The Keap1-Nrf2 system plays pivotal roles in defense mechanisms by regulating cellular redox homeostasis. Nrf2 is an inducible transcription factor that activates a battery of genes encoding ...antioxidant proteins and phase II enzymes in response to oxidative stress and electrophilic xenobiotics. The activity of Nrf2 is regulated by Keap1, which promotes the ubiquitination and subsequent degradation of Nrf2 under normal conditions and releases the inhibited Nrf2 activity upon exposure to the stresses. Though an impressive contribution of the Keap1-Nrf2 system to the protection from exogenous and endogenous electrophilic insults has been well established, a line of evidence has suggested that the Keap1-Nrf2 system has various novel functions, particularly in cell proliferation and differentiation. Because the proliferation and differentiation of diverse cell types are often influenced and modulated by the cellular redox balance, Nrf2 has been considered to control these cellular processes by regulating the cellular levels of reactive oxygen species (ROS). In addition, analyses of the genome-wide distribution of Nrf2 have identified new sets of Nrf2 target genes whose products are involved in cell proliferation and differentiation but not necessarily in the regulation of oxidative stress. Considering the most characteristic features of Nrf2 as an inducible transcription factor, a newly emerged concept proposes that the Keap1-Nrf2 system translates environmental stresses into regulatory network signals in cell fate determination. In this review, we introduce the contribution of Nrf2 to lineage-specific differentiation, maintenance and differentiation of stem cells, and proliferation of normal and cancer cells, and we discuss how the response to fluctuating environments modulates cell behavior through the Keap1-Nrf2 system.
•Keap1–Nrf2 system plays a key role in the regulation of redox homeostasis.•Nrf2 also regulates cell proliferation and differentiation.•Keap1–Nrf2 system translates oxidative stress into cell fate determination signals.
•The irregular strip packing problem of rasterized shapes in high-resolution•A double scanline representation for computing coordinate penetration depths•Coordinate descent heuristics based on a line ...search algorithm with a corner detection•Computational results on irregular shapes including some curve segments and holes
We consider the irregular strip packing problem of rasterized shapes, where a given set of pieces of irregular shapes represented in pixels should be placed into a rectangular container without overlap. The rasterized shapes provide simple procedures of the intersection test without any exceptional handling due to geometric issues, while they often require much memory and computational effort in high-resolution. To reduce the complexity of rasterized shapes, we propose a pair of scanlines representation called the double scanline representation that merges consecutive pixels in each row and column into strips with unit width, respectively. Based on this, we develop coordinate descent heuristics for the raster model that repeat a line search in the horizontal and vertical directions alternately, where we also introduce a corner detection technique used in computer vision to reduce the search space. Computational results for test instances show that the proposed algorithm obtains sufficiently dense layouts of rasterized shapes in high-resolution within a reasonable computation time.
Abstract
Oxidative stress is one of the major causes of the age-related functional decline in cells and tissues. The KEAP1–NRF2 system plays a central role in the regulation of redox balance, and ...NRF2 activation exerts antiageing effects by controlling oxidative stress in aged tissues. α-Klotho was identified as an ageing suppressor protein based on the premature ageing phenotypes of its mutant mice, and its expression is known to gradually decrease during ageing. Because α-klotho has been shown to possess antioxidant function, ageing-related phenotypes of α-klotho mutant mice seem to be attributable to increased oxidative stress at least in part. To examine whether NRF2 activation antagonizes ageing-related phenotypes caused by α-klotho deficiency, we crossed α-klotho–deficient (Kl−/−) mice with a Keap1-knockdown background, in which the NRF2 pathway is constitutively activated in the whole body. NRF2 pathway activation in Kl−/− mice extended the lifespan and dramatically improved ageing-related renal phenotypes. With elevated expression of antioxidant genes accompanied by an oxidative stress decrease, the antioxidant effects of NRF2 seem to make a major contribution to the attenuation of ageing-related renal phenotypes of Kl−/− mice. Thus, NRF2 is expected to exert an antiageing function by partly compensating for the functional decline of α-Klotho during physiological ageing.
Graphical Abstract
Graphical Abstract
NRF2 (nuclear factor erythroid 2-related factor 2) is a key transcriptional activator that mediates the inducible expression of antioxidant genes. NRF2 is normally ubiquitinated by KEAP1 (Kelch-like ...ECH-associated protein 1) and subsequently degraded by proteasomes. Inactivation of KEAP1 by oxidative stress or electrophilic chemicals allows NRF2 to activate transcription through binding to antioxidant response elements (AREs) and recruiting histone acetyltransferase CBP (CREB-binding protein). Whereas KEAP1-dependent regulation is a major determinant of NRF2 activity, NRF2-mediated transcriptional activation varies from context to context, suggesting that other intracellular signaling cascades may impact NRF2 function. To identify a signaling pathway that modifies NRF2 activity, we immunoprecipitated endogenous NRF2 and its interacting proteins from mouse liver and identified glucocorticoid receptor (GR) as a novel NRF2-binding partner. We found that glucocorticoids, dexamethasone and betamethasone, antagonize diethyl maleate-induced activation of NRF2 target genes in a GR-dependent manner. Dexamethasone treatment enhanced GR recruitment to AREs without affecting chromatin binding of NRF2, resulting in the inhibition of CBP recruitment and histone acetylation at AREs. This repressive effect was canceled by the addition of histone deacetylase inhibitors. Thus, GR signaling decreases NRF2 transcriptional activation through reducing the NRF2-dependent histone acetylation. Consistent with these observations, GR signaling blocked NRF2-mediated cytoprotection from oxidative stress. This study suggests that an impaired antioxidant response by NRF2 and a resulting decrease in cellular antioxidant capacity account for the side effects of glucocorticoids, providing a novel viewpoint for the pathogenesis of hypercorticosteroidism.
Tissue stem cells are maintained in quiescence under physiological conditions but proliferate and differentiate to replenish mature cells under stressed conditions. The KEAP1-NRF2 system plays an ...essential role in stress response and cytoprotection against redox disturbance. To clarify the role of the KEAP1-NRF2 system in tissue stem cells, we focused on hematopoiesis in this study and used Keap1-deficient mice to examine the effects of persistent activation of NRF2 on long-term hematopoietic stem cells (LT-HSCs). We found that persistent activation of NRF2 due to Keap1 deficiency did not change the number of LT-HSCs but reduced their quiescence in steady-state hematopoiesis. During hematopoietic regeneration after bone marrow (BM) transplantation, persistent activation of NRF2 reduced the BM reconstitution capacity of LT-HSCs, suggesting that NRF2 reduces the quiescence of LT-HSCs and promotes their differentiation, leading to eventual exhaustion. Transient activation of NRF2 by an electrophilic reagent also promotes the entry of LT-HSCs into the cell cycle. Taken together, our findings show that NRF2 drives the cell cycle entry and differentiation of LT-HSCs at the expense of their quiescence and maintenance, an effect that appears to be beneficial for prompt recovery from blood loss. We propose that the appropriate control of NRF2 activity by KEAP1 is essential for maintaining HSCs and guarantees their stress-induced regenerative response.
Cells are often exposed to exogenous and endogenous redox disturbances and exert their protective mechanisms in response to stimuli. The KEAP1-NRF2 system plays pivotal roles in counteracting ...oxidative damage. Due to the transient nature of NRF2 activation, the identification of cells in which NRF2 is activated in response to systemic stimuli is sometimes not easy. To examine the electrophilic stress response at a single-cell resolution, we aimed to develop a new reporter mouse in this study. A cell-tracing strategy exploiting Cre recombinase-mediated activation of a reporter gene was chosen for stable detection of reporter expression instead of real-time monitoring of the cellular response. We established a transgenic mouse line expressing the Neh2-Cre recombinase fusion protein. As Neh2 is an amino-terminal domain of NRF2 that serves as a degron and mediates KEAP1-dependent degradation and electrophile-inducible stabilization, Neh2-Cre was expected to be activated in response to electrophiles. The Neh2-Cre transgenic mouse was crossed with the ROSA26-loxP-stop-loxP-tdTomato reporter mouse (ROSA-LSL-tdTomato mouse). The compound mutant reporter mice exhibited accumulation of tdTomato-positive cells in various organs after repeated administration of CDDO-Im, one of the NRF2-inducing electrophiles. The mice were also successfully used for the detection of cells that experienced a cisplatin-induced electrophilic stress response.
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•Stabilization of Neh2-Cre fusion protein is regulated by KEAP1.•Transgenic mice expressing Neh2-Cre were generated.•Neh2-Cre:tdTomato mouse was used to monitor the exposure history to electrophiles.•Repetitive CDDO-Im treatment induces tdTomato expression in various tissues.•Repetitive cisplatin treatment induces tdTomato expression in inner ear and kidney.
Noise-induced hearing loss (NIHL) is one of the most common sensorineural hearing deficits. Recent studies have demonstrated that the pathogenesis of NIHL is closely related to ischemia-reperfusion ...injury of cochlea, which is caused by blood flow decrease and free radical production due to excessive noise. This suggests that protecting the cochlea from oxidative stress is an effective therapeutic approach for NIHL. NRF2 is a transcriptional activator playing an essential role in the defense mechanism against oxidative stress. To clarify the contribution of NRF2 to cochlear protection, we examined Nrf2(-/-) mice for susceptibility to NIHL. Threshold shifts of the auditory brainstem response at 7 days post-exposure were significantly larger in Nrf2(-/-) mice than wild-type mice. Treatment with CDDO-Im, a potent NRF2-activating drug, before but not after the noise exposure preserved the integrity of hair cells and improved post-exposure hearing levels in wild-type mice, but not in Nrf2(-/-) mice. Therefore, NRF2 activation is effective for NIHL prevention. Consistently, a human NRF2 SNP was significantly associated with impaired sensorineural hearing levels in a cohort subjected to occupational noise exposure. Thus, high NRF2 activity is advantageous for cochlear protection from noise-induced injury, and NRF2 is a promising target for NIHL prevention.
NF-E2-related factor 2 (NRF2) plays a crucial role in the maintenance of cellular homeostasis by regulating various enzymes and proteins that are involved in the redox reactions utilizing sulfur. ...While substantial impacts of NRF2 on mitochondrial activity have been described, the precise mechanism by which NRF2 regulates mitochondrial function is still not fully understood. Here, we demonstrated that NRF2 increased intracellular persulfides by upregulating the cystine transporter xCT encoded by Slc7a11, a well-known NRF2 target gene. Persulfides have been shown to play an important role in mitochondrial function. Supplementation with glutathione trisulfide (GSSSG), which is a form of persulfide, elevated the mitochondrial membrane potential (MMP), increased the oxygen consumption rate (OCR) and promoted ATP production. Persulfide-mediated mitochondrial activation was shown to require the mitochondrial sulfur oxidation pathway, especially sulfide quinone oxidoreductase (SQOR). Consistently, NRF2-mediated mitochondrial activation was also dependent on SQOR activity. This study clarified that the facilitation of persulfide production and sulfur metabolism in mitochondria by increasing cysteine availability is one of the mechanisms for NRF2-dependent mitochondrial activation.
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•Cystine restriction reduces persulfides and inhibits mitochondrial activity.•Persulfide supplementation promotes mitochondrial activity.•NRF2 activation increases persulfides and promotes mitochondrial activity.•Upregulation of xCT induced by NRF2 activation results in perusulfide increase.•NRF2-mediated mitochondrial activation relies on persulfide metabolism.
Nucleomethylin (NML) has been shown to contribute to ribosome formation through regulating transcription and post-transcriptional modification of rRNA. Based on the observation that NML-/- mice are ...frequently embryonic lethal, we analyzed NML-/- embryos to clarify the role of NML in embryogenesis. We found that NML deficiency leads to lethality at the time point between E10.5 and E12.5. Most of E10.5 NML-/- embryos exhibited growth retardation and/or malformation with marked impairment of erythropoiesis. Consistent with a previous study, the m1A in 28S rRNA was dramatically reduced in NML-/- foetal liver (FL) cells. Because the previous study demonstrated p53-dependent apoptosis of NML-knockdown cells, and because we observed upregulation of p21, one of the p53 target genes, in NML-/- FL cells, we tested whether p53 disruption cancelled the NML-deficient phenotypes. Contrary to our expectation, suppression of p53 did not rescue the lethality or impaired erythropoiesis of NML-/- embryos, suggesting that p53-independent mechanisms underlie the NML-deficient phenotypes. These results clarify an essential role of NML during embryogenesis, particularly in erythropoiesis. We surmise that embryonic erythropoiesis is particularly sensitive to impaired protein synthesis, which is caused by the defective methylation of rRNA and consequent failure of ribosome formation.
Nrf2 is a major transcriptional activator of cytoprotective genes against oxidative/electrophilic stress, and Keap1 negatively regulates Nrf2. Emerging works have also suggested a role for Nrf2 as a ...regulator of differentiation in various cells, but the contribution of Nrf2 to the differentiation of hematopoietic stem cells (HSCs) remains elusive. Clarifying this point is important to understand Nrf2 functions in the development and/or resolution of inflammation. Here, we established two transgenic reporter mouse lines that allowed us to examine Nrf2 expression precisely in HSCs. Nrf2 was abundantly transcribed in HSCs, but its activity was maintained at low levels due to the Keap1‐mediated degradation of Nrf2 protein. When we characterized Keap1‐deficient mice, their bone marrow cells showed enhanced granulocyte‐monocyte differentiation at the expense of erythroid and lymphoid differentiation. Importantly, Keap1‐null HSCs showed lower expression of erythroid and lymphoid genes than did control HSCs, suggesting granulocyte‐monocyte lineage priming in Keap1‐null HSCs. This abnormal lineage commitment was restored by a concomitant deletion of Nrf2, demonstrating the Nrf2‐dependency of the skewing. Analysis of Nrf2‐deficient mice revealed that the physiological level of Nrf2 is sufficient to contribute to the lineage commitment. This study unequivocally shows that the Keap1‐Nrf2 system regulates the cell fate determination of HSCs.
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