Metabolism alteration in follicular niche: The nexus among intermediary metabolism,mitochondrial function,and classic polycystic ovary syndrome

Classic polycystic ovary syndrome (PCOS) is a high-risk phenotype accompanied by increased risks of reproductive and metabolic abnormalities; however, the local metabolism characteristics of the ovaries and their effects on germ cell development are unclear. The present study used targeted metabolomics to detect alterations in the intermediate metabolites of follicular fluid from classic PCOS patients, and the results indicated that hyperandrogenism but not obesity induced the changed intermediate metabolites in classic PCOS patients. Regarding the direct contact, we identified mitochondrial function, redox potential, and oxidative stress in cumulus cells which were necessary to support oocyte growth before fertilization, and suggested dysfunction of mitochondria, imbalanced redox potential, and increased oxidative stress in cumulus cells of classic PCOS patients. Follicular fluid intermediary metabolic profiles provide signatures of classic PCOS ovary local metabolism and establish a close link with mitochondria dysfunction of cumulus cells, highlighting the role of metabolic signal and mitochondrial cross talk involved in the pathogenesis of classic PCOS.     

PGAM5: A crucial role in mitochondrial dynamics and programmed cell death

In response to mitochondrial damage, mitochondria activate mitochondrial dynamics to maintain normal functions, and an imbalance in mitochondrial dynamics triggers multiple programmed cell death processes. Recent studies have shown that phosphoglycerate mutase 5 (PGAM5) is associated with mitochondrial damage. PGAM5 activates mitochondrial biogenesis and mitophagy to promote a cellular compensatory response when mitochondria are mildly damaged, whereas severe damage to mitochondria leads to PGAM5 inducing excessive mitochondria fission, disruption to mitochondrial movement, and amplification of apoptosis, necroptosis and mitophagic death signals, which eventually evoke cell death. PGAM5 functions mainly through protein-protein interactions and specific Ser/Thr/His protein phosphatase activity. PGAM5 is also regulated by mitochondrial proteases. Detection of PGAM5 and its interacting protein partners should enable a more accurate evaluation of mitochondrial damage and a more precise method for the diagnosis and treatment of diseases.     

Rhomboid Protease PARL Mediates the Mitochondrial Membrane Potential Loss-induced Cleavage of PGAM5

Regulated intramembrane proteolysis is a widely conserved mechanism for controlling diverse biological processes. Considering that proteolysis is irreversible, it must be precisely regulated in a context-dependent manner. Here, we show that phosphoglycerate mutase 5 (PGAM5), a mitochondrial Ser/Thr protein phosphatase, is cleaved in its N-terminal transmembrane domain in response to mitochondrial membrane potential (ΔΨ_m) loss. This ΔΨ_m loss-dependent cleavage of PGAM5 was mediated by presenilin-associated rhomboid-like (PARL). PARL is a mitochondrial resident rhomboid serine protease and has recently been reported to mediate the cleavage of PINK1, a mitochondrial Ser/Thr protein kinase, in healthy mitochondria with intact ΔΨ_m. Intriguingly, we found that PARL dissociated from PINK1 and reciprocally associated with PGAM5 in response to ΔΨ_m loss. These results suggest that PARL mediates differential cleavage of PINK1 and PGAM5 depending on the health status of mitochondria. Our data provide a prototypical example of stress-dependent regulation of PARL-mediated regulated intramembrane proteolysis.     

IFN‐β rescues neurodegeneration by regulating mitochondrial fission via STAT5, PGAM5, and Drp1

Mitochondrial homeostasis is essential for providing cellular energy, particularly in resource‐demanding neurons, defects in which cause neurodegeneration, but the function of interferons (IFNs) in regulating neuronal mitochondrial homeostasis is unknown. We found that neuronal IFN‐β is indispensable for mitochondrial homeostasis and metabolism, sustaining ATP levels and preventing excessive ROS by controlling mitochondrial fission. IFN‐β induces events that are required for mitochondrial fission, phosphorylating STAT5 and upregulating PGAM5, which phosphorylates serine 622 of Drp1. IFN‐β signaling then recruits Drp1 to mitochondria, oligomerizes it, and engages INF2 to stabilize mitochondria–endoplasmic reticulum (ER) platforms. This process tethers damaged mitochondria to the ER to separate them via fission. Lack of neuronal IFN‐β in the Ifnb ^–/– model of Parkinson disease (PD) disrupts STAT5‐PGAM5‐Drp1 signaling, impairing fission and causing large multibranched, damaged mitochondria with insufficient ATP production and excessive oxidative stress to accumulate. In other PD models, IFN‐β rescues dopaminergic neuronal cell death and pathology, associated with preserved mitochondrial homeostasis. Thus, IFN‐β activates mitochondrial fission in neurons through the pSTAT5/PGAM5/^S622Drp1 pathway to stabilize mitochondria/ER platforms, constituting an essential neuroprotective mechanism.     

Yeast replicative life span--the mitochondrial connection

Mitochondria have been associated with aging in many experimental systems through the damaging action of reactive oxygen species. There is more, however, to the connection between mitochondria and Saccharomyces cerevisiae longevity and aging. Induction of the retrograde response, a pathway signaling mitochondrial dysfunction, results in the extension of life span and postponement of the manifestations of aging, changing the metabolic and stress resistance status of the cell. A paradox associated with the retrograde response is the simultaneous triggering of extrachromosomal ribosomal DNA circle (ERC) production, because of the deleterious effect these circles have on yeast longevity. The retrograde response gene RTG2 appears to play a pivotal role in ERC production, linking metabolism and genome stability. In addition to mother cell aging, mitochondria are important in establishment of age asymmetry between mother and daughter cells. The results more generally point to the existence of a mechanism to "filter" damaged components from daughter cells, a form of checkpoint control. Mitochondrial integrity is affected by the PHB1 and PHB2 genes, which encode inner mitochondrial membrane chaperones called prohibitins. The Phb1/2 proteins protect the cell from imbalances in the production of mitochondrial proteins. Such imbalances appear to cause a stochastic stratification of the yeast population with the appearance of short-lived cells. Ras2p impacts this process. Maintenance of mitochondrial membrane potential and the provision of Krebs cycle intermediates for biosyntheses appear to be crucial elements in yeast longevity. In sum, it is clear that mitochondria lie at the nexus of yeast longevity and aging.     

The fine structure of the cumulus oophorus during follicular development in sheep

Summary The cumulus and membrana granulosa of non-atretic ovarian follicles from primordial up to a stage shortly before ovulation were studied by electron microscopy.The follicular cells of primordial follicles were undifferentiated and rested on a thick basal lamina. In secondary follicles the endoplasmic reticulum had proliferated forming an anastomosing network. In early antral and antral follicles (0.5–2.0 mm dia.) the ER was composed of short cisternae, the mitochondria had elongated and gap junctions were first observed. In late antral follicles (3.0–5.9 mm dia.) gap junctions were frequent. In the cumulus the glycogen was associated with electron lucent areas whereas in the granulosa it was invariably associated with membranes. In large antral follicles large membrane bound bodies were present in the basal cells of the cumulus. At early oestrus a distinctive mitochondrial morphology was noted in the granulosa but not elsewhere in the follicles. At mid oestrus numerous annular nexuses were present in the granulosa but not in the cumulus. At late oestrus numerous lipid droplets were formed in both cumulus and granulosa, the boundary with theca interna became indistinct and the basal lamina became incomplete.Deceased     

Aging shifts mitochondrial dynamics toward fission to promote germline stem cell loss

Changes in mitochondrial dynamics (fusion and fission) are known to occur during stem cell differentiation; however, the role of this phenomenon in tissue aging remains unclear. Here, we report that mitochondrial dynamics are shifted toward fission during aging of Drosophila ovarian germline stem cells (GSCs), and this shift contributes to aging‐related GSC loss. We found that as GSCs age, mitochondrial fragmentation and expression of the mitochondrial fission regulator, Dynamin‐related protein (Drp1), are both increased, while mitochondrial membrane potential is reduced. Moreover, preventing mitochondrial fusion in GSCs results in highly fragmented depolarized mitochondria, decreased BMP stemness signaling, impaired fatty acid metabolism, and GSC loss. Conversely, forcing mitochondrial elongation promotes GSC attachment to the niche. Importantly, maintenance of aging GSCs can be enhanced by suppressing Drp1 expression to prevent mitochondrial fission or treating with rapamycin, which is known to promote autophagy via TOR inhibition. Overall, our results show that mitochondrial dynamics are altered during physiological aging, affecting stem cell homeostasis via coordinated changes in stemness signaling, niche contact, and cellular metabolism. Such effects may also be highly relevant to other stem cell types and aging‐induced tissue degeneration.     

Myostatin induces mitochondrial metabolic alteration and typical apoptosis in cancer cells

Myostatin, a member of the transforming growth factor-β superfamily, regulates the glucose metabolism of muscle cells, while dysregulated myostatin activity is associated with a number of metabolic disorders, including muscle cachexia, obesity and type II diabetes. We observed that myostatin induced significant mitochondrial metabolic alterations and prolonged exposure of myostatin induced mitochondria-dependent apoptosis in cancer cells addicted to glycolysis. To address the underlying mechanism, we found that the protein levels of Hexokinase II (HKII) and voltage-dependent anion channel 1 (VDAC1), two key regulators of glucose metabolisms as well as metabolic stress-induced apoptosis, were negatively correlated. In particular, VDAC1 was dramatically upregulated in cells that are sensitive to myostatin treatment whereas HKII was downregulated and dissociated from mitochondria. Myostatin promoted the translocation of Bax from cytosol to mitochondria, and knockdown of VDAC1 inhibited myostatin-induced Bax translocation and apoptosis. These apoptotic changes can be partially rescued by repletion of ATP, or by ectopic expression of HKII, suggesting that perturbation of mitochondrial metabolism is causally linked with subsequent apoptosis. Our findings reveal novel function of myostatin in regulating mitochondrial metabolism and apoptosis in cancer cells.     

The loss of PGAM5 suppresses the mitochondrial degeneration caused by inactivation of PINK1 in Drosophila

PTEN-induced kinase 1 (PINK1), which is required for mitochondrial homeostasis, is a gene product responsible for early-onset Parkinson's disease (PD). Another early onset PD gene product, Parkin, has been suggested to function downstream of the PINK1 signalling pathway based on genetic studies in Drosophila. PINK1 is a serine/threonine kinase with a predicted mitochondrial target sequence and a probable transmembrane domain at the N-terminus, while Parkin is a RING-finger protein with ubiquitin-ligase (E3) activity. However, how PINK1 and Parkin regulate mitochondrial activity is largely unknown. To explore the molecular mechanism underlying the interaction between PINK1 and Parkin, we biochemically purified PINK1-binding proteins from human cultured cells and screened the genes encoding these binding proteins using Drosophila PINK1 (dPINK1) models to isolate a molecule(s) involved in the PINK1 pathology. Here we report that a PINK1-binding mitochondrial protein, PGAM5, modulates the PINK1 pathway. Loss of Drosophila PGAM5 (dPGAM5) can suppress the muscle degeneration, motor defects, and shorter lifespan that result from dPINK1 inactivation and that can be attributed to mitochondrial degeneration. However, dPGAM5 inactivation fails to modulate the phenotypes of parkin mutant flies. Conversely, ectopic expression of dPGAM5 exacerbated the dPINK1 and Drosophila parkin (dParkin) phenotypes. These results suggest that PGAM5 negatively regulates the PINK1 pathway related to maintenance of the mitochondria and, furthermore, that PGAM5 acts between PINK1 and Parkin, or functions independently of Parkin downstream of PINK1.     

Mitochondrial dysfunction and apoptosis in cumulus cells of type I diabetic mice

Impaired oocyte quality has been demonstrated in diabetic mice; however, the potential pathways by which maternal diabetes exerts its effects on the oocyte are poorly understood. Cumulus cells are in direct contact with the oocyte via gap junctions and provide essential nutrients to support oocyte development. In this study, we investigated the effects of maternal diabetes on the mitochondrial status in cumulus cells. We found an increased frequency of fragmented mitochondria, a decreased transmembrane potential and an aggregated distribution of mitochondria in cumulus cells from diabetic mice. Furthermore, while mitochondrial biogenesis in cumulus cells was induced by maternal diabetes, their metabolic function was disrupted as evidenced by lower ATP and citrate levels. Moreover, we present evidence suggesting that the mitochondrial impairments induced by maternal diabetes, at least in part, lead to cumulus cell apoptosis through the release of cytochrome c. Together the deleterious effects on cumulus cells may disrupt trophic and signaling interactions with the oocyte, contributing to oocyte incompetence and thus poor pregnancy outcomes in diabetic females.     

Is cancer a metabolic rebellion against host aging?: In the quest for immortality,tumor cells try to save themselves by boosting mitochondrial metabolism

Aging drives large systemic reductions in oxidative mitochondrial function, shifting the entire body metabolically toward aerobic glycolysis, a.k.a, the Warburg effect. Aging is also one of the most significant risk factors for the development of human cancers, including breast tumors. How are these two findings connected? One simplistic idea is that cancer cells rebel against the aging process by increasing their capacity for oxidative mitochondrial metabolism (OXPHOS). Then, local and systemic aerobic glycolysis in the aging host would provide energy-rich mitochondrial fuels (such as L-lactate and ketones) to directly “fuel” tumor cell growth and metastasis. This would establish a type of parasite-host relationship or “two-compartment tumor metabolism,” with glycolytic/oxidative metabolic coupling. The cancer cells (“the seeds”) would flourish in this nutrient-rich microenvironment (“the soil”), which has been fertilized by host aging. In this scenario, cancer cells are only trying to save themselves from the consequences of aging by engineering a metabolic mutiny, through the amplification of mitochondrial metabolism. We discuss the recent findings of Drs. Ron DePinho (MD Anderson) and Craig Thomspson (Sloan-Kettering) that are also consistent with this new hypothesis, linking cancer progression with metabolic aging. Using data mining and bioinformatics approaches, we also provide key evidence of a role for PGC1a/NRF1 signaling in the pathogenesis of (1) two-compartment tumor metabolism and (2) mitochondrial biogenesis in human breast cancer cells.Key words: aging, mitochondria, cancer metabolism, autophagy, mitophagy, aerobic glycolysis, oxidative phosphorylation, Metformin, drug resistance, chemoresistance, Warburg effect, metabolic compartments, parasite, PGC1a, PGC1b, NRF1, two-compartment tumor metabolism     

Pyrroline-5-carboxylate synthase senses cellular stress and modulates metabolism by regulating mitochondrial respiration

Pyrroline-5-carboxylate synthase (P5CS) catalyzes the synthesis of pyrroline-5-carboxylate (P5C), a key precursor for the synthesis of proline and ornithine. P5CS malfunction leads to multiple human diseases; however, the molecular mechanism underlying these diseases is unknown. We found that P5CS localizes in mitochondria in rod- and ring-like patterns but diffuses inside the mitochondria upon cellular starvation or exposure to oxidizing agents. Some of the human disease-related mutant forms of P5CS also exhibit diffused distribution. Multimerization (but not the catalytic activity) of P5CS regulates its localization. P5CS mutant cells have a reduced proliferation rate and are sensitive to cellular stresses. Flies lacking P5CS have reduced eclosion rates. Lipid droplets accumulate in the eyes of the newly eclosed P5CS mutant flies, which degenerate with aging. The loss of P5CS in cells leads to abnormal purine metabolism and lipid-droplet accumulation. The reduced lipid-droplet consumption is likely due to decreased expression of the fatty acid transporter, CPT1, and few β-oxidation-related genes following P5CS knockdown. Surprisingly, we found that P5CS is required for mitochondrial respiratory complex organization and that the respiration defects in P5CS knockout cells likely contribute to the metabolic defects in purine synthesis and lipid consumption. This study links amino acid synthesis with mitochondrial respiration and other key metabolic processes, whose imbalance might contribute to P5CS-related disease conditions.Subject terms: Enzyme mechanisms, Cell biology     

The membrane scaffold SLP2 anchors a proteolytic hub in mitochondria containing PARL and the i‐AAA protease YME1L

The SPFH (stomatin, prohibitin, flotillin, HflC/K) superfamily is composed of scaffold proteins that form ring‐like structures and locally specify the protein–lipid composition in a variety of cellular membranes. Stomatin‐like protein 2 (SLP2) is a member of this superfamily that localizes to the mitochondrial inner membrane (IM) where it acts as a membrane organizer. Here, we report that SLP2 anchors a large protease complex composed of the rhomboid protease PARL and the i‐AAA protease YME1L, which we term the SPY complex (for SLP2–PARL–YME1L). Association with SLP2 in the SPY complex regulates PARL‐mediated processing of PTEN‐induced kinase PINK1 and the phosphatase PGAM5 in mitochondria. Moreover, SLP2 inhibits the stress‐activated peptidase OMA1, which can bind to SLP2 and cleaves PGAM5 in depolarized mitochondria. SLP2 restricts OMA1‐mediated processing of the dynamin‐like GTPase OPA1 allowing stress‐induced mitochondrial hyperfusion under starvation conditions. Together, our results reveal an important role of SLP2 membrane scaffolds for the spatial organization of IM proteases regulating mitochondrial dynamics, quality control, and cell survival.     

Adaptations of Energy Metabolism Associated with Increased Levels of Mitochondrial Cholesterol in Niemann-Pick Type C1-deficient Cells

Niemann-Pick type C1 (NPC1) is a late endosomal transmembrane protein, which, together with NPC2 in the endosome lumen, mediates the transport of endosomal cholesterol to the plasma membrane and endoplasmic reticulum. Loss of function of NPC1 or NPC2 leads to cholesterol accumulation in late endosomes and causes neuronal dysfunction and neurodegeneration. Recent studies indicate that cholesterol also accumulates in mitochondria of NPC1-deficient cells and brain tissue and that NPC1 deficiency leads to alterations in mitochondrial function and energy metabolism. Here, we have investigated the effects of increased mitochondrial cholesterol levels on energy metabolism, using RNA interference to deplete Chinese hamster ovary cells of NPC1 alone or in combination with MLN64, which mediates endosomal cholesterol transport to mitochondria. Mitochondrial cholesterol levels were also altered by depletion of NPC2 in combination with the expression of NPC2 mutants. We found that the depletion of NPC1 increased lactate secretion, decreased glutamine-dependent mitochondrial respiration, and decreased ATP transport across mitochondrial membranes. These metabolic alterations did not occur when transport of endosomal cholesterol to mitochondria was blocked. In addition, the elevated mitochondrial cholesterol levels in NPC1-depleted cells and in NPC2-depleted cells expressing mutant NPC2 that allows endosomal cholesterol trafficking to mitochondria were associated with increased expression of the antioxidant response factor Nrf2. Antioxidant treatment not only prevented the increase in Nrf2 mRNA levels but also prevented the increased lactate secretion in NPC1-depleted cells. These results suggest that mitochondrial cholesterol accumulation can increase oxidative stress and in turn cause increased glycolysis to lactate and other metabolic alterations.     

Decline in cytochrome c oxidase activity in rat-brain mitochondria with aging. Role of peroxidized cardiolipin and beneficial effect of melatonin

Reactive oxygen species (ROS) are considered a key factor in mitochondrial dysfunction associated with brain aging process. Mitochondrial respiration is an important source of ROS and hence a potential contributor to brain functional changes with aging. In this study, we examined the effect of aging on cytochrome c oxidase activity and other bioenergetic processes such as oxygen consumption, membrane potential and ROS production in rat brain mitochondria. We found a significant age-dependent decline in the cytochrome c oxidase activity which was associated with parallel changes in state 3 respiration, membrane potential and with an increase in H₂O₂ generation. The cytochrome aa₃ content was practically unchanged in mitochondria from young and aged animals. The age-dependent decline of cytochrome c oxidase activity could be restored, in situ, to the level of young animals, by exogenously added cardiolipin. In addition, exposure of brain mitochondria to peroxidized cardiolipin resulted in an inactivation of this enzyme complex. It is suggested that oxidation/depletion of cardiolipin could be responsible, at least in part, for the decline of cytochrome c oxidase and mitochondrial dysfunction in brain aging. Melatonin treatment of old animals largely prevented the age-associated alterations of mitochondrial bioenergetic parameters. These results may prove useful in elucidating the molecular mechanisms underlying mitochondrial dysfunction associated with brain aging process, and may have implications in etiopathology of age-associated neurodegenerative disorders and in the development of potential treatment strategies.     

Mitochondrial translocation of Nur77 induced by ROS contributed to cardiomyocyte apoptosis in metabolic syndrome

Metabolic syndrome is a major risk factor for cardiovascular diseases, and increased cardiomyocyte apoptosis which contributes to cardiac dysfunction after myocardial ischemia/reperfusion (MI/R) injury. Nur77, a nuclear orphan receptor, is involved in such various cellular events as apoptosis, proliferation, and glucose and lipid metabolism in several cell types. Apoptosis is positively correlated with mitochondrial translocation of Nur77 in the cancer cells. However, the roles of Nur77 on cardiac myocytes in patients with metabolic syndrome remain unclear. The objective of this study was to determine whether Nur77 may contribute to cardiac apoptosis in patients with metabolic syndrome after I/R injury, and, if so, to identify the underlying molecular mechanisms responsible. We used leptin-deficient (ob/ob) mice to make metabolic syndrome models. In this report, we observed that, accompanied by the substantial decline in apoptosis inducer Nur77, MI/R induced cardiac dysfunction was manifested as cardiomyopathy and increased ROS. Using the neonatal rat cardiac myocytes cultured in a high-glucose and high-fat medium, we found that excessive H₂O₂ led to the significant alteration in mitochondrial membrane potential and translocation of Nur77 from the nucleus to the mitochondria. However, inhibition of the relocation of Nur77 to mitochondria via Cyclosporin A reversed the changes in membrane potential mediated by H₂O₂ and reduced myocardial cell injury. Therefore, these data provide a potential underlying mechanism for cardiac dysfunction in metabolic syndrome and the suppression of Nur77 translocation may provide an effective approach to reduce cardiac injury in the process.     

K-ras(G12V) transformation leads to mitochondrial dysfunction and a metabolic switch from oxidative phosphorylation to glycolysis

Increased aerobic glycolysis and oxidative stress are important features of cancer cell metabolism, but the underlying biochemical and molecular mechanisms remain elusive. Using a tetracycline inducible model, we show that activation of K-ras(G12V) causes mitochondrial dysfunction, leading to decreased respiration, elevated glycolysis, and increased generation of reactive oxygen species. The K-RAS protein is associated with mitochondria, and induces a rapid suppression of respiratory chain complex-I and a decrease in mitochondrial transmembrane potential by affecting the cyclosporin-sensitive permeability transition pore. Furthermore, pre-induction of K-ras(G12V) expression in vitro to allow metabolic adaptation to high glycolytic metabolism enhances the ability of the transformed cells to form tumor in vivo. Our study suggests that induction of mitochondrial dysfunction is an important mechanism by which K-ras(G12V) causes metabolic changes and ROS stress in cancer cells, and promotes tumor development.     

The role of compartmentalized signaling pathways in the control of mitochondrial activities in cancer cells

Mitochondria are the powerhouse organelles present in all eukaryotic cells. They play a fundamental role in cell respiration, survival and metabolism. Stimulation of G-protein coupled receptors (GPCRs) by dedicated ligands and consequent activation of the cAMP·PKA pathway finely couple energy production and metabolism to cell growth and survival. Compartmentalization of PKA signaling at mitochondria by A-Kinase Anchor Proteins (AKAPs) ensures efficient transduction of signals generated at the cell membrane to the organelles, controlling important aspects of mitochondrial biology. Emerging evidence implicates mitochondria as essential bioenergetic elements of cancer cells that promote and support tumor growth and metastasis. In this context, mitochondria provide the building blocks for cellular organelles, cytoskeleton and membranes, and supply all the metabolic needs for the expansion and dissemination of actively replicating cancer cells. Functional interference with mitochondrial activity deeply impacts on cancer cell survival and proliferation. Therefore, mitochondria represent valuable targets of novel therapeutic approaches for the treatment of cancer patients. Understanding the biology of mitochondria, uncovering the molecular mechanisms regulating mitochondrial activity andmapping the relevant metabolic and signaling networks operating in cancer cells will undoubtly contribute to create a molecular platform to be used for the treatment of proliferative disorders.Here, we will highlight the emerging roles of signaling pathways acting downstream to GPCRs and their intersection with the ubiquitin proteasome system in the control of mitochondrial activity in different aspects of cancer cell biology.