Exendin-4 protects against high glucose-induced mitochondrial dysfunction and oxidative stress in SH-SY5Y neuroblastoma cells through GLP-1 receptor/Epac/Akt signaling.

Mitochondrial dysfunction under diabetic condition leads to the development and progression of neurodegenerative complications. Recently, the beneficial effects of glucagon-like peptide-1 (GLP-1) receptor agonists on diabetic neuropathies have been widely recognized. However, molecular mechanisms underlying the neuroprotective effects of GLP-1 receptor agonists against high glucose (HG)-induced neuronal damages is not completely elucidated. Here, we investigated the underlying mechanisms of GLP-1 receptor agonist treatment against oxidative stress, mitochondrial dysfunction, and neuronal damages under HG conditions mimicking a diabetic hyperglycemic state in SH-SY5Y neuroblastoma cells. We revealed that treatment with exendin-4, a GLP-1 receptor agonist, not only increased the expression of survival markers, phospho-Akt/Akt and Bcl-2, but also decreased the expression of pro-apoptotic marker, Bax, and reduced the levels of reactive oxygen species (ROS) defense markers (catalase, SOD-2, and HO-1) under HG conditions. The expressions of mitochondrial function associated genes, MCU and UCP3, and mitochondrial fission genes, DRP1 and FIS1, were decreased by exendin-4 compared to non-treated levels, while the protein expression levels of mitochondrial homeostasis regulators, Parkin and PINK1, were enhanced. In addition, blockade of Epac and Akt activities was able to antagonize these neuroprotective effects of exendin-4. Collectively, we demonstrated that stimulation of GLP-1 receptor propagates a neuroprotective cascade against the oxidative stress and mitochondrial dysfunction as well as augments survival through the Epac/Akt-dependent pathway. Therefore, the revealed mechanisms underlying GLP-1 receptor pathway by preserving mitochondrial homeostasis would be a therapeutic candidate to alleviate neuronal dysfunctions and delay the progression of diabetic neuropathies.

Modulation of Reactive Oxygen Species Homeostasis as a Pleiotropic Effect of Commonly Used Drugs.

Age-associated diseases represent a growing burden for global health systems in our aging society. Consequently, we urgently need innovative strategies to counteract these pathological disturbances. Overwhelming generation of reactive oxygen species (ROS) is associated with age-related damage, leading to cellular dysfunction and, ultimately, diseases. However, low-dose ROS act as crucial signaling molecules and inducers of a vaccination-like response to boost antioxidant defense mechanisms, known as mitohormesis. Consequently, modulation of ROS homeostasis by nutrition, exercise, or pharmacological interventions is critical in aging. Numerous nutrients and approved drugs exhibit pleiotropic effects on ROS homeostasis. In the current review, we provide an overview of drugs affecting ROS generation and ROS detoxification and evaluate the potential of these effects to counteract the development and progression of age-related diseases. In case of inflammation-related dysfunctions, cardiovascular- and neurodegenerative diseases, it might be essential to strengthen antioxidant defense mechanisms in advance by low ROS level rises to boost the individual ROS defense mechanisms. In contrast, induction of overwhelming ROS production might be helpful to fight pathogens and kill cancer cells. While we outline the potential of ROS manipulation to counteract age-related dysfunction and diseases, we also raise the question about the proper intervention time and dosage.

ALG-2 and peflin regulate COPII targeting and secretion in response to calcium signaling.

ER-to-Golgi transport is the first step in the constitutive secretory pathway, which, unlike regulated secretion, is believed to proceed nonstop independent of Ca2+ flux. However, here we demonstrate that penta-EF hand (PEF) proteins ALG-2 and peflin constitute a hetero-bifunctional COPII regulator that responds to Ca2+ signaling by adopting one of several distinct activity states. Functionally, these states can adjust the rate of ER export of COPII-sorted cargos up or down by ∼50%. We found that at steady-state Ca2+, ALG-2/peflin hetero-complexes bind to ER exit sites (ERES) through the ALG-2 subunit to confer a low, buffered secretion rate, while peflin-lacking ALG-2 complexes markedly stimulate secretion. Upon Ca2+ signaling, ALG-2 complexes lacking peflin can either increase or decrease the secretion rate depending on signaling intensity and duration-phenomena that could contribute to cellular growth and intercellular communication following secretory increases or protection from excitotoxicity and infection following decreases. In epithelial normal rat kidney (NRK) cells, the Ca2+-mobilizing agonist ATP causes ALG-2 to depress ER export, while in neuroendocrine PC12 cells, Ca2+ mobilization by ATP results in ALG-2-dependent enhancement of secretion. Furthermore, distinct Ca2+ signaling patterns in NRK cells produce opposing ALG-2-dependent effects on secretion. Mechanistically, ALG-2-dependent depression of secretion involves decreased levels of the COPII outer shell and increased peflin targeting to ERES, while ALG-2-dependent enhancement of secretion involves increased COPII outer shell and decreased peflin at ERES. These data provide insights into how PEF protein dynamics affect secretion of important physiological cargoes such as collagen I and significantly impact ER stress.

Green tea catechins EGCG and ECG enhance the fitness and lifespan of Caenorhabditis elegans by complex I inhibition.

Green tea catechins are associated with a delay in aging. We have designed the current study to investigate the impact and to unveil the target of the most abundant green tea catechins, epigallocatechin gallate (EGCG) and epicatechin gallate (ECG). Experiments were performed in Caenorhabditis elegans to analyze cellular metabolism, ROS homeostasis, stress resistance, physical exercise capacity, health- and lifespan, and the underlying signaling pathways. Besides, we examined the impact of EGCG and ECG in isolated murine mitochondria. A concentration of 2.5 µM EGCG and ECG enhanced health- and lifespan as well as stress resistance in C. elegans. Catechins hampered mitochondrial respiration in C. elegans after 6-12 h and the activity of complex I in isolated rodent mitochondria. The impaired mitochondrial respiration was accompanied by a transient drop in ATP production and a temporary increase in ROS levels in C. elegans. After 24 h, mitochondrial respiration and ATP levels got restored, and ROS levels even dropped below control conditions. The lifespan increases induced by EGCG and ECG were dependent on AAK-2/AMPK and SIR-2.1/SIRT1, as well as on PMK-1/p38 MAPK, SKN-1/NRF2, and DAF-16/FOXO. Long-term effects included significantly diminished fat content and enhanced SOD and CAT activities, required for the positive impact of catechins on lifespan. In summary, complex I inhibition by EGCG and ECG induced a transient drop in cellular ATP levels and a temporary ROS burst, resulting in SKN-1 and DAF-16 activation. Through adaptative responses, catechins reduced fat content, enhanced ROS defense, and improved healthspan in the long term.

PCK2 opposes mitochondrial respiration and maintains the redox balance in starved lung cancer cells.

Cancer cells frequently lack nutrients like glucose, due to insufficient vascular networks. Mitochondrial phosphoenolpyruvate carboxykinase, PCK2, has recently been found to mediate partial gluconeogenesis and hence anabolic metabolism in glucose starved cancer cells. Here we show that PCK2 acts as a regulator of mitochondrial respiration and maintains the redox balance in nutrient-deprived human lung cancer cells. PCK2 silencing increased the abundance and interconversion of tricarboxylic acid (TCA) cycle intermediates, augmented mitochondrial respiration and enhanced glutathione oxidation under glucose and serum starvation, in a PCK2 re-expression reversible manner. Moreover, enhancing the TCA cycle by PCK2 inhibition severely reduced colony formation of lung cancer cells under starvation. As a conclusion, PCK2 contributes to maintaining a reduced glutathione pool in starved cancer cells besides mediating the biosynthesis of gluconeogenic/glycolytic intermediates. The study sheds light on adaptive responses in cancer cells to nutrient deprivation and shows that PCK2 confers protection against respiration-induced oxidative stress.

Near-UV Light Induced ROS Production Initiates Spatial Ca2+ Spiking to Fire NFATc3 Translocation.

Ca2+-dependent gene regulation controls several functions to determine the fate of the cells. Proteins of the nuclear factor of activated T-cells (NFAT) family are Ca2+ sensitive transcription factors that control the cell growth, proliferation and insulin secretion in ß-cells. Translocation of NFAT proteins to the nucleus occurs in a sequence of events that starts with activating calmodulin-dependent phosphatase calcineurin in a Ca2+-dependent manner, which dephosphorylates the NFAT proteins and leads to their translocation to the nucleus. Here, we examined the role of IP3-generating agonists and near-UV light in the induction of NFATc3 migration to the nucleus in the pancreatic ß-cell line INS-1. Our results show that IP3 generation yields cytosolic Ca2+ rise and NFATc3 translocation. Moreover, near-UV light exposure generates reactive oxygen species (ROS), resulting in cytosolic Ca2+ spiking via the L-type Ca2+ channel and triggers NFATc3 translocation to the nucleus. Using the mitochondria as a Ca2+ buffering tool, we showed that ROS-induced cytosolic Ca2+ spiking, not the ROS themselves, was the triggering mechanism of nuclear import of NFATc3. Collectively, this study reveals the mechanism of near-UV light induced NFATc3 migration.

Sigma-1 Receptor Promotes Mitochondrial Bioenergetics by Orchestrating ER Ca2+ Leak during Early ER Stress.

The endoplasmic reticulum (ER) is a complex, multifunctional organelle of eukaryotic cells and responsible for the trafficking and processing of nearly 30% of all human proteins. Any disturbance to these processes can cause ER stress, which initiates an adaptive mechanism called unfolded protein response (UPR) to restore ER functions and homeostasis. Mitochondrial ATP production is necessary to meet the high energy demand of the UPR, while the molecular mechanisms of ER to mitochondria crosstalk under such stress conditions remain mainly enigmatic. Thus, better understanding the regulation of mitochondrial bioenergetics during ER stress is essential to combat many pathologies involving ER stress, the UPR, and mitochondria. This article investigates the role of Sigma-1 Receptor (S1R), an ER chaperone, has in enhancing mitochondrial bioenergetics during early ER stress using human neuroblastoma cell lines. Our results show that inducing ER stress with tunicamycin, a known ER stressor, greatly enhances mitochondrial bioenergetics in a time- and S1R-dependent manner. This is achieved by enhanced ER Ca2+ leak directed towards mitochondria by S1R during the early phase of ER stress. Our data point to the importance of S1R in promoting mitochondrial bioenergetics and maintaining balanced H2O2 metabolism during early ER stress.

Dynamic Control of Mitochondrial Ca2+ Levels as a Survival Strategy of Cancer Cells.

Cancer cells have increased energy requirements due to their enhanced proliferation activity. This energy demand is, among others, met by mitochondrial ATP production. Since the second messenger Ca2+ maintains the activity of Krebs cycle dehydrogenases that fuel mitochondrial respiration, proper mitochondrial Ca2+ uptake is crucial for a cancer cell survival. However, a mitochondrial Ca2+ overload induces mitochondrial dysfunction and, ultimately, apoptotic cell death. Because of the vital importance of balancing mitochondrial Ca2+ levels, a highly sophisticated machinery of multiple proteins manages mitochondrial Ca2+ homeostasis. Notably, mitochondria sequester Ca2+ preferentially at the interaction sites between mitochondria and the endoplasmic reticulum (ER), the largest internal Ca2+ store, thus, pointing to mitochondrial-associated membranes (MAMs) as crucial hubs between cancer prosperity and cell death. To investigate potential regulatory mechanisms of the mitochondrial Ca2+ uptake routes in cancer cells, we modulated mitochondria-ER tethering and the expression of UCP2 and analyzed mitochondrial Ca2+ homeostasis under the various conditions. Hence, the expression of contributors to mitochondrial Ca2+ regulation machinery was quantified by qRT-PCR. We further used data from The Cancer Genome Atlas (TCGA) to correlate these in vitro findings with expression patterns in human breast invasive cancer and human prostate adenocarcinoma. ER-mitochondrial linkage was found to support a mitochondrial Ca2+ uptake route dependent on uncoupling protein 2 (UCP2) in cancer cells. Notably, combined overexpression of Rab32, a protein kinase A-anchoring protein fostering the ER-mitochondrial tethering, and UCP2 caused a significant drop in cancer cells' viability. Artificially enhanced ER-mitochondrial tethering further initiated a sudden decline in the expression of UCP2, probably as an adaptive response to avoid mitochondrial Ca2+ overload. Besides, TCGA analysis revealed an inverse expression correlation between proteins stabilizing mitochondrial-ER linkage and UCP2 in tissues of human breast invasive cancer and prostate adenocarcinoma. Based on these results, we assume that cancer cells successfully manage mitochondrial Ca2+ uptake to stimulate Ca2+-dependent mitochondrial metabolism while avoiding Ca2+-triggered cell death by fine-tuning ER-mitochondrial tethering and the expression of UCP2 in an inversed manner. Disruption of this equilibrium yields cancer cell death and may serve as a treatment strategy to specifically kill cancer cells.

Targeting cellular senescence based on interorganelle communication, multilevel proteostasis, and metabolic control.

Cellular senescence, a stable cell division arrest caused by severe damage and stress, is a hallmark of aging in vertebrates including humans. With progressing age, senescent cells accumulate in a variety of mammalian tissues, where they contribute to tissue aging, identifying cellular senescence as a major target to delay or prevent aging. There is an increasing demand for the discovery of new classes of small molecules that would either avoid or postpone cellular senescence by selectively eliminating senescent cells from the body (i.e., 'senolytics') or inactivating/switching damage-inducing properties of senescent cells (i.e., 'senostatics/senomorphics'), such as the senescence-associated secretory phenotype. Whereas compounds with senolytic or senostatic activity have already been described, their efficacy and specificity has not been fully established for clinical use yet. Here, we review mechanisms of senescence that are related to mitochondria and their interorganelle communication, and the involvement of proteostasis networks and metabolic control in the senescent phenotype. These cellular functions are associated with cellular senescence in in vitro and in vivo models but have not been fully exploited for the search of new compounds to counteract senescence yet. Therefore, we explore possibilities to target these mechanisms as new opportunities to selectively eliminate and/or disable senescent cells with the aim of tissue rejuvenation. We assume that this research will provide new compounds from the chemical space which act as mimetics of caloric restriction, modulators of calcium signaling and mitochondrial physiology, or as proteostasis optimizers, bearing the potential to counteract cellular senescence, thereby allowing healthy aging.

Interrelation between ROS and Ca2+ in aging and age-related diseases.

Calcium (Ca2+) and reactive oxygen species (ROS) are versatile signaling molecules coordinating physiological and pathophysiological processes. While channels and pumps shuttle Ca2+ ions between extracellular space, cytosol and cellular compartments, short-lived and highly reactive ROS are constantly generated by various production sites within the cell. Ca2+ controls membrane potential, modulates mitochondrial adenosine triphosphate (ATP) production and affects proteins like calcineurin (CaN) or calmodulin (CaM), which, in turn, have a wide area of action. Overwhelming Ca2+ levels within mitochondria efficiently induce and trigger cell death. In contrast, ROS comprise a diverse group of relatively unstable molecules with an odd number of electrons that abstract electrons from other molecules to gain stability. Depending on the type and produced amount, ROS act either as signaling molecules by affecting target proteins or as harmful oxidative stressors by damaging cellular components. Due to their wide range of actions, it is little wonder that Ca2+ and ROS signaling pathways overlap and impact one another. Growing evidence suggests a crucial implication of this mutual interplay on the development and enhancement of age-related disorders, including cardiovascular and neurodegenerative diseases as well as cancer.

tBHP treatment as a model for cellular senescence and pollution-induced skin aging.

Accumulation of senescent cells promotes the development of age-related pathologies and deterioration. In human skin, senescent cells potentially impair structure and function by secreting a mixture of signaling molecules and proteases that influence neighboring cells and degrade extracellular matrix components, such as elastin and collagen. One of the key underlying mechanisms of senescence and extrinsic skin aging is the increase of intracellular reactive oxygen species and resulting oxidative stress. Tert-butyl hydroperoxide (tBHP) is a known inducer of oxidative stress and cellular damage, acting at least in part by depleting the antioxidant glutathione. Here, we provide a detailed characterization of tBHP-induced senescence in human dermal fibroblasts in monolayer culture. In addition, results obtained with more physiological experimental models revealed that tBHP treated 3D reconstructed skin and ex vivo skin developed signs of chronic tissue damage, displaying reduced epidermal thickness and collagen fiber thinning. We, therefore, propose that tBHP treatment can be used as a model to study the effects of extrinsic skin aging, focusing mainly on the influence of environmental pollution.

Myeloperoxidase and Septic Conditions Disrupt Sphingolipid Homeostasis in Murine Brain Capillaries In Vivo and Immortalized Human Brain Endothelial Cells In Vitro.

During inflammation, activated leukocytes release cytotoxic mediators that compromise blood-brain barrier (BBB) function. Under inflammatory conditions, myeloperoxidase (MPO) is critically involved in inflicting BBB damage. We used genetic and pharmacological approaches to investigate whether MPO induces aberrant lipid homeostasis at the BBB in a murine endotoxemia model. To corroborate findings in a human system we studied the impact of sera from sepsis and non-sepsis patients on brain endothelial cells (hCMEC/D3). In response to endotoxin, the fatty acid, ceramide, and sphingomyelin content of isolated mouse brain capillaries dropped and barrier dysfunction occurred. In mice, genetic deficiency or pharmacological inhibition of MPO abolished these alterations. Studies in metabolic cages revealed increased physical activity and less pronounced sickness behavior of MPO-/- compared to wild-type mice in response to sepsis. In hCMEC/D3 cells, exogenous tumor necrosis factor α (TNFα) potently regulated gene expression of pro-inflammatory cytokines and a set of genes involved in sphingolipid (SL) homeostasis. Notably, treatment of hCMEC/D3 cells with sera from septic patients reduced cellular ceramide concentrations and induced barrier and mitochondrial dysfunction. In summary, our in vivo and in vitro data revealed that inflammatory mediators including MPO, TNFα induce dysfunctional SL homeostasis in brain endothelial cells. Genetic and pharmacological inhibition of MPO attenuated endotoxin-induced alterations in SL homeostasis in vivo, highlighting the potential role of MPO as drug target to treat inflammation-induced brain dysfunction.

Tracking intra- and inter-organelle signaling of mitochondria.

Mitochondria are as highly specialized organelles and masters of the cellular energy metabolism in a constant and dynamic interplay with their cellular environment, providing adenosine triphosphate, buffering Ca2+ and fundamentally contributing to various signaling pathways. Hence, such broad field of action within eukaryotic cells requires a high level of structural and functional adaptation. Therefore, mitochondria are constantly moving and undergoing fusion and fission processes, changing their shape and their interaction with other organelles. Moreover, mitochondrial activity gets fine-tuned by intra- and interorganelle H+ , K+ , Na+ , and Ca2+ signaling. In this review, we provide an up-to-date overview on mitochondrial strategies to adapt and respond to, as well as affect, their cellular environment. We also present cutting-edge technologies used to track and investigate subcellular signaling, essential to the understanding of various physiological and pathophysiological processes.

Mitochondrial-Endoplasmic Reticulum Interplay: A Lifelong On-Off Relationship?

This article comments recent publications that highlight an intriguing importance of specific settings in the interaction between the mitochondria and the endoplasmic reticulum to ensure cell-specific functions like the responsiveness to elevated glucose in pancreatic ß-cells. Hence, alterations of the mitochondria-endoplasmic reticulum communications under various pathological conditions like aging or cancer often come with enhanced Ca2+ transfer that, in turn, yields stimulation of basal mitochondrial activity to meet the increasing adenosine triphosphate demand of the very cell. Such observations identify mitochondria-associated membranes as potential target for new therapeutic strategies against aging or cancer.

MICU1 controls cristae junction and spatially anchors mitochondrial Ca2+ uniporter complex.

Recently identified core proteins (MICU1, MCU, EMRE) forming the mitochondrial Ca2+ uniporter complex propelled investigations into its physiological workings. Here, we apply structured illumination microscopy to visualize and localize these proteins in living cells. Our data show that MICU1 localizes at the inner boundary membrane (IBM) due to electrostatic interaction of its polybasic domain. Moreover, this exclusive localization of MICU1 is important for the stability of cristae junctions (CJ), cytochrome c release and mitochondrial membrane potential. In contrast to MICU1, MCU and EMRE are homogeneously distributed at the inner mitochondrial membrane under resting conditions. However, upon Ca2+ elevation MCU and EMRE dynamically accumulate at the IBM in a MICU1-dependent manner. Eventually, our findings unveil an essential function of MICU1 in CJ stabilization and provide mechanistic insights of how sophistically MICU1 controls the MCU-Complex while maintaining the structural mitochondrial membrane framework.

Glycogen Synthase Kinase 3 Beta Controls Presenilin-1-Mediated Endoplasmic Reticulum Ca²âº Leak Directed to Mitochondria in Pancreatic Islets and ß-Cells.

BACKGROUND/AIMS: In pancreatic ß-cells, the intracellular Ca²âº homeostasis is an essential regulator of the cells major functions. The endoplasmic reticulum (ER) as interactive intracellular Ca²âº store balances cellular Ca²âº. In this study basal ER Ca²âº homeostasis was evaluated in order to reveal potential ß-cell-specificity of ER Ca²âº handling and its consequences for mitochondrial Ca²âº, ATP and respiration. METHODS: The two pancreatic cell lines INS-1 and MIN-6, freshly isolated pancreatic islets, and the two non-pancreatic cell lines HeLA and EA.hy926 were used. Cytosolic, ER and mitochondrial Ca²âº and ATP measurements were performed using single cell fluorescence microscopy and respective (genetically-encoded) sensors/dyes. Mitochondrial respiration was monitored by respirometry. GSK3ß activity was measured with ELISA. RESULTS: An atypical ER Ca²âº leak was observed exclusively in pancreatic islets and ß-cells. This continuous ER Ca²âº efflux is directed to mitochondria and increases basal respiration and organellar ATP levels, is established by GSK3ß-mediated phosphorylation of presenilin-1, and is prevented by either knockdown of presenilin-1 or an inhibition/knockdown of GSK3ß. Expression of a presenlin-1 mutant that mimics GSK3ß-mediated phosphorylation established a ß-cell-like ER Ca²âº leak in HeLa and EA.hy926 cells. The ER Ca²âº loss in ß-cells was compensated at steady state by Ca²âº entry that is linked to the activity of TRPC3. CONCLUSION: Pancreatic ß-cells establish a cell-specific ER Ca²âº leak that is under the control of GSK3ß and directed to mitochondria, thus, reflecting a cell-specific intracellular Ca²âº handling for basal mitochondrial activity.

Enhanced inter-compartmental Ca2+ flux modulates mitochondrial metabolism and apoptotic threshold during aging.

BACKGROUND: Senescence is characterized by a gradual decline in cellular functions, including changes in energy homeostasis and decreased proliferation activity. As cellular power plants, contributors to signal transduction, sources of reactive oxygen species (ROS) and executors of programmed cell death, mitochondria are in a unique position to affect aging-associated processes of cellular decline. Notably, metabolic activation of mitochondria is tightly linked to Ca2+ due to the Ca2+ -dependency of several enzymes in the Krebs cycle, however, overload of mitochondria with Ca2+ triggers cell death pathways. Consequently, a machinery of proteins tightly controls mitochondrial Ca2+ homeostasis as well as the exchange of Ca2+ between the different cellular compartments, including Ca2+ flux between mitochondria and the endoplasmic reticulum (ER). METHODS: In this study, we investigated age-related changes in mitochondrial Ca2+ homeostasis, mitochondrial-ER linkage and the activity of the main ROS production site, the mitochondrial respiration chain, in an in vitro aging model based on porcine aortic endothelial cells (PAECs), using high-resolution live cell imaging, proteomics and various molecular biological methods. RESULTS: We describe that in aged endothelial cells, increased ER-mitochondrial Ca2+ crosstalk occurs due to enhanced ER-mitochondrial tethering. The close functional inter-organelle linkage increases mitochondrial Ca2+ uptake and thereby the activity of the mitochondrial respiration, but also makes senescent cells more vulnerable to mitochondrial Ca2+-overload-induced cell death. Moreover, we identified the senolytic properties of the polyphenol resveratrol, triggering cell death via mitochondrial Ca2+ overload exclusively in senescent cells. CONCLUSION: By unveiling aging-related changes in the inter-organelle tethering and Ca2+ communications we have advanced the understanding of endothelial aging and highlighted a potential basis to develop drugs specifically targeting senescent cells.

Real-Time Imaging of Mitochondrial ATP Dynamics Reveals the Metabolic Setting of Single Cells.

Reprogramming of metabolic pathways determines cell functions and fate. In our work, we have used organelle-targeted ATP biosensors to evaluate cellular metabolic settings with high resolution in real time. Our data indicate that mitochondria dynamically supply ATP for glucose phosphorylation in a variety of cancer cell types. This hexokinase-dependent process seems to be reversed upon the removal of glucose or other hexose sugars. Our data further verify that mitochondria in cancer cells have increased ATP consumption. Similar subcellular ATP fluxes occurred in young mouse embryonic fibroblasts (MEFs). However, pancreatic beta cells, senescent MEFs, and MEFs lacking mitofusin 2 displayed completely different mitochondrial ATP dynamics, indicative of increased oxidative phosphorylation. Our findings add perspective to the variability of the cellular bioenergetics and demonstrate that live cell imaging of mitochondrial ATP dynamics is a powerful tool to evaluate metabolic flexibility and heterogeneity at a single-cell level.

Cytosolic Aspartate Availability Determines Cell Survival When Glutamine Is Limiting.

Mitochondrial function is important for aspartate biosynthesis in proliferating cells. Here, we show that mitochondrial aspartate export via the aspartate-glutamate carrier 1 (AGC1) supports cell proliferation and cellular redox homeostasis. Insufficient cytosolic aspartate delivery leads to cell death when TCA cycle carbon is reduced following glutamine withdrawal and/or glutaminase inhibition. Moreover, loss of AGC1 reduces allograft tumor growth that is further compromised by treatment with the glutaminase inhibitor CB-839. Together, these findings argue that mitochondrial aspartate export sustains cell survival in low-glutamine environments and AGC1 inhibition can synergize with glutaminase inhibition to limit tumor growth.

Targeting Mitochondria to Counteract Age-Related Cellular Dysfunction.

Senescence is related to the loss of cellular homeostasis and functions, which leads to a progressive decline in physiological ability and to aging-associated diseases. Since mitochondria are essential to energy supply, cell differentiation, cell cycle control, intracellular signaling and Ca2+ sequestration, fine-tuning mitochondrial activity appropriately, is a tightrope walk during aging. For instance, the mitochondrial oxidative phosphorylation (OXPHOS) ensures a supply of adenosine triphosphate (ATP), but is also the main source of potentially harmful levels of reactive oxygen species (ROS). Moreover, mitochondrial function is strongly linked to mitochondrial Ca2+ homeostasis and mitochondrial shape, which undergo various alterations during aging. Since mitochondria play such a critical role in an organism's process of aging, they also offer promising targets for manipulation of senescent cellular functions. Accordingly, interventions delaying the onset of age-associated disorders involve the manipulation of mitochondrial function, including caloric restriction (CR) or exercise, as well as drugs, such as metformin, aspirin, and polyphenols. In this review, we discuss mitochondria's role in and impact on cellular aging and their potential to serve as a target for therapeutic interventions against age-related cellular dysfunction.