Characterization of Common Minke Whale (Balaenoptera Acutorostrata) Cell Lines Immortalized with the Expression of Cell Cycle Regulators.

Primary cultured cells cannot proliferate infinite. The overcoming of this limit can be classified as immortalization. Bypass of p16 senescence protein induces efficient immortalization various types of mammalians is previously reported. However, the Cetacea species is not known. Here, that common minke whale-derived cells can be immortalized with a combination of human genes, mutant cyclin-dependent kinase 4 (CDK4R24C ), cyclin D1, and Telomerase Reverse Transcriptase (TERT) is reported. These results indicate that the function of cell cycle regulators in premature senescence is evolutionarily conserved. This study describes the conserved roles of cell cycle regulators in the immortalization of cells from humans to Cetacea species. Furthermore, using RNA-seq based on next-generation sequencing, the gene expression profiles of immortalized cells are compared with parental cells as well as those immortalized with SV40 large T antigen, which is once a popular method for cellular immortalization. The profiling results show that newly established common minke-whale-derived immortaliozed cells have completely different profiles from SV40 cells. This result indicates that the expression of mutant CDK4, cyclin D1, and TERT enables to establish immortalized cell lines with different biological nature from SV40 expressing cells.

Sheep-derived cell immortalization through the expression of cell cycle regulators and biological characterization using transcriptomes.

Sheep are important domestic animals for the production of wool and meat. Although numerous cultured cell lines from humans and mice have been established, the number of cell lines derived from sheep is limited. To overcome this issue, the efficient establishment of a sheep-derived cell line and its biological characterization is reported. Mutant cyclin-dependent kinase 4, cyclin D1, and telomerase reverse transcriptase were introduced into sheep muscle-derived cells in an attempt to immortalize primary cells using the K4DT method. Furthermore, the SV40 large T oncogene was introduced into the cells. The successful immortalization of sheep muscle-derived fibroblasts was shown using the K4DT method or SV40 large T antigen. Furthermore, the expression profile of established cells showed close biological characteristics of ear-derived fibroblasts. This study provides a useful cellular resource for veterinary medicine and cell biology.

Properties of a Single Amino Acid Residue in the Third Transmembrane Domain Determine the Kinetics of Ambient Light-Sensitive Channelrhodopsin.

Channelrhodopsins have been utilized in gene therapy to restore vision in patients with retinitis pigmentosa and their channel kinetics are an important factor to consider in such applications. We investigated the channel kinetics of ComV1 variants with different amino acid residues at the 172nd position. Patch clamp methods were used to record the photocurrents induced by stimuli from diodes in HEK293 cells transfected with plasmid vectors. The channel kinetics (τon and τoff) were considerably altered by the replacement of the 172nd amino acid and was dependent on the amino acid characteristics. The size of amino acids at this position correlated with τon and decay, whereas the solubility correlated with τon and τoff. Molecular dynamic simulation indicated that the ion tunnel constructed by H172, E121, and R306 widened due to H172A variant, whereas the interaction between A172 and the surrounding amino acids weakened compared with H172. The bottleneck radius of the ion gate constructed with the 172nd amino acid affected the photocurrent and channel kinetics. The 172nd amino acid in ComV1 is a key residue for determining channel kinetics as its properties alter the radius of the ion gate. Our findings can be used to improve the channel kinetics of channelrhodopsins.

Comprehensive transcriptome data to identify downstream genes of testosterone signalling in dermal papilla cells.

Testosterone-related steroid hormones are associated with various types of diseases, including prostate cancer and androgenetic alopecia (AGA). The testosterone or dihydroxy testosterone (DHT) circulates through the blood, binds to the androgen receptor (AR) in the cytoplasm, and finally enters the nucleus to activate downstream target genes. We previously found that immortalized dermal papilla cells (DPCs) lost AR expression, which may be explained by the repeated cell passages of DPCs. To compensate for the AR expression, DPCs that express AR exogenously were established. In this study, we performed an RNA-Seq analysis of the AR-expressing and non-AR-expressing DPCs in the presence or absence of DHT to identify the downstream target genes regulated by AR signalling. Furthermore, we treated DPCs with minoxidil sulphate, which has the potential to treat AGA. This is the first comprehensive analysis to identify the downstream genes involved in testosterone signalling in DPCs. Our manuscript provides high-priority data for the discovery of molecular targets for prostate cancer and AGA.

SIG-1451, a Novel, Non-Steroidal Anti-Inflammatory Compound, Attenuates Light-Induced Photoreceptor Degeneration by Affecting the Inflammatory Process.

Age-related macular degeneration is a progressive retinal disease that is associated with factors such as oxidative stress and inflammation. In this study, we evaluated the protective effects of SIG-1451, a non-steroidal anti-inflammatory compound developed for treating atopic dermatitis and known to inhibit Toll-like receptor 4, in light-induced photoreceptor degeneration. SIG-1451 was intraperitoneally injected into rats once per day before exposure to 1000 lx light for 24 h; one day later, optical coherence tomography showed a decrease in retinal thickness, and electroretinogram (ERG) amplitude was also found to have decreased 3 d after light exposure. Moreover, SIG-1451 partially protected against this decrease in retinal thickness and increase in ERG amplitude. One day after light exposure, upregulation of inflammatory response-related genes was observed, and SIG-1451 was found to inhibit this upregulation. Iba-1, a microglial marker, was suppressed in SIG-1451-injected rats. To investigate the molecular mechanism underlying these effects, we used lipopolysaccharide (LPS)-stimulated rat immortalised Müller cells. The upregulation of C-C motif chemokine 2 by LPS stimulation was significantly inhibited by SIG-1451 treatment, and Western blot analysis revealed a decrease in phosphorylated I-κB levels. These results indicate that SIG-1451 indirectly protects photoreceptor cells by attenuating light damage progression, by affecting the inflammatory responses.

Lentiviral expression of calpain-1 C2-like domain peptide prevents glutamate-induced cell death in mouse hippocampal neuronal HT22 cells.

Glutamate neurotoxicity is involved in neurodegenerative diseases, including Alzheimer's and Parkinson's diseases. Excess glutamate causes caspase-independent programmed cell death via oxidative stress and calcium influx. Our previous study showed that calpain-1 localizes to both the cytoplasm and mitochondria, where apoptosis-inducing factor (AIF) is cleaved by calpain-1 and translocates to the nucleus to induce DNA fragmentation. The autoinhibitory region of calpain-1 conjugated with the cell-penetrating peptide HIV1-Tat (namely Tat-µCL) specifically prevents the activity of mitochondrial calpain-1 and attenuates neuronal cell death in animal models of retinitis pigmentosa, as well as glutamate-induced cell death in mouse hippocampal HT22 cells. In the present study, we constructed a lentiviral vector expressing the Tat-µCL peptide and evaluated its protective effect against glutamate-induced cell death in HT22 cells. Lentiviral transduction with Tat-µCL significantly suppressed glutamate-induced nuclear translocation of AIF and DNA fragmentation. The findings of the present study suggest that the stable expression of Tat-µCL may be a potential gene therapy modality for neurodegenerative diseases.

Mitochondrial localization of calpain-13 in mouse brain.

Calpains are Ca2+-dependent cysteine proteases involved in various intercellular physiological functions. Although most calpains exist in the cytosol, four isoforms of calpain (calpains-1, -2, -5, -10) are also localized in the mitochondria. In the present study, we examined the mitochondrial localization of calpain-13, as a novel mitochondrial calpain, in C57BL/6J mice. The tissue distribution and mitochondrial subfractionation of calpain-13 were investigated using western blotting. Calpain-13 was present in both the mitochondrial membrane (outer membrane and inner membrane) and soluble (intermembrane space and matrix) fractions. Through immunohistochemistry, calpain-13 was found to be expressed in the cerebral cortex and hippocampus of the mouse brain. We further confirmed the localization of calpain-13 in the mitochondria of the mouse brain using immunoelectron microscopy. Our present study thus revealed that calpain-13 is localized in the mitochondria, in addition to the cytosol, in the mouse brain. Future studies investigating the enzymatic properties and physiological functions of both cytosolic and mitochondrial calpain-13 will shed light on the potential involvement of calpain-13 in neurodegenerative diseases including Parkinson's disease and Alzheimer's disease.

Mitochondrial calpain-5 truncates caspase-4 during endoplasmic reticulum stress.

Calpains are cysteine proteases activated in response to intracellular calcium signaling. Activated calpains regulate various cellular functions by degrading substrate molecules in a site-specific manner. Although most calpains are localized in the cytosol, we previously reported that calpain-5 exists in the mitochondria. The mitochondrial calpain-5 is activated during endoplasmic reticulum (ER) stress. However, the substrate of calpain-5, as well as the physiological significance of calpain-5 activation, has not yet been elucidated. In the present study, we treated HeLa cells with A23187, tunicamycin, or hydrogen peroxide to induce intracellular calcium increase, resulting in cell death. The cells treated with A23187 or tunicamycin exhibited the activation of calpain-5 and truncation of caspase-4. The truncation of caspase-4 was inhibited by the repression of calpain-5 expression with the appropriate siRNA. Additionally, both calpain-5 and caspase-4 were observed in the mitochondria. Our study is the first to demonstrate that the activation of mitochondrial calpain-5 triggers the truncation of caspase-4, suggesting that mitochondrial calpain-5 regulates the downstream pathway of caspase-4, including cell death and the inflammatory cascade. The results of the present study provide new insights into ER-stress-related diseases such as Alzheimer's disease and cancer. These perspectives allow us to propose new therapeutic strategies such as the development of inhibitors or activators of calpain-5, which may be useful in the development of treatment for ER-stress-related diseases.

Detailed chromosome analysis of wild-type, immortalized fibroblasts with SV40T, E6E7, combinational introduction of cyclin dependent kinase 4, cyclin D1, telomerase reverse transcriptase.

Cell immortalization enables us to expand the cultured cell infinitely. However, the process of immortalization sometimes changes the nature of the original cell. In this study, we established immortalized embryonic fibroblasts with oncogenic SV40T and human papilla virus-derived E6E7, combinational expression of mutant cyclin-dependent kinase 4 (CDK4), cyclin D1, and telomerase reverse transcriptase (TERT) from identical primary wild-type human embryonic fibroblasts (HE16). After the establishment of immortalized cells, we compared the details of chromosome condition with the G-banding and Q-banding methods. There is no example of detailed analysis so far about chromosome abnormalities, such as trisomy, ring chromosome, reciprocal translocation, and dicentric chromosomes. The detailed chromosome analysis revealed that immortalized cells with SV40T and E6E7 showed intensive chromosome abnormalities, such as gain or loss of the chromosomes all through the genome. Furthermore, we detected that the incidence of chromosome abnormities in the immortalized cell with the combinational introduction of R24C mutant of CDK4, cyclin D1, and TERT is almost identical to that of wild-type cell. Furthermore, short tandem repeat analysis demonstrated that the origin of K4DT cell is primary HE16. These results showed that cellular immortalization with CDK4, cyclin D1, and TERT is more advantageous in keeping the chromosome's original condition than oncogenic immortalization methods.

Calpain-1 C2L domain peptide protects mouse hippocampus-derived neuronal HT22 cells against glutamate-induced oxytosis.

Calpains are Ca2+-dependent cysteine proteases; their aberrant activation is associated with several neurodegenerative diseases. The µ-calpain catalytic subunit, calpain-1, is located in the cytoplasm as well as in the mitochondria. Mitochondrial calpain-1 cleaves apoptosis-inducing factor (AIF), leading to apoptotic cell death. We have previously reported that short peptides of calpain-1 C2-like domain conjugated with cell penetrating peptide HIV-Tat (Tat-µCL) selectively inhibit mitochondrial calpain-1 and effectively prevent neurodegenerative diseases of the eye. In this study, we determined whether mitochondrial calpain-1 mediates oxytosis (oxidative glutamate toxicity) in hippocampal HT22 cells using Tat-µCL and newly generated polyhistidine-conjugated µCL peptide and compared their efficacies in preventing oxytosis. TUNEL assay and single strand DNA staining revealed that both µCL peptides inhibited glutamate-induced oxytosis. Additionally, both the peptides suppressed the mitochondrial AIF translocation into the nucleus. All polyhistidine-µCL peptides (containing 4-16 histidine residues) showed higher cell permeability than Tat-µCL. Notably, tetrahistidine (H4)-µCL exerted the highest cytoprotective activity. Thus, H4-µCL may be a potential peptide drug for calpain-1-mediated neurodegenerative diseases such as Alzheimer's disease.

Phototoxicities Caused by Continuous Light Exposure Were Not Induced in Retinal Ganglion Cells Transduced by an Optogenetic Gene.

The death of photoreceptor cells is induced by continuous light exposure. However, it is unclear whether light damage was induced in retinal ganglion cells with photosensitivity by transduction of optogenetic genes. In this study, we evaluated the phototoxicities of continuous light exposure on retinal ganglion cells after transduction of the optogenetic gene mVChR1 using an adeno-associated virus vector. Rats were exposed to continuous light for a week, and visually evoked potentials (VEPs) were recorded. The intensities of continuous light (500, 1000, 3000, and 5000 lx) increased substantially after VEP recordings. After the final recording of VEPs, retinal ganglion cells (RGCs) were retrogradely labeled with a fluorescein tracer, FluoroGold, and the number of retinal ganglion cells was counted under a fluorescent microscope. There was no significant reduction in the amplitudes of VEPs and the number of RGCs after exposure to any light intensity. These results indicated that RGCs were photosensitive after the transduction of optogenetic genes and did not induce any phototoxicity by continuous light exposure.

Characterization of mitochondrial calpain-5.

Calpain, a Ca2+-dependent cysteine protease, plays a significant role in gene expression, signal transduction, and apoptosis. Mutations in human calpain-5 cause autosomal dominant neovascular inflammatory vitreoretinopathy and the inhibition of calpain-5 activity may constitute an effective therapeutic strategy for this condition. Although calpain-5 is ubiquitously expressed in mammalian tissues and was recently found to be present in the mitochondria as well as in the cytosol, its physiological function and enzymological properties require further elucidation. The objective of the current study was to determine the characteristics of mitochondrial calpain-5 in porcine retinas, human HeLa cells, and C57BL/6J mice using subcellular fractionation. We found that mitochondrial calpain-5 was proteolyzed/autolyzed at low Ca2+ concentrations in mitochondria isolated from porcine retinas and by thapsigargin-induced endoplasmic reticulum (ER) stress in HeLa cells. Further, mitochondrial calpain-5, as opposed to cytosolic calpain-5, was activated during the early stages of ER stress in C57BL/6J mice. These results showed that mitochondrial calpain-5 was activated at low Ca2+ concentrations in vitro and in response to ER stress in vivo. The present study provides new insights into a novel calpain system in the mitochondria that includes stress responses during the early phases of ER stress. Further, activation of mitochondrial calpain-5 by treatment using low-molecular-weight compounds may have therapeutic potential for diseases related to ER stress, including neurodegenerative diseases, metabolic syndromes, diabetes, and cancer.

Emerging Regulatory Role of Nrf2 in Iron, Heme, and Hemoglobin Metabolism in Physiology and Disease.

Iron has played an important role in energy production since the beginning of life, as iron-catalyzed redox reactions are required for energy production. Oxygen, a highly efficient electron acceptor with high reduction potential, facilitates highly efficient energy production in eukaryotic cells. However, the increasing atmospheric oxygen concentration produces new threats to the organism, as oxygen reacts with iron and produces reactive oxygen species unless its levels are strictly regulated. As the size of multicellular organisms increases, these organisms must transport oxygen to the peripheral tissues and begin to employ red blood cells containing hemoglobin. This system is potentially a double-edged sword, as hemoglobin autoxidation occurs at a certain speed and releases free iron into the cytoplasm. Nrf2 belongs to the CNC transcription factor family, in which NF-E2p45 is the founding member. NF-E2p45 was first identified as a transcription factor that binds to the erythroid gene regulatory element NF-E2 located in the promoter region of the heme biosynthetic porphobilinogen deaminase gene. Human Nrf2 was also identified as a transcription factor that binds to the regulatory region of the ß-globin gene. Despite these original findings, NF-E2p45 and Nrf2 knockout mice exhibit few erythroid phenotypes. Nrf2 regulates the expression of a wide range of antioxidant and detoxification enzymes. In this review article, we describe and discuss the roles of Nrf2 in various iron-mediated bioreactions and its possible coevolution with iron and oxygen.

Presence of calpain-5 in mitochondria.

Calpains are Ca2+-dependent cysteine proteases that are widely distributed in animal tissues and modulate a variety of cellular processes. There are 15 members of the calpain family in mammals. In animal cells, there are three types of calpains, viz., calpain-1, calpain-2, and calpain-10 in the mitochondria. The three types of calpains have been shown to play significant roles in pathophysiological conditions, including in apoptosis- and necrosis-like cell death. One of the severe retinal diseases, autosomal dominant neovascular inflammatory vitreoretinopathy, is known to be induced by mutations of the calpain-5 gene. However, the distribution of calpain-5 in the retina has not been elucidated. Therefore, in the present study, we determined the localization of calpain-5 in the porcine retina. We detected calpain-5 in the inner segment of photoreceptor cells using immunohistochemistry. With immunoelectron microscopy, calpain-5 was localized in the mitochondria of photoreceptor cells. Western blot analyses showed that calpain-5 was present in each mitochondrial subfraction. Furthermore, we showed that the molecular weight of mitochondrial calpain-5 was slightly smaller than cytosolic one. Our results demonstrated that a novel mitochondrial calpian, calpain-5, was localized in the mitochondria of retinal photoreceptor cells.

Visual Responses of Photoreceptor-Degenerated Rats Expressing Two Different Types of Channelrhodopsin Genes.

Optogenetic technologies are expected to be applicable for clinical use in restoring vision. However, the degree of recovered visual function is highly dependent on the function of the chosen optogenetic gene. To investigate the effect on visual function of dual expression of genes with different wavelength sensitivities, we transduced a modified Volvox-derived channelrhodopsin gene (mVChR1) via an adeno-associated virus vector into transgenic rats harbouring the ChR2 gene in retinal ganglion cells. These transgenic rats were given an intraperitoneal injection of N-methyl-N-nitrosourea to induce the degeneration of native photoreceptor cells prior to transduction of mVChR1. Optical coherence tomography images indicated the degeneration of the native photoreceptor cells after the N-methyl-N-nitrosourea injection. Complete loss of function of the native photoreceptor cells was confirmed using electroretinograms. In the ChR2 transgenic rats, visually evoked potentials were clearly detectable in spite of native photoreceptor function abolishment; however the responses were limited to within blue wavelengths. In contrast, the limited wavelength sensitivities were improved by the additional transduction of mVChR1, which exhibited sensitivities to green and red. Thus, the transductions of dual genes encoding channelrhodopsins that exhibit different wavelength sensitivities represents a promising candidate method to expand and to enhance rescued wavelength sensitivities in blind subjects.

Improved transduction efficiencies of adeno-associated virus vectors by synthetic cell-permeable peptides.

Various serotypes of adeno-associated virus (AAV) vectors have been used for gene therapy and as research tools. Among these serotypes, the AAV type 2 vector has been used successfully in human gene therapies. However, the transduction efficiency of AAV2 depends on the cell type, and this poses a problem in the efficacy of gene therapy. To improve the transduction efficiency of AAV2, we designed a small peptide consisting of epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor peptide and the HIV-Tat sequence Tat-Y1068. Pre- or co-treatment of CYNOM-K1 cells from cynomolgus monkey embryo skin with Tat-Y1068 increased the transduction efficiencies in a dose-dependent manner and caused p38 phosphorylation. The transduction efficiency of AAV2 into the rat fibroblast cell line RAT-1 highly expressing EGFR was less than the transduction efficiency of AAV2 into CYNOM-K1 cells. Tat-Y1068 increased the transduction efficiency in RAT-1 cells in the same manner as in CYNOM-K1 cells. In conclusion, cell-permeable peptides possessing the EGFR tyrosine kinase inhibitor function might serve as a useful ingredient of AAV2 vector solution for increasing the transduction efficiency of gene therapies.

The protection of rat retinal ganglion cells from ischemia/reperfusion injury by the inhibitory peptide of mitochondrial µ-calpain.

Intracellular Ca(2+)-dependent cysteine proteases such as calpains have been suggested as critical factors in retinal ganglion cell (RGC) death. However, it is unknown whether mitochondrial calpains are involved in RGC death. The purpose of the present study was to determine whether the inhibition of mitochondrial µ-calpain activity protects against RGC death during ischemia/reperfusion (I/R) injury. This study used a well-established rat model of experimental acute glaucoma involving I/R injury. A specific peptide inhibitor of mitochondrial µ-calpain, Tat-µCL, was topically applied to rats via eye drops three times a day for 5 days after I/R. RGC death was determined by the terminal deoxynucleotidyl transferase dUTP nick end labeling assay. The truncation of apoptosis-inducing factor (AIF) was determined by western blot analyses. Retinal morphology was determined after staining with hematoxyline and eosin. In addition, the number of Fluoro Gold-labeled RGCs in flat-mounted retinas was used to determine the percentage of surviving RGCs after I/R injury. After 1 day of I/R, RGC death was observed in the ganglion cell layer. Treatment with Tat-µCL eye drops significantly prevented the death of RGCs and the truncation of AIF. After 5 days of I/R, RGC death decreased by approximately 40%. However, Tat-µCL significantly inhibited the decrease in the retinal sections and flat-mounted retinas. The results suggested that mitochondrial µ-calpain is associated with RGC death during I/R injury via truncation of AIF. In addition, the inhibition of mitochondrial µ-calpain activity by Tat-µCL had a neuroprotective effect against I/R-induced RGC death.

HSP90 Regulation of P2X7 Receptor Function Requires an Intact Cytoplasmic C-Terminus.

P2X7 receptors (P2X7Rs) are ATP-gated ion channels that display the unusual property of current facilitation during long applications of agonists. Here we show that facilitation disappears in chimeric P2X7Rs containing the C-terminus of the P2X2 receptor (P2X2R), and in a truncated P2X7R missing the cysteine-rich domain of the C-terminus. The chimeric and truncated receptors also show an apparent decreased permeability to N-methyl-d-glucamine(+) (NMDG(+)). The effects of genetic modification of the C-terminus on NMDG(+) permeability were mimicked by preapplication of the HSP90 antagonist geldanamycin to the wild-type receptor. Further, the geldanamycin decreased the shift in the reversal potential of the ATP-gated current measured under bi-ionic NMDG(+)/Na(+) condition without affecting the ability of the long application of agonist to facilitate current amplitude. Taken together, the results suggest that HSP90 may be essential for stabilization and function of P2X7Rs through an action on the cysteine-rich domain of the cytoplasmic the C-terminus.

Delivery of Topically Applied Calpain Inhibitory Peptide to the Posterior Segment of the Rat Eye.

We developed an inhibitory peptide that specifically acts against mitochondrial µ-calpain (Tat-µCL, 23 amino acid, 2857.37 Da) and protects photoreceptors in retinal dystrophic rats. In the present study, we topically administered Tat-µCL to the eyes of Sprague-Dawley rats for 7 days to determine both the delivery route of the peptide to the posterior segment of the eye and the kinetics after topical application in adult rats. Distribution of the peptide was determined by immunohistochemical analysis, and enzyme-linked immune-absorbent assay was used to quantify the accumulation in the retina. Peptides were prominently detected in both the anterior and posterior segments of the eye at 1 h after the final eye drop application. Immunohistochemically positive reactions were observed in the retina, optic nerve, choroid, sclera and the retrobulbar tissues, even in the posterior portion of the eye. Immunoactivities gradually diminished at 3 and 6 h after the final eye drop. Quantitative estimations of the amount of peptide in the retina were 15.3, 5.8 and 1.0 pg/µg protein at 1, 3 and 6 h after the final instillation, respectively. Current results suggest that while the topically applied Tat-µCL peptide reaches the posterior segment of the retina and the optic nerve, the sufficient concentration (> IC50) is maintained for at least 6 h in the rat retina. Our findings suggest that delivery of topically applied peptide to the posterior segment and optic nerve occurs through the conjunctiva, periocular connective tissue, sclera and optic nerve sheath.

Emerging functional cross-talk between the Keap1-Nrf2 system and mitochondria.

Nuclear factor erythroid-derived 2-related factor 2 (Nrf2) was originally identified as a positive regulator of drug detoxifying enzyme gene expression during exposure to environmental electrophiles. Currently, Nrf2 is known to regulate the expression of hundreds of cytoprotective genes to counteract endogenously or exogenously generated oxidative stress. Furthermore, when activated in human tumors by somatic mutations, Nrf2 confers growth advantages and chemoresistance by regulating genes involved in various processes such as the pentose phosphate pathway and nucleotide synthesis in addition to antioxidant proteins. Interestingly, increasing evidence shows that Nrf2 is associated with mitochondrial biogenesis during environmental stresses in certain tissues such as the heart. Furthermore, SKN-1, a functional homolog of Nrf2 in C. elegans, is activated by mitochondrial reactive oxygen species and extends life span by promoting mitochondrial homeostasis (i.e., mitohormesis). Similarly, Nrf2 activation was recently observed in the heart of surfeit locus protein 1 (Surf1) -/- mice in which cellular respiration was decreased due to cytochrome c oxidase defects. In this review, we critically examine the relationship between Nrf2 and mitochondria and argue that the Nrf2 stress pathway intimately communicates with mitochondria to maintain cellular homeostasis during oxidative stress.