals and nuclear gene programs driving mitochondrial function, antioxidant enzymes, Ca2+ handling, and action potential repolarization, suggesting new targets for therapeutic intervention. mROS drive both acute emergent events, such as electrical instability responsible for SCD, and those that mediate chronic HF remodeling, characterized by suppression or altered phosphorylation of metabolic, antioxidant, and ion transport protein networks. In vivo reduction of mROS prevents and reverses electrical instability, SCD, and HF. Our findings support the feasibility of targeting the mitochondria as a potential new therapy for HF and SCD while identifying new mROS-sensitive protein modifications.

P-291 - Using redox proteomics to identify oxidized proteins in Arabidopsis roots that modulate development

Reactive oxygen species (ROS) are second messengers that reversibly oxidize cysteine residues to sulfenic acids to alter the activity of redox-sensitive proteins. To cope with high levels of ROS, plants have evolved elaborate mechanisms including synthesis of ROS scavenging proteins and small molecule antioxidants, such as flavonols. The Arabidopsis tt4 mutant produces no flavonols and has increased numbers of root hairs and lateral roots. Consistent with the absence of flavonol antioxidants, tt4 has increased fluorescence from two ROS sensors, DCF and Peroxy Orange 1, which detect all ROS and H2O2, respectively. A mutation in the gene encoding RBOHE leads to reduced ROS and decreased lateral roots. These results are consistent with flavonol-modulated ROS altering lateral root emergence. We are using a dimedone probe, DCP-BIOTIN1, which binds to sulfenic acids to isolate oxidized proteins. The abundance of oxidized proteins is elevated in tt4 compared to wild type. A proteomic approach is being used to identify which proteins are differentially oxidized between tt4 and wild-type. The function of these proteins will be examined using mutants with the goal of identifying the ROS signaling network that defines root architecture.

Role of mitochondrial reactive oxygen species (ROS) and TRPV4 activation in microvascular endothelial cell dysfunction in PAH.

RationaleIn pulmonary arterial hypertension (PAH), endothelial cells (ECs) exhibit increased migration/proliferation leading to formation of vaso‐occlusive lesions. The mechanisms underlying this pro‐migratory/proliferative state remain unclear. Increased ROS and intracellular Ca2+ concentration ([Ca2+]i) are known to promote migration and proliferation in other vascular beds, and there is evidence for increased oxidant stress and basal [Ca2+]i in ECs from humans with PAH. Although the exact source of increased EC oxidant stress in PAH ECs is not fully known, mitochondria are a significant source of ROS in ECs, and mitochondrial dysfunction is present in human PAH ECs. However, the links between ROS production, elevated [Ca2+]i and enhanced migration and proliferation in PAH ECs are incompletely understood.We previously showed that exogenous ROS increased [Ca2+]i in lung microvascular endothelial cells (MVECs) via the transient receptor potential vanilloid‐4 (TRPV4) calcium channel. To study endogenous ROS production and the possible role of ROS‐induced Ca2+ influx in promoting MVEC migration and proliferation in PAH, we isolated MVECs from the lungs of rats subjected to normoxia (N‐MVEC) or SU5416/hypoxia (SuHx‐MVEC), a robust model of PAH, and examined ROS production, [Ca2+]i, mitochondrial morphology/function and cellular migration and proliferation.MethodsCa2+ was measured using Fura‐2AM‐loaded MLMVEC in a flow chamber perfused with Krebs buffer. Mitochondrial morphology measurements were made using validated algorithms on confocal images of mitochondria in N‐ and SuHx‐MVEC transfected with MitoRFP. ROS measurements were made using the ratiometric ROS sensor roGFP. Migration and proliferation were measured using transwell assay and BrDU incorporation, respectively.ResultsROS levels were increased in SuHx‐MVEC and attenuated following treatment with MitoQ (MQ) a mitochondrion‐specific antioxidant. In SuHx‐MVECs, mitochondrial number, network fragmentation and activation of the fission‐inducing protein dynamin‐related protein 1 were increased, while oxidative phosphorylation and mitochondrial membrane potential were both decreased. Basal [Ca2+]i, migration and proliferation were all increased in SuHx‐MVEC, but similarly attenuated following treatment with: 1) MQ; 2) a global ROS scavenger (TEMPOL); or 3) HC‐067047, a specific TRPV4 inhibitor. While total TRPV4 expression was unchanged between N‐ and SuHx‐MVEC, activation of membrane bound TRPV4 (using the specific agonist GSK1016790A ‐ GSK) produced greater [Ca2+]i influx in SuHx‐MVEC. Biotinylation of TRPV4 revealed greater membrane‐bound expression of TRPV4 in SuHx‐MVEC. Interestingly, quenching mitochondrial ROS attenuated GSK‐induced [Ca2+]i influx in SuHx‐MVEC.ConclusionThese data suggest that, in SuHx‐MVEC, mitochondrial ROS promote membrane translocation of TRPV4, leading to increased basal [Ca2+]i and enhanced migration and proliferation.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

On suitable observables for the chaotic Peroxidase-Oxidase autocatalytic reaction

Neurodegenerative diseases such as Alzheimer’s and Parkinson present oxidative damage of neurocytes generally caused by reactive oxygen species (ROS). Heme peroxidases consume and produce ROS thus serving as suppresors and promoters of ROS-related pathology A core reaction network of production and consumption of hydrogen peroxide and superoxide is the Peroxidase-Oxidase (PO). PO reaction was among the first biochemical oscillators to be experimentally and numerically studied, with dynamical scenarios such as multiple steady states, complex oscillations and chaos. In particular, the oxyferrous peroxidase (compound III) and O2 have been used as observable variables in experiments to record the chaotic dynamics. Moreover, PO reaction has been embedded in nanoparticles in order to serve as a sensor for ROS. In this contribution we quantified, using observability coefficients, which chemical species is more suitable to observe and reconstruct the characteristic complex dynamics of the PO reaction.

Oxeiptosis, a ROS-induced caspase-independent apoptosis-like cell-death pathway.

Reactive oxygen species (ROS) are generated by virally-infected cells however the physiological significance of ROS generated under these conditions is unclear. Here we show that inflammation and cell death induced by exposure of mice or cells to sources of ROS is not altered in the absence of canonical ROS-sensing pathways or known cell death pathways. ROS-induced cell death signaling involves interaction between the cellular ROS sensor and antioxidant factor KEAP1, the phosphatase PGAM5 and the proapoptotic factor AIFM1. Pgam5−/− mice show exacerbated lung inflammation and proinflammatory cytokines in an ozone exposure model. Similarly, challenge with influenza A virus leads to increased virus infiltration, lymphocytic bronchiolitis and reduced survival of Pgam5−/− mice. This pathway, which we term ‘oxeiptosis’, is a ROS-sensitive, caspase independent, non-inflammatory cell death pathway and is important to protect against inflammation induced by ROS or ROS-generating agents such as viral pathogens.

Abscisic Acid-Induced Reactive Oxygen Species Are Modulated by Flavonols to Control Stomata Aperture.

Abscisic acid (ABA) increases reactive oxygen species (ROS) in guard cells to close Arabidopsis (Arabidopsis thaliana) stomata. In tomato (Solanum lycopersicum), we find that ABA-increased ROS is followed by stomatal closure and that both responses are blocked by inhibitors of ROS-producing respiratory burst oxidase enzymes. ABA-induced ROS sensor fluorescence accumulates in the nucleus, chloroplasts, and endomembranes. The accumulation of flavonol antioxidants in guard cells, but not surrounding pavement cells, was visualized by confocal microscopy using a flavonol-specific fluorescent dye. Decreased flavonols in guard cells in the anthocyanin reduced (are) mutant and elevated levels in the anthocyanin without (aw) mutant were quantified by confocal microscopy and in leaf extracts by mass spectrometry. Consistent with flavonols acting as antioxidants, higher levels of ROS were detected in guard cells of the tomato are mutant and lower levels were detected in aw both at homeostasis and after treatment with ABA. These results demonstrate the inverse relationship between flavonols and ROS. Guard cells of are show greater ABA-induced closure than the wild type, reduced light-dependent guard cell opening, and reduced water loss, with aw having opposite responses. Ethylene treatment of wild-type tomato plants increased flavonol accumulation in guard cells; however, no flavonol increases were observed in Neverripe (Nr), an ethylene receptor mutant. Consistent with lower levels of ROS due to elevated flavonols, ethylene treatments decreased ABA-induced stomatal closure in the wild type, but not Nr, with ethylene responses attenuated in the are mutant. Together, these results are consistent with flavonols dampening the ABA-dependent ROS burst that drives stomatal closure and facilitating stomatal opening to modulate leaf gas exchange.

Ascorbic acid alters cell fate commitment of human neural progenitors in a WNT/\u03b2-catenin/ROS signaling dependent manner

BackgroundImproving the neuronal yield from in vitro cultivated neural progenitor cells (NPCs) is an essential challenge in transplantation therapy in neurological disorders. In this regard, Ascorbic acid (AA) is widely used to expand neurogenesis from NPCs in cultures although the mechanisms of its action remain unclear. Neurogenesis from NPCs is regulated by the redox-sensitive WNT/β-catenin signaling pathway. We therefore aimed to investigate how AA interacts with this pathway and potentiates neurogenesis.MethodsEffects of 200 μM AA were compared with the pro-neurogenic reagent and WNT/β-catenin signaling agonist lithium chloride (LiCl), and molecules with antioxidant activities i.e. N-acetyl-L-cysteine (NAC) and ruthenium red (RuR), in differentiating neural progenitor ReNcell VM cells. Cells were supplemented with reagents for two periods of treatment: a full period encompassing the whole differentiation process versus an early short period that is restricted to the cell fate commitment stage. Intracellular redox balance and reactive oxygen species (ROS) metabolism were examined by flow cytometry using redox and ROS sensors. Confocal microscopy was performed to assess cell viability, neuronal yield, and levels of two proteins: Nucleoredoxin (NXN) and the WNT/β-catenin signaling component Dishevelled 2 (DVL2). TUBB3 and MYC gene responses were evaluated by quantitative real-time PCR. DVL2-NXN complex dissociation was measured by fluorescence resonance energy transfer (FRET).ResultsIn contrast to NAC which predictably exhibited an antioxidant effect, AA treatment enhanced ROS metabolism with no cytotoxic induction. Both drugs altered ROS levels only at the early stage of the differentiation as no changes were held beyond the neuronal fate commitment stage. FRET studies showed that AA treatment accelerated the redox-dependent release of the initial pool of DVL2 from its sequestration by NXN, while RuR treatment hampered the dissociation of the two proteins. Accordingly, AA increased WNT/β-catenin signaling output i.e. MYC mRNA level, whereas RuR attenuated it. Moreover, AA improved neurogenesis as much as LiCl as both TUBB3-positive cell yield and TUBB3 mRNA level increased, while NAC or RuR attenuated neurogenesis. Markedly, the neurogenesis outputs between the short and the full treatment with either NAC or AA were found unchanged, supporting our model that neuronal yield is altered by events taking place at the early phase of differentiation.ConclusionsOur findings demonstrate that AA treatment elevates ROS metabolism in a non-lethal manner prior to the NPCs commitment to their neuronal fate. Such effect stimulates the redox-sensitive DVL2 activation and WNT/β-catenin signaling response that would enhance the ensuing neuronal cell differentiation.

CaMKII oxidation causes increased atrial fibrillation in diabetic mice

Background: Diabetes mellitus (DM) is an independent risk factor for atrial fibrillation (AF), the most frequent clinical arrhythmia, but the mechanism(s) underlying this clinical association are unknown. Increased reactive oxygen species (ROS) and O-GlcNAcylation (OGN) is a hallmark of DM. Hyperactivated CaMKII promotes AF, and CaMKII may become constitutively active and proarrhythmic by oxidation (ox-CaMKII) or OGN (OGN-CaMKII), post translational modifications to adjacent amino acids in the regulatory domain. We hypothesize that CaMKII promotes diabetes-induced AF through ox-CaMKII and OGN-CaMKII. Methods and Results: We generated knock-in mouse models, based on the predominant CaMKII isoform in myocardium (CaMKIIδ), to selectively eliminate ox-CaMKII and OGN-CaMKII. We used the pancreatic β-cell toxin, streptozocin (STZ) or a combination of high fat diet and STZ, to generate validated mouse models of T1D and T2D, and an established right atrial pacing model of paroxysmal AF. T1D and T2D mice showed increased AF [70% (14/20), p < 0.05; and 61% (11/18), p < 0.05] compared to non-diabetic controls [25% (5/20)], suggesting diabetes was a risk factor for AF in mice. We focused on T1D because it is a simpler model system, compared to T2D. We found that enhanced AF risk was prevented in mice with T1D by myocardial CaMKII inhibition due to transgenic expression of a CaMKII inhibitory peptide, AC3-I, suggesting that CaMKII is required for AF in diabetes. Atria from T1D mice showed increased ROS, OGN, and ox-CaMKII. Mice selectively resistant to ox-CaMKII due to knock-in replacement of regulatory domain methionines with valines (MMVV) with T1D were resistant to AF. In contrast, knock-in mice with putative OGN resistant CaMKII (S280A) and wild type (WT) littermate mice with T1D showed similar increases in AF, indicating that OGN-CaMKII did not contribute to AF. WT T1D atrial myocytes, but not MMVV T1D atrial myocytes, showed increased RyR2 Ca2+ leak, delayed afterdepolarizations (DADs) and triggered action potentials, indicating ox-CaMKII coupled ROS to a fundamental proarrhythmic cellular pathway. Addition of an OGN antagonist prevented increased AF, RyR2 Ca2+ leak, DADs and triggered action potentials. Knock-in mice lacking a CaMKII phosphorylation site on RyR2 (S2814A) with T1D were resistant to AF, compared to WT T1D controls, establishing the contribution of CaMKII and RyR2 to AF in this model, and suggesting that OGN is proarrhythmic by a CaMKII independent pathway involving RyR2. Conclusions: These studies establish CaMKII as a critical ROS sensor and proarrhythmic signal in AF, and suggest that ox-CaMKII and OGN converge on RyR2 to promote AF in diabetes. These provide new insights for understanding important clinical phenotypes that currently lack clear mechanisms and adequate therapies.

PML is a ROS sensor activating p53 upon oxidative stress.

Promyelocytic leukemia (PML) nuclear bodies (NBs) recruit partner proteins, including p53 and its regulators, thereby controlling their abundance or function. Investigating arsenic sensitivity of acute promyelocytic leukemia, we proposed that PML oxidation promotes NB biogenesis. However, physiological links between PML and oxidative stress response in vivo remain unexplored. Here, we identify PML as a reactive oxygen species (ROS) sensor. Pml-/- cells accumulate ROS, whereas PML expression decreases ROS levels. Unexpectedly, Pml-/- embryos survive acute glutathione depletion. Moreover, Pml-/- animals are resistant to acetaminophen hepatotoxicity or fasting-induced steatosis. Molecularly, Pml-/- animals fail to properly activate oxidative stress-responsive p53 targets, whereas the NRF2 response is amplified and accelerated. Finally, in an oxidative stress-prone background, Pml-/- animals display a longevity phenotype, likely reflecting decreased basal p53 activation. Thus, similar to p53, PML exerts basal antioxidant properties but also drives oxidative stress-induced changes in cell survival/proliferation or metabolism in vivo. Through NB biogenesis, PML therefore couples ROS sensing to p53 responses, shedding a new light on the role of PML in senescence or stem cell biology.

Early and Late Induction of KRAS and HRAS Proto-Oncogenes by Reactive Oxygen Species in Primary Astrocytes

Astrocytes, one of the predominant types of glial cells, function as both supportive and metabolic cells for the brain. Among mammalian tissues, the highest levels of p21Ras protein are detected in the brain. Here, we investigated the expression of KRAS and HRAS proto-oncogenes in primary astrocytes following acute oxidative stimulation. Reactive oxygen species (ROS) changed the expression of proto-oncogenes at both transcriptional and translational levels. De novo protein synthesis analysis measured approximate values of proteins half-life, ranging from 1–4 h, of the different H- and K- isoforms by western blot analysis. Quantitative gene expression analysis of KRAS and HRAS revealed an unexpected short-term induction of KRAS mRNA in primary astrocytes in response to acute stimulation. Indeed, cultured astrocytes responded to proteasomal inhibition by preventing the reduction of c-K-Ras. A fraction of K-Ras protein accumulated in the presence of ROS and cycloheximide, while a substantial proportion was continuously synthesized. These data indicate that ROS regulate in a complementary fashion p21Ras isoforms in primary astrocytes: K-Ras is rapidly and transiently induced by post-translational and post-transcriptional mechanisms, while H-Ras is stably induced by mRNA accumulation. We suggest that K-Ras and H-Ras are ROS sensors that adapt cells to metabolic needs and oxidative stress.

Bound by Fate: The Role of Reactive Oxygen Species in Receptor-Like Kinase Signaling.

In plants, receptor-like kinases (RLKs) and extracellular reactive oxygen species (ROS) contribute to the communication between the environment and the interior of the cell. Apoplastic ROS production is a frequent result of RLK signaling in a multitude of cellular processes; thus, by their nature, these two signaling components are inherently linked. However, it is as yet unclear how ROS signaling downstream of receptor activation is executed. In this review, we provide a broad view of the intricate connections between RLKs and ROS signaling and describe the regulatory events that control and coordinate extracellular ROS production. We propose that concurrent initiation of ROS-dependent and -independent signaling linked to RLKs might be a critical element in establishing cellular responses. Furthermore, we discuss the possible ROS sensing mechanisms in the context of the biochemical environment in the apoplast. We suggest that RLK-dependent modulation of apoplastic and intracellular conditions facilitates ROS perception and signaling. Based on data from plant and animal models, we argue that specific RLKs could be components of the ROS sensing machinery or ROS sensors. The importance of the crosstalk between RLK and ROS signaling is discussed in the context of stomatal immunity. Finally, we highlight challenges in the understanding of these signaling processes and provide perspectives for future research.

Mitochondrial and Oxidative Stress-Mediated Activation of Protein Kinase D1 and Its Importance in Pancreatic Cancer

Due to alterations in their metabolic activity and decreased mitochondrial efficiency, cancer cells often show increased generation of reactive oxygen species (ROS), but at the same time, to avoid cytotoxic signaling and to facilitate tumorigenic signaling, have mechanism in place that keep ROS in check. This requires signaling molecules that convey increases in oxidative stress to signal to the nucleus to upregulate antioxidant genes. Protein kinase D1 (PKD1), the serine/threonine kinase, is one of these ROS sensors. In this mini-review, we highlight the mechanisms of how PKD1 is activated in response to oxidative stress, so far known downstream effectors, as well as the importance of PKD1-initiated signaling for development and progression of pancreatic cancer.

Detection of Reactive Oxygen Species by a Carbon-Dot-Ascorbic Acid Hydrogel.

Detection of reactive oxygen species (ROS) is important in varied biological processes, disease diagnostics, and chemotherapeutic drug screening. We constructed a ROS sensor comprising an ascorbic-acid-based hydrogel encapsulating luminescent amphiphilic carbon-dots (C-dots). The sensing mechanism is based upon ROS-induced oxidation of the ascorbic acid units within the hydrogel scaffold; as a consequence, the hydrogel framework collapses resulting in aggregation of the C-dots and quenching of their luminescence. The C-dot-hydrogel platform exhibits high sensitivity and detected ROS generated chemically in solution and in actual cell environments. We demonstrate application of the C-dot-hydrogel for evaluating the efficacy of a chemotherapeutic substance, underscoring the potential of the system for drug screening applications.