Oxygen-sensing Mechanism Research Articles

Functional MRI during hyperbaric oxygen: Effects of oxygen on neurovascular coupling and BOLD fMRI signals

Hyperbaric oxygen (HBO) therapy is used to treat a number of ailments. Improved understanding of how HBO affects neuronal activity, cerebral blood flow (CBF) and blood-oxygenation-level dependent (BOLD) changes could shed light on the role of oxygen in neurovascular coupling and help guide HBO treatments. The goal of this study was to test two hypotheses: i) activation-induced CBF fMRI response is not dependent on hemoglobin deoxygenation, and ii) activation-induced BOLD fMRI is markedly attenuated under HBO. CBF and BOLD fMRI of forepaw stimulation in anesthetized rats under HBO at 3 atmospheres absolute (ATA) were compared with normobaric air. Robust BOLD and CBF fMRI were detected under HBO. Inflow effects and spin-density changes did not contribute significantly to the BOLD fMRI signal under HBO. Analysis of the T2⁎-weighted signal at normobaric air and 1, 2 and 3ATA oxygen in the tissue and the superior sagittal sinus showed a strong dependence on increasing inhaled [O2]. Spontaneous electrophysiological activity and evoked local-field potentials were reduced under HBO. The differences between normobaric air and HBO in basal and evoked electrical activity could not fully account for the strong BOLD responses under HBO. We concluded that activation-induced CBF regulation in the brain does not operate through an oxygen-sensing mechanism and that stimulus-evoked BOLD responses and the venous T2⁎-weighted signals still have room to increase under 3ATA HBO. To our knowledge, this is the first fMRI study under HBO, providing insights into the effects of HBO on neural activity, neurovascular coupling, tissue oxygenation, and the BOLD signal.

Antioxidants and Biomarkers of Oxidative Stress in Preterm Infants with Symptomatic Patent Ductus Arteriosus.

Immature antioxidant and oxygen-sensing mechanisms are involved in the pathogenesis of the patent ductus arteriosus (PDA). We conducted a prospective, observational, pilot study to test the hypothesis that antioxidant activity is low at birth in preterm infants at risk for symptomatic PDA. Blood and urine samples were collected within 24 to 48 hours of life in 53 preterm infants (≤32 weeks' gestation) who developed early PDA symptoms and in 30 term (≥37 weeks' gestation) control infants. Thirty preterm infants developed hemodynamically significant PDA (hsPDA) and required pharmacologic treatment and/or PDA ligation. For these infants, blood and urine samples were also collected at 24 hours posttreatment. Samples were analyzed for biomarkers of antioxidant activity, oxidative stress, and lipid peroxidation. At 24 to 48 hours after birth, plasma superoxide dismutase (SOD), urinary catalase, and plasma and urinary 8-isoPGF2α were significantly lower in preterm infants who developed hsPDA. Plasma 8-isoPGF2α levels rebounded post-PDA treatment, while urinary prostaglandin E2, plasma and urinary thromboxane B2, and plasma SOD declined. The antioxidant status is low in preterm infants at risk for developing hsPDA. SOD may be a key antioxidant regulating functional ductus arteriosus closure. Therefore, low levels may result in persistence of a hsPDA.

A Trihelix DNA Binding Protein Counterbalances Hypoxia-Responsive Transcriptional Activation in Arabidopsis

Author SummaryRespiratory metabolism in land plants requires oxygen availability to be able to generate ATP, which is essential for biosynthetic processes. Cellular hypoxia can be triggered as a consequence of environmental events (mainly floods), anatomical constraints (low tissue permeability to gases), or elevated cellular respiration, and it is unfavorable to growth due to the resultant decline in ATP. The adaptation of plants to fluctuating oxygen levels inside tissues requires the dynamic regulation of mechanisms that ensure cell viability and ultimately organism survival, but only a few molecular components of this homeostatic network are known. Direct hypoxia-sensing entails the posttranslational stabilization of a subgroup of plant ethylene-responsive factor (ERF) transcription factors, which coordinate the expression of hypoxia-inducible genes. Turnover of these ERFs is determined by an oxygen-dependent pathway of proteasomal degradation. Here, we demonstrate that the hypoxia-inducible transcription factor gene HRA1 is transcriptionally activated upon ERF-VII RAP2.12 stabilization and encodes a trihelix DNA binding protein that functionally interacts with RAP2.12 to curtail its activity. In addition to its negative regulation of RAP2.12, HRA1 negatively regulates activation of its own promoter. This RAP2.12-HRA1 control unit allows plants to modulate the extent of the response to hypoxia, including anaerobic enzyme production, to levels that improve endurance of the stress. Our results emphasize the importance of a strategy that can counterbalance energy-inefficient survival responses.

Diversity of Magneto-Aerotactic Behaviors and Oxygen Sensing Mechanisms in Cultured Magnetotactic Bacteria

Microorganisms living in gradient environments affect large-scale processes, including the cycling of elements such as carbon, nitrogen or sulfur, the rates and fate of primary production, and the generation of climatically active gases. Aerotaxis is a common adaptation in organisms living in the oxygen gradients of stratified environments. Magnetotactic bacteria are such gradient-inhabiting organisms that have a specific type of aerotaxis that allows them to compete at the oxic-anoxic interface. They biomineralize magnetosomes, intracellular membrane-coated magnetic nanoparticles, that comprise a permanent magnetic dipole that causes the cells to align along magnetic field lines. The magnetic alignment enables them to efficiently migrate toward an optimal oxygen concentration in microaerobic niches. This phenomenon is known as magneto-aerotaxis. Magneto-aerotaxis has only been characterized in a limited number of available cultured strains. In this work, we characterize the magneto-aerotactic behavior of 12 magnetotactic bacteria with various morphologies, phylogenies, physiologies, and flagellar apparatus. We report six different magneto-aerotactic behaviors that can be described as a combination of three distinct mechanisms, including the reported (di-)polar, axial, and a previously undescribed mechanism we named unipolar. We implement a model suggesting that the three magneto-aerotactic mechanisms are related to distinct oxygen sensing mechanisms that regulate the direction of cells’ motility in an oxygen gradient.

How Does the Heart Sense Changes in Oxygen Tension: A Role for Ion Channels?

Oxygen plays a key role in cellular metabolism and function. Oxygen delivery to cells is crucial, and a lack of oxygen such as that which occurs during myocardial infarction can be lethal. Cells should, therefore, be able to respond to changes in oxygen tension. Since the first studies examining the acute cellular effect of hypoxia on activation of transmitter release from glomus or type I chemoreceptor cells, it is now known that virtually all cells are able to respond to changes in oxygen tension. Despite advances made in characterizing hypoxic responses, the identity of the "oxygen sensor" remains debated. Recently, more evidence has evolved as to how cardiac myocytes sense acute changes in oxygen. This review will examine the available evidence in support of acute oxygen-sensing mechanisms providing a brief historical perspective and then more detailed insights into the heart and the role of cardiac ion channels in hypoxic responses. A further understanding of these cellular processes should result in interventions that assist in preventing the deleterious effects of acute changes in oxygen tension such as alterations in contractile function and cardiac arrhythmia.

Cellular Mechanisms of Oxygen Sensing in Astrocytes

In this study we used fluorescence imaging techniques to characterize the effects of an acute short-term hypoxia on intracellular [Ca2+] signalling in primary cultures of astrocytes from the rat brain stem, midbrain, hippocampus and cortex. It was found that astrocytes respond with strong elevation in intracellular [Ca2+] when PO2 in the incubation media decreases below 15 mmHg. Removal of external Ca2+ had no effect on hypoxia-evoked [Ca2+]i signalling in astrocytes, while application of SERCA inhibitor thapsigargin completely blocked the responses. Cellular mechanism of PO2 sensing involves activation of the PLC pathway as demonstrated in the experiments showing blockade of the hypoxia-induced responses by various inhibitors of PLC downstream processes (U73122, 2-APB or Xestospongine C). Hypoxia-induced [Ca2+]i responses were found to be completely blocked by depolarization of astroglial mitochondria using uncoupler FCCP and, therefore, appear to be dependent on the mitochondrial membrane potential. Ca2+ responses induced in astrocytes by decreases in PO2 lead to fusion of ATP and polyphosphate containing vesicular compartments. These data indicate that astrocytes are sensitive to physiological changes in PO2 and this sensitivity may have a functional significance in the control of local blood flow and the neuronal activity.

The Aryl Hydrocarbon Receptor Nuclear Translocator (ARNT) Family of Proteins: Transcriptional Modifiers with Multi-Functional Protein Interfaces

The basic Helix-Loop-Helix/PER-ARNT-SIM (bHLH-PAS) domain family of transcription factors mediates cellular responses to a variety of internal and external stimuli. As functional transcription factors, these proteins act as bHLH-PAS heterodimers and can be further sub-classified into sensory/activated subunits and regulatory or ARNT-like proteins. This class of proteins act as master regulators of the bHLH-PAS superfamily of transcription factors that mediate circadian rhythm gene programs, innate and adaptive immune responses, oxygen-sensing mechanisms and compensate for deleterious environmental exposures. Some contribute to the etiology of human pathologies including cancer because of their effects on cell growth and metabolism. We will review the canonical roles of ARNT and ARNT-like proteins with an emphasis on coactivator selectivity and recruitment. We will also discuss recent advances in our understanding of noncanonical DNA-binding independent or off-target roles of ARNT that are uncoupled from its classic heterodimeric bHLH-PAS binding partners. Understanding the DNA binding-independent functions of ARNT may identify novel therapeutic options for the treatment of a large spectrum of disease states.

Thiosulfate: a readily accessible source of hydrogen sulfide in oxygen sensing

H2S derived from organic thiol metabolism has been proposed serve as an oxygen sensor in a variety of systems because of its susceptibility to oxidation and its ability to mimic hypoxic responses in numerous oxygen-sensing tissues. Thiosulfate, an intermediate in oxidative H2S metabolism can alternatively be reduced and regenerate H2S. We propose that this contributes to the H2S-mediated oxygen-sensing mechanism. H2S formation from thiosulfate in buffers and in a variety of mammalian tissues and in lamprey dorsal aorta was examined in real time using a polarographic H2S sensor. Inferences of intracellular H2S production were made by examining hypoxic pulmonary vasoconstriction (HPV) in bovine pulmonary arteries under conditions in which increased H2S production would be expected and in mouse and rat aortas, where reducing conditions should mediate vasorelaxation. In Krebs-Henseleit (mammalian) and Cortland (lamprey) buffers, H2S was generated from thiosulfate in the presence of the exogenous reducing agent, DTT, or the endogenous reductant dihydrolipoic acid (DHLA). Both the magnitude and rate of H2S production were greatly increased by these reductants in the presence of tissue, with the most notable effects occurring in the liver. H2S production was only observed when tissues were hypoxic; exposure to room air, or injecting oxygen inhibited H2S production and resulted in net H2S consumption. Both DTT and DHLA augmented HPV, and DHLA dose-dependently relaxed precontracted mouse and rat aortas. These results indicate that thiosulfate can contribute to H2S signaling under hypoxic conditions and that this is not only a ready source of H2S production but also serves as a means of recycling sulfur and thereby conserving biologically relevant thiols.

Abstract 2307: Small GTPase R-Ras promotes normalization of tumor vasculature: a potential oxygen-sensing independent mechanism.

Abstract Newly formed blood vessels in tumors fail to mature into fully functional vessels due to the chronically angiogenic microenvironment. The functional impairment of these vessels hampers drug delivery, thereby diminishing the efficacy of anti-tumor therapies. Excessive vessel permeability associated with tumors also causes clinical complications such as cerebral edema in brain cancer patients. The ability to control vessel maturity in tumors, therefore, provides a potential therapeutic opportunity. We show that R-Ras is essential for the establishment of mature, functional blood vessels in tumors. R-Ras is a small GTPase of the Ras family with antiangiogenic activity. We found that malformation and malfunction of tumor vessels are exacerbated by genetic disruption of R-Ras. R-Ras-deficient vessels exhibited severely impaired pericyte and basement membrane supports, disrupted adherens junctions, and excessive blood leakage in tumors. Tumor cell penetration into the blood circulation increased in R-Ras-deficient mice, supporting a role for R-Ras in controlling tumor vessel permeability. Decreased blood perfusion of tumor vessels in R-Ras-deficient mice elevated tumor hypoxia, which in turn led to decreased efficacy of radiotherapy in these mice. In contrast to R-Ras disruption, R-Ras gain-of-function improved vessel structure and blood perfusion and blocked plasma leakage by enhancing endothelial barrier function and pericyte association with nascent blood vessels. The studies of cell type-specific expression of R-Ras revealed that R-Ras in endothelial cells and in pericytes both contribute individually to the vessel regulation. For instance, endothelial R-Ras limits vascular permeability by inhibiting VE-cadherin internalization induced by VEGF without affecting VE-cadherin gene expression level. These findings indicate that R-Ras is critical for controlling blood circulation and vascular permeability in solid tumors. It has been shown that haplodeficiency of the oxygen-sensor PHD2 leads to normalization of tumor vasculature through stabilization of endothelial HIF-2 and consequent upregulation of soluble VEGFR1 and VE-cadherin gene expression in endothelial cells. Our studies of endothelial cells in hypoxia and normoxia indicate that R-Ras is not involved in oxygen-sensing mechanisms of vessel regulations mediated by the PHD2/HIF-2 pathway. There are important differences between the vessel regulations by PHD2/HIF-2 and by R-Ras. PHD2 haplodeficiency does not affect vessel density, area, or vessel dilation while it normalizes the endothelial barrier and stability. In comparison, R-Ras does normalize all of these vessel parameters while halting angiogenic sprouting. Our observations suggest that an alternative, oxygen sensing-independent vascular normalization phenomenon. Citation Format: Masanobu Komatsu, Junko Sawada, Takeo Urakami, Fangfei Li, Akane Urakami. Small GTPase R-Ras promotes normalization of tumor vasculature: a potential oxygen-sensing independent mechanism. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2307. doi:10.1158/1538-7445.AM2013-2307

Hypoxia and cytoplasmic alkalinization upregulate growth hormone expression in lymphocytes

We report here that culture of lymphoid cells under hypoxic conditions showed an increase in both luciferase expression from a GH-promoter luciferase construct and the levels of lymphocyte GH. The effect was mimicked by treatment of cells with cobalt chloride consistent with a specific oxygen-sensing mechanism. We identified a putative hypoxia response element (HRE) in the GH promoter at the region −176bp to −172bp that contains a copy of the hypoxia-inducible factor-1 (Hif-1) binding motif (5′-ACGTG-3′). The results also showed that culture of primary rat spleen cells with different doses of TMA induced a dose-dependent increase in lymphocyte GH by Western blot analysis. Greater levels of GH are induced in T cell-enriched populations compared to B cell-enriched populations after treatment with CoCl2 or TMA. Our results suggest that the stressful cellular conditions likely to occur at sites of inflammation or tumor growth may induce the synthesis of lymphocyte GH.

Hypoxia-inducible factor prolyl 4-hydroxylases: common and specific roles

Hypoxia-inducible transcription factor (HIF), an αβ dimer, is the key inducer of hypoxia-responsive genes that operate both during normal development and pathological processes in association with decreased oxygen availability. The products of HIF target genes function in, e.g., hematopoiesis, angiogenesis, iron transport, glucose utilization, resistance to oxidative stress, cell proliferation, survival and apoptosis, extracellular matrix homeostasis, and tumorigenesis and metastasis. HIF is accumulated in hypoxia, whereas it is rapidly degraded in normoxic cells. The oxygen-sensing mechanism behind this phenomenon is provided by HIF prolyl 4-hydroxylases (HIF-P4Hs, commonly known as PHDs and EglNs) that require oxygen in their reaction. In normoxia, two prolines in the oxygen-dependent degradation domain of the HIFα subunit become hydroxylated by the HIF-P4Hs. The 4-hydroxyproline residues formed serve as recognition sites for the von Hippel-Lindau E3 ubiquitin ligase complex and result in subsequent ubiquitination and instant proteasomal degradation of HIFα in normoxia. The HIF-P4H reaction is inhibited in hypoxia. HIFα evades degradation and forms a functional dimer with HIFβ, leading to activation of the HIF target genes. The central role of HIF-P4Hs in the regulation of the hypoxia response pathway has provided an attractive possibility as a drug candidate for treatment of, e.g., severe anemias and ischemic conditions, and several companies are currently carrying out clinical studies on the use of HIF-P4H inhibitors to treat anemia in patients with a kidney disease. Therefore, it is important to understand the effects of individual HIF-P4H isoenzymes on the hypoxia response and potential other pathways in vivo. The common and specific functions of the HIF-P4H isoenzymes are discussed in this review on the basis of available data from cell biological studies and gene-modified animals.

Sensing Hypoxia by Mitochondria: A Unifying Hypothesis Involving S-Nitrosation

Sudden hypoxia requires a rapid response in tissues with high energy demand. Mitochondria are rapid sensors for a lack of oxygen, but no consistent mechanism for the sensing process and the subsequent counter-regulation has been described. In the present hypothesis review, we suggest an oxygen-sensing mechanism by mitochondria that is initiated at low oxygen tension by electrons from the respiratory chain, leading to the reduction of intracellular nitrite to nitric oxide ((•)NO) that would subsequently compete with oxygen for binding to cytochrome c oxidase. This allows superoxide ((•)O2(-)) formation in hypoxic areas, leading to S-nitrosation and the inhibition of mitochondrial Krebs cycle enzymes. With more formation of (•)O2(-), peroxynitrite is generated and known to damage the connection between the mitochondrial matrix and the outer membrane. A fundamental question on a regulatory mechanism is its reversibility. Readmission of oxygen and opening of the mitochondrial KATP-channel would allow electrons from glycerol-3-phosphate to selectively reduce the ubiquinone pool to generate (•)O2(-) at both sides of the inner mitochondrial membrane. On the cytosolic side, superoxide is dismutated and will support H2O2/Fe(2+)-dependent transcription processes and on the mitochondrial matrix side, it could lead to the one-electron reduction and reactivation of S-nitrosated proteins. It remains to be elucidated up to which stage the herein proposed silencing of mitochondria remains reversible and when irreversible changes that ultimately lead to classical reperfusion injury are initiated.

Regulation of SREBP during hypoxia requires Ofd1-mediated control of both DNA bindingand degradation

Cells adapt to changes in ambient oxygen by changing their gene expression patterns. In fission yeast, the sterol regulatory element-binding protein Sre1 is proteolytically cleaved under low oxygen, and its N-terminal segment (Sre1N) serves as a hypoxic transcription factor. When oxygen is present, the prolyl hydroxylase Ofd1 down-regulates Sre1N activity in two ways: first, by inhibiting its binding to DNA, and second, by accelerating its degradation. Here we use a mathematical model to assess what each of these two regulatory functions contributes to the hypoxic response of the cell. By disabling individual regulatory functions in the model, which would be difficult in vivo, we found that the Ofd1 function of inhibiting Sre1N binding to DNA is essential for oxygen-dependent Sre1N regulation. The other Ofd1 function of accelerating Sre1N degradation is necessary for the yeast to quickly turn off its hypoxic response when oxygen is restored. In addition, the model predicts that increased Ofd1 production at low oxygen plays an important role in the hypoxic response, and the model indicates that the Ofd1 binding partner Nro1 tunes the response to oxygen. This model quantifies our understanding of a novel oxygen-sensing mechanism that is widely conserved.

Relationships Between Vascular Oxygen Sensing Mechanisms and Hypertensive Disease Processes

Many pathological conditions, like sleep apnea (SA), preeclampsia, high altitude sickness, and chronic obstructive pulmonary disease, that cause intermittent or chronic hypoxia, are often associated with the development of either systemic or pulmonary hypertension.1 Mechanisms sensing hypoxia may be important contributors to the development of pulmonary hypertension and to tissue injury and repair associated with hypertensive disease and in integrated aspects of cardiovascular function. This review will focus on highlighting how some of the better documented oxygen-sensing mechanisms associated with vascular regulation could be influenced by poorly understood interactions between hypoxia and hypertension. It will also consider some of the identified integrated interactions between hypoxia and hypertensive disease processes that could contribute to the progression of disease processes, such as renal hypertension. High altitude and chronic obstructive pulmonary disease are thought to promote pulmonary hypertension by exposing the pulmonary circulation to chronic hypoxia as a result of the low partial pressure of oxygen at high altitude and increases in the diffusion distance for oxygen, respectively.2,3 Although high altitude can initially promote increased systemic blood pressure, this change appears to reverse with time.4 Interestingly, hypobaric hypoxia conditions of high altitude were observed to prevent the development of hypertension and skeletal muscle arteriolar rarefaction in spontaneously hypertensive rats.5 Living at high altitude seems to lower blood pressure only in children,6 and it did not appear to attenuate hypertension in obese adults consuming high-fat diets.7 Based on studies in animal models of acute and chronic hypoxia, hypoxic pulmonary vasoconstriction (HPV) is initially activated. Eventually endothelial dysfunction associated with a loss of NO and increased reactive oxygen species (ROS), endothelin, serotonin, and contractile prostaglandins contribute to maintaining elevated pulmonary artery pressure through increased intracellular Ca2+ levels and rho kinase-associated myofilament Ca2+ sensitivity.8 Although …

Perspective in Chronic Kidney Disease: Targeting Hypoxia-Inducible Factor (HIF) as Potential Therapeutic Approach

Tissue hypoxia is a pathologic feature of many human diseases like cancer, myocardial infarction, stroke, and kidney disease. Convincing data from clinical studies in patients with chronic renal failure point to chronic hypoxia of kidneys as the end result of multiple processes and mechanisms. In acute as well as chronic diseases, tissue hypoxia not only implies a risk of energy deprivation but also induces regulatory mechanisms with profound influence on gene expression. Moreover, once established, accumulating evidence points to this chronic hypoxia as the central player along with final common pathway to end-stage renal disease (ESRD). An evolutionarily preserved oxygen-sensing mechanism enables cells to adapt and maintain homeostasis under hypoxic conditions by transcriptional activation of a host of genes mediating metabolic adaptation, angiogenesis, energy conservation, erythropoiesis, in addition to cell survival. The endogenous oxygen-sensing mechanism incorporates hypoxia-inducible factors (HIFs) that hub cellular response to hypoxia and comprises a family of oxygen-sensitive basic helix–loop–helix proteins that control the cellular transcriptional response to hypoxia. Hypoxia-inducible factor 1 (HIF-1) is thus a significant mediator of physiological responses to acute and chronic hypoxia. Since HIF is activated to suboptimal levels in pathogenic renal states, therapeutic activation holds a promising novel and effective approach to the treatment of ESRD. Current insights into the regulation of HIF may augment the understanding of the role of hypoxia in renal failure progression and may unbolt new options to improve hypoxia tolerance and induce nephroprotection.

Dysregulated Oxygen Metabolism of the Kidney by Uremic Toxins: Review

Because kidneys consume a large amount of oxygen and are relatively inefficient in oxygen uptake, they are susceptible to hypoxia, especially in patients with advanced chronic kidney disease accompanied by loss of peritubular capillaries. Accumulating evidence suggests that chronic tubulointerstitial hypoxia acts as a final common pathway leading to end-stage renal disease. Some biologically active uremic retention molecules, considered as uremic toxins, accumulate as the renal function declines, and at this moment, more than 90 bioactive uremic toxins have been identified. Uremic toxins per se have been proven to accelerate the progression of renal failure. However, the causal relationship between uremic toxin and tubulointerstitial hypoxia remains unclear. Our studies provided direct evidence that uremic toxin dysregulates oxygen metabolism in the kidney. Indoxyl sulfate (IS), a representative protein-bound uremic toxin, increased oxygen consumption in proximal renal tubules, decreased renal oxygenation, and consequently aggravated hypoxia in the remnant rat kidneys. The increase in tubular oxygen consumption by IS was dependent on sodium-potassium adenosine triphosphatase and oxidative stress. Our work also indicated a possible connection between IS and the desensitization of the oxygen-sensing mechanism in erythropoietin-producing cells, which may partly explain inadequate erythropoietin production in hypoxic kidneys of end-stage renal disease patients. Studies of uremic toxins will open a new avenue in development of novel therapeutic approaches of kidney disease.

Plant Oxygen Sensing Is Mediated by the N-End Rule Pathway: A Milestone in Plant Anaerobiosis

Like all aerobic organisms, plants require molecular oxygen for respiratory energy production. In plants, hypoxic conditions can occur during natural events (e.g., flooding), during developmental processes (e.g., seed germination), and in cells of compact tissues with high metabolic rates. Plant acclimation responses to hypoxia involve a modulation of gene expression leading to various biochemical, physiological, and morphological changes that stave off eventual anoxia. In contrast with the animal kingdom, a direct oxygen-sensing mechanism in plants has been elusive so far. However, two recent independent studies show that oxygen sensing in plants operates via posttranslational regulation of key hypoxia response transcription factors by the N-end rule pathway. The N-end rule is an evolutionarily conserved pathway for protein degradation that relates the fate of a protein with the identity of its N-terminal residues. Results from these studies demonstrate that oxygen-dependent modification and targeted proteolysis of members of the ethylene response factor group VII transcription factor family regulate hypoxia-responsive gene expression in Arabidopsis thaliana. The discovery of this plant hypoxia-sensing mechanism sets the stage for further research on plant homeostatic response to oxygen, which could be relevant to understanding plant distributions in flood-prone ecosystems and improving hypoxia tolerance of crops.

Indoxyl sulfate, a representative uremic toxin, suppresses erythropoietin production in a HIF-dependent manner

Advanced chronic kidney disease (CKD) patients encounter anemia through insufficient erythropoietin (EPO) production by peritubular fibroblasts. Recent studies showed an increase in EPO production by pharmacological activation of hypoxia-inducible transcription factors (HIFs) in dialysis patients, suggesting that desensitization of the oxygen-sensing mechanism is responsible for the development of renal anemia. Our recent work demonstrated that indoxyl sulfate (IS), a uremic toxin, dysregulates oxygen metabolism in tubular cells. Here we provide evidence of an additional property that IS impairs oxygen sensing in EPO-producing cells. HepG2 cells were stimulated with cobalt chloride (CoCl2) or hypoxia under varying concentrations of IS. EPO mRNA was evaluated by quantitative PCR. Nuclear accumulation of HIF-α was evaluated by western blotting. Transcriptional activity of HIF was checked by hypoxia-responsive element (HRE)-luciferase reporter assay. The impact of IS was further evaluated in vivo by administering rats with indole, a metabolic precursor of IS, and subjecting them to CoCl2 stimulation, in which renal EPO mRNA as well as plasma EPO levels were measured by quantitative PCR and enzyme-linked immunosorbent assay, respectively. Although IS induced cellular toxicity at relatively high concentrations (⩾2.5 mM), EPO mRNA expression was significantly suppressed by IS at concentrations below cytotoxic ranges. In HepG2 cells, IS treatment decreased nuclear accumulation of HIF-α proteins and suppressed HRE-luciferase activity following hypoxia. Furthermore, administration of rats with indole suppressed renal EPO mRNA expression and plasma EPO levels, corroborating in vitro findings. Results of the present study provide a possible connection between a uremic toxin and the desensitization of the oxygen-sensing mechanism in EPO-producing cells, which may partly explain inadequate EPO production in hypoxic kidneys of CKD patients.

Oxygen sensing in plants is mediated by an N-end rule pathway for protein destabilization

The majority of eukaryotic organisms rely on molecular oxygen for respiratory energy production. When the supply of oxygen is compromised, a variety of acclimation responses are activated to reduce the detrimental effects of energy depletion. Various oxygen-sensing mechanisms have been described that are thought to trigger these responses, but they each seem to be kingdom specific and no sensing mechanism has been identified in plants until now. Here we show that one branch of the ubiquitin-dependent N-end rule pathway for protein degradation, which is active in both mammals and plants, functions as an oxygen-sensing mechanism in Arabidopsis thaliana. We identified a conserved amino-terminal amino acid sequence of the ethylene response factor (ERF)-transcription factor RAP2.12 to be dedicated to an oxygen-dependent sequence of post-translational modifications, which ultimately lead to degradation of RAP2.12 under aerobic conditions. When the oxygen concentration is low-as during flooding-RAP2.12 is released from the plasma membrane and accumulates in the nucleus to activate gene expression for hypoxia acclimation. Our discovery of an oxygen-sensing mechanism opens up new possibilities for improving flooding tolerance in crops.

The HIF Pathway and Erythrocytosis

Because of the central role that red blood cells play in the delivery of oxygen to tissues of the body, red blood cell mass must be controlled at precise levels. The glycoprotein hormone erythropoietin (EPO) regulates red blood cell mass. EPO transcription, in turn, is regulated by a distinctive oxygen-sensing mechanism. In this pathway, prolyl hydroxylase domain protein (PHD) site-specifically hydroxylates the α-subunit of the transcription factor hypoxia-inducible factor α (HIF-α), thereby targeting the latter for degradation by the von Hippel-Lindau tumor-suppressor protein (VHL). Under hypoxic conditions, this posttranslational modification of HIF-α is inhibited, which stabilizes it and promotes the transcriptional activation of genes, including that for EPO. Rare patients with erythrocytosis have mutations in the genes encoding for PHD2, HIF-2α, and VHL, which implicates these proteins as critical to the proper control of red blood cell mass in humans.