Cell-specific modulation of mitochondrial respiration and metabolism by the pro-apoptotic Bcl-2 family members Bax and Bak.

Proteins from the Bcl-2 family play an essential role in the regulation of apoptosis. However, they also possess cell death-unrelated activities that are less well understood. This prompted us to study apoptosis-unrelated activities of the Bax and Bak, pro-apoptotic members of the Bcl-2 family. We prepared Bax/Bak-deficient human cancer cells of different origin and found that while respiration in the glioblastoma U87 Bax/Bak-deficient cells was greatly enhanced, respiration of Bax/Bak-deficient B lymphoma HBL-2 cells was slightly suppressed. Bax/Bak-deficient U87 cells also proliferated faster in culture, formed tumours more rapidly in mice, and showed modulation of metabolism with a considerably increased NAD+/NADH ratio. Follow-up analyses documented increased/decreased expression of mitochondria-encoded subunits of respiratory complexes and stabilization/destabilization of the mitochondrial transcription elongation factor TEFM in Bax/Bak-deficient U87 and HBL-2 cells, respectively. TEFM downregulation using shRNAs attenuated mitochondrial respiration in Bax/Bak-deficient U87 as well as in parental HBL-2 cells. We propose that (post)translational regulation of TEFM levels in Bax/Bak-deficient cells modulates levels of subunits of mitochondrial respiratory complexes that, in turn, contribute to respiration and the accompanying changes in metabolism and proliferation in these cells.

Sperm mitochondria dysfunction in response to testicular cancer.

Testicular cancer is the most common form of cancer in young men of reproductive age and its incidence is increasing globally. With the currently successful treatment and 95% survival rate, there is a need for deeper understanding of testicular cancer-related infertility. Most patients with testicular cancer experience semen abnormalities prior to cancer therapy. However, the exact mechanism of the effect of testicular cancer on sperm anomalies is not known. Mitochondria are organelles that play a crucial role in both tumorigenesis and spermatogenesis and their malfunction may be an important factor resulting in sperm abnormalities in testicular cancer patients. Within the scope of this review, we will discuss current knowledge of testicular cancer-related alterations in the ATP production pathway, a possible pathophysiological switch from oxidative phosphorylation (OXPHOS) to glycolysis, as well as the role of oxidative stress promoting sperm dysfunction. In this regard, the review provides a summary of the impact of testicular cancer on sperm quality as a possible consequence of impaired mitochondrial function including the energy metabolic pathways that are known to be altered in the sperm of testicular cancer patients.

Mitochondrial HER2 stimulates respiration and promotes tumorigenicity.

BACKGROUND: Amplification of HER2, a receptor tyrosine kinase and a breast cancer-linked oncogene, is associated with aggressive disease. HER2 protein is localised mostly at the cell membrane, but a fraction translocates to mitochondria. Whether and how mitochondrial HER2 contributes to tumorigenicity is currently unknown. METHODS: We enriched the mitochondrial (mt-)HER2 fraction in breast cancer cells using an N-terminal mitochondrial targeting sequence and analysed how this manipulation impacts bioenergetics and tumorigenic properties. The role of the tyrosine kinase activity of mt-HER2 was assessed in wild type, kinase-dead (K753M) and kinase-enhanced (V659E) mtHER2 constructs. RESULTS: We document that mt-HER2 associates with the oxidative phosphorylation system, stimulates bioenergetics and promotes larger respiratory supercomplexes. mt-HER2 enhances proliferation and invasiveness in vitro and tumour growth and metastatic potential in vivo, in a kinase activity-dependent manner. On the other hand, constitutively active mt-HER2 provokes excessive mitochondria ROS generation, sensitises to cell death, and restricts growth of primary tumours, suggesting that regulation of HER2 activity in mitochondria is required for the maximal pro-tumorigenic effect. CONCLUSIONS: mt-HER2 promotes tumorigenicity by supporting bioenergetics and optimal redox balance.

Miro proteins connect mitochondrial function and intercellular transport.

Mitochondria are organelles present in most eukaryotic cells, where they play major and multifaceted roles. The classical notion of the main mitochondrial function as the powerhouse of the cell per se has been complemented by recent discoveries pointing to mitochondria as organelles affecting a number of other auxiliary processes. They go beyond the classical energy provision via acting as a relay point of many catabolic and anabolic processes, to signaling pathways critically affecting cell growth by their implication in de novo pyrimidine synthesis. These additional roles further underscore the importance of mitochondrial homeostasis in various tissues, where its deregulation promotes a number of pathologies. While it has long been known that mitochondria can move within a cell to sites where they are needed, recent research has uncovered that mitochondria can also move between cells. While this intriguing field of research is only emerging, it is clear that mobilization of mitochondria requires a complex apparatus that critically involves mitochondrial proteins of the Miro family, whose role goes beyond the mitochondrial transfer, as will be covered in this review.

Chemical Biopsy for GNMT as Noninvasive and Tumorigenesis-Relevant Diagnosis of Liver Cancer.

Early diagnosis of hepatocellular carcinoma (HCC) is difficult; the lack of convenient biomarker-based diagnostic modalities renders high-risk HCC patients burdened by life-long periodical examinations. Here, a new chemical biopsy approach was developed for noninvasive diagnosis of HCC using urine samples. Bioinformatic screening for tumor suppressors yielded glycine N-methyltransferase (GNMT) as a biomarker with clinical relevance to HCC tumorigenesis. A liquid chromatography-mass spectrometry (LC-MS)-based chemical biopsy detecting nonradioactive 13C-sarcosine from 13C-glycine was designed to noninvasively assess liver GNMT activity extrahepatically. 13C-Sarcosine showed a strong correlation with GNMT in normal and cancerous liver cells. In an autochthonous animal model developing visible cancer nodules at 17 weeks, the urinary 13C-sarcosine chemical biopsy exhibited notable changes as early as 8 weeks, showing significant correlations with liver GNMT and molecular pathological changes. Our chemical biopsy approach should facilitate early and noninvasive diagnosis of HCC, with direct relevance to tumorigenesis, which can be straightforwardly applied to other diseases.

Repurposing of MitoTam: Novel Anti-Cancer Drug Candidate Exhibits Potent Activity against Major Protozoan and Fungal Pathogens.

Many of the currently available anti-parasitic and anti-fungal frontline drugs have severe limitations, including adverse side effects, complex administration, and increasing occurrence of resistance. The discovery and development of new therapeutic agents is a costly and lengthy process. Therefore, repurposing drugs with already established clinical application offers an attractive, fast-track approach for novel treatment options. In this study, we show that the anti-cancer drug candidate MitoTam, a mitochondria-targeted analog of tamoxifen, efficiently eliminates a wide range of evolutionarily distinct pathogens in vitro, including pathogenic fungi, Plasmodium falciparum, and several species of trypanosomatid parasites, causative agents of debilitating neglected tropical diseases. MitoTam treatment was also effective in vivo and significantly reduced parasitemia of two medically important parasites, Leishmania mexicana and Trypanosoma brucei, in their respective animal infection models. Functional analysis in the bloodstream form of T. brucei showed that MitoTam rapidly altered mitochondrial functions, particularly affecting cellular respiration, lowering ATP levels, and dissipating mitochondrial membrane potential. Our data suggest that the mode of action of MitoTam involves disruption of the inner mitochondrial membrane, leading to rapid organelle depolarization and cell death. Altogether, MitoTam is an excellent candidate drug against several important pathogens, for which there are no efficient therapies and for which drug development is not a priority.

Mitochondrial Function as Related to Psychological Distress in Health Care Professionals.

OBJECTIVE: The present study evaluated the association of psychological distress and radiation exposure as a work-related stressor with mitochondrial function in health care professionals. METHODS: Health care professionals at a regional hospital in Italy were evaluated for physical health and psychological measures using self-report questionnaires (n = 41; mean age = 47.6 [13.1] years; 66% women). In a second sample, individuals exposed to elevated levels of ionizing radiation (IR; likely effective dose exceeding 6 mSv/y; n = 63, mean age = 45.8 [8.8] years; 62% women) were compared with health care workers with low IR (n = 57; mean age = 47.2 [9.5] years; 65% women) because exposure to a toxic agent might act as a (work-related) stressor. Associations were examined between psychological factors (12-item General Health Questionnaire, Perceived Stress Scale), work ability (Work Ability Index), and IR exposure at the workplace with markers of mitochondrial function, including mitochondrial redox activity, mitochondrial membrane potential, mitochondrial DNA (mtDNA) copy number, biogenesis, and mtDNA damage response measured from peripheral blood mononuclear cells. RESULTS: All participants were in good physical health. Individuals reporting high levels of psychological distress showed lower mitochondrial biogenesis as indicated by peroxisome proliferator-activated receptor-γ coactivator 1-α and lower nuclear factor erythroid 2-related factor 2 (NRF2) expression (2.5 [1.0] versus 1.0 [0.9] relative expression [rel exp], p = .035, and 31.5 [5.0] versus 19.4 [6.9] rel exp, p = .013, respectively). However, exposure to toxic agents (IR) was primarily associated with mitochondrial metabolism and reduced mtDNA integrity. Participants with IR exposure displayed higher mitochondrial redox activity (4480 [1202] mean fluorescence intensity [MFI]/min versus 3376 [983] MFI/min, p < .001) and lower mitochondrial membrane potential (0.89 [0.09] MFI versus 0.95 [0.11] MFI, p = .001), and reduced mtDNA integrity (1.18 [0.21] rel exp versus 3.48 [1.57] rel exp, p < .001) compared with nonexposed individuals. CONCLUSIONS: This study supports the notion that psychological distress and potential stressors related to toxic agents might influence various aspects of mitochondrial biology, and that chronic stress exposure can lead to molecular and functional recalibrations among mitochondria.

Quantitative analysis of neuronal mitochondrial movement reveals patterns resulting from neurotoxicity of rotenone and 6-hydroxydopamine.

Alterations in mitochondrial dynamics, including their intracellular trafficking, are common early manifestations of neuronal degeneration. However, current methodologies used to study mitochondrial trafficking events rely on parameters that are primarily altered in later stages of neurodegeneration. Our objective was to establish a reliable applied statistical analysis to detect early alterations in neuronal mitochondrial trafficking. We propose a novel quantitative analysis of mitochondria trajectories based on innovative movement descriptors, including straightness, efficiency, anisotropy, and kurtosis. We evaluated time- and dose-dependent alterations in trajectory descriptors using biological data from differentiated SH-SY5Y cells treated with the mitochondrial toxicants 6-hydroxydopamine and rotenone. MitoTracker Red CMXRos-labelled mitochondria movement was analyzed by total internal reflection fluorescence microscopy followed by computational modelling to describe the process. Based on the aforementioned trajectory descriptors, this innovative analysis of mitochondria trajectories provides insights into mitochondrial movement characteristics and can be a consistent and sensitive method to detect alterations in mitochondrial trafficking occurring in the earliest time points of neurodegeneration.

Oxidative stress and Rho GTPases in the biogenesis of tunnelling nanotubes: implications in disease and therapy.

Tunnelling nanotubes (TNTs) are an emerging route of long-range intercellular communication that mediate cell-to-cell exchange of cargo and organelles and contribute to maintaining cellular homeostasis by balancing diverse cellular stresses. Besides their role in intercellular communication, TNTs are implicated in several ways in health and disease. Transfer of pathogenic molecules or structures via TNTs can promote the progression of neurodegenerative diseases, cancer malignancy, and the spread of viral infection. Additionally, TNTs contribute to acquiring resistance to cancer therapy, probably via their ability to rescue cells by ameliorating various pathological stresses, such as oxidative stress, reactive oxygen species (ROS), mitochondrial dysfunction, and apoptotic stress. Moreover, mesenchymal stem cells play a crucial role in the rejuvenation of targeted cells with mitochondrial heteroplasmy and oxidative stress by transferring healthy mitochondria through TNTs. Recent research has focussed on uncovering the key regulatory molecules involved in the biogenesis of TNTs. However further work will be required to provide detailed understanding of TNT regulation. In this review, we discuss possible associations with Rho GTPases linked to oxidative stress and apoptotic signals in biogenesis pathways of TNTs and summarize how intercellular trafficking of cargo and organelles, including mitochondria, via TNTs plays a crucial role in disease progression and also in rejuvenation/therapy.

Mitochondrial Complex II: At the Crossroads.

Mitochondrial complex II (CII), also called succinate dehydrogenase (SDH), is a central purveyor of the reprogramming of metabolic and respiratory adaptation in response to various intrinsic and extrinsic stimuli and abnormalities. In this review we discuss recent findings regarding SDH biogenesis, which requires four known assembly factors, and modulation of its enzymatic activity by acetylation, succinylation, phosphorylation, and proteolysis. We further focus on the emerging role of both genetic and epigenetic aberrations leading to SDH dysfunction associated with various clinical manifestations. This review also covers the recent discovery of the role of SDH in inflammation-linked pathologies. Conceivably, SDH is a potential target for several hard-to-treat conditions, including cancer, that remains to be fully exploited.

Mitocans Revisited: Mitochondrial Targeting as Efficient Anti-Cancer Therapy.

Mitochondria are essential cellular organelles, controlling multiple signalling pathways critical for cell survival and cell death. Increasing evidence suggests that mitochondrial metabolism and functions are indispensable in tumorigenesis and cancer progression, rendering mitochondria and mitochondrial functions as plausible targets for anti-cancer therapeutics. In this review, we summarised the major strategies of selective targeting of mitochondria and their functions to combat cancer, including targeting mitochondrial metabolism, the electron transport chain and tricarboxylic acid cycle, mitochondrial redox signalling pathways, and ROS homeostasis. We highlight that delivering anti-cancer drugs into mitochondria exhibits enormous potential for future cancer therapeutic strategies, with a great advantage of potentially overcoming drug resistance. Mitocans, exemplified by mitochondrially targeted vitamin E succinate and tamoxifen (MitoTam), selectively target cancer cell mitochondria and efficiently kill multiple types of cancer cells by disrupting mitochondrial function, with MitoTam currently undergoing a clinical trial.

TRAIL induces apoptosis but not necroptosis in colorectal and pancreatic cancer cells preferentially via the TRAIL-R2/DR5 receptor.

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is a cytokine that can trigger apoptosis in many types of human cancer cells via engagement of its two pro-apoptotic receptors TRAIL-R1 (DR4) and TRAIL-R2 (DR5). TRAIL can also activate several other signaling pathways such as activation of stress kinases, canonical NF-κB signaling and necroptosis. Though both receptors are ubiquitously expressed, their relative participation in TRAIL-induced signaling is still largely unknown. To analyze TRAIL receptor-specific signaling, we prepared Strep-tagged, trimerized variants of recombinant human TRAIL with high affinity for either DR4 or DR5 receptor. Using these receptor-specific ligands, we examined the contribution of individual pro-apoptotic receptors to TRAIL-induced signaling pathways. We found that in TRAIL-resistant colorectal HT-29 cells but not in pancreatic PANC-1 cancer cells, DISC formation and initial caspase-8 processing proceeds comparably via both DR4- and DR5-activated signaling. TRAIL-induced apoptosis, enhanced by the inhibitor of the Bcl-2 family ABT-737, or by the translation inhibitor homoharringtonine, proceeded in both cell lines predominantly via the DR5 receptor. ShRNA-mediated downregulation of DR4 or DR5 receptors in HT-29 cells also pointed to a stronger contribution of DR5 in TRAIL-induced apoptosis. In contrast to apoptosis, necroptotic signaling was activated similarly by both DR4- or DR5-specific ligands. Activation of auxiliary signaling pathways involving NF-κB or stress kinases proceeded under apoptotic conditions mainly in a DR5-dependent manner, while these signaling pathways were during necroptosis similarly activated by either of these ligands. Our study provides the first systematic insight into DR4-/DR5-specific signaling in colorectal and pancreatic cancer cells.

Mitochondria-driven elimination of cancer and senescent cells.

Mitochondria and oxidative phosphorylation (OXPHOS) are emerging as intriguing targets for the efficient elimination of cancer cells. The specificity of this approach is aided by the capacity of non-proliferating non-cancerous cells to withstand oxidative insult induced by OXPHOS inhibition. Recently we discovered that mitochondrial targeting can also be employed to eliminate senescent cells, where it breaks the interplay between OXPHOS and ATP transporters that appear important for the maintenance of mitochondrial morphology and viability in the senescent setting. Hence, mitochondria/OXPHOS directed pharmacological interventions show promise in several clinically-relevant scenarios that call for selective removal of cancer and senescent cells.

MiR-126 in intestinal-type sinonasal adenocarcinomas: exosomal transfer of MiR-126 promotes anti-tumour responses.

BACKGROUND: Intestinal-type sinonasal adenocarcinomas (ITACs) are aggressive malignancies related to wood dust and leather exposure. ITACs are generally associated with advanced stage at presentation due to the insidious growth pattern and non-specific symptoms. Therefore, biomarkers that can detect the switch from the benign disease to malignancy are needed. Essential for tumour growth, angiogenesis is an important step in tumour development and progression. This process is strictly regulated, and MiR-126 considered its master modulator. METHODS: We have investigated MiR-126 levels in ITACs and compared them to benign sinonasal lesions, such as sinonasal-inverted papillomas (SIPs) and inflammatory polyps (NIPs). The tumour-suppressive functions of MiR-126 were also evaluated. RESULTS: We found that MiR-126 can significantly distinguish malignancy from benign nasal forms. The low levels of MiR-126 in ITACs point to its role in tumour progression. In this context, restoration of MiR-126 induced metabolic changes, and inhibited cell growth and the tumorigenic potential of MNSC cells. CONCLUSIONS: We report that MiR-126 delivered via exosomes from endothelial cells promotes anti-tumour responses. This paracrine transfer of MiRs may represent a new approach towards MiR-based therapy.

The Assembly Factor SDHAF2 Is Dispensable for Flavination of the Catalytic Subunit of Mitochondrial Complex II in Breast Cancer Cells.

Mitochondrial complex II or succinate dehydrogenase (SDH) is at the crossroads of oxidative phosphorylation and the tricarboxylic acid cycle. It has been shown that Sdh5 (SDHAF2/SDH5 in mammals) is required for flavination of the subunit Sdh1 (SDHA in human cells) in yeast. Here we demonstrate that in human breast cancer cells, SDHAF2/SDH5 is dispensable for SDHA flavination. In contrast to yeast, CRISPR-Cas9 nickase-mediated SDHAF2 KO breast cancer cells feature flavinated SDHA and retain fully assembled and functional complex II, as well as normal mitochondrial respiration. Our data show that SDHA flavination is independent of SDHAF2 in breast cancer cells, employing an alternative mechanism.

The mobility of mitochondria: Intercellular trafficking in health and disease.

The view that genes are constrained within somatic cells is challenged by in vitro evidence, and more recently by in vivo studies which demonstrate that mitochondria with their mitochondrial DNA (mtDNA) payload not only can, but do move between cells in tumour models and in mouse models of tissue damage. Using mouse tumour cell models without mtDNA to reflect mtDNA damage, we have shown that these cells grow tumours only after acquiring mtDNA from cells in the local microenvironment resulting in respiration recovery, tumorigenesis and metastasis. Mitochondrial transfer between cells has also been demonstrated following ischaemia-induced injury in the heart and brain and in lung epithelium, and following lung inflammation. In vitro investigations suggest that stem cells may be mitochondrial donors. The ability of mitochondria to move between cells appears to be an evolutionarily-conserved phenomenon, relevant to diseases with compromised mitochondrial function including neurodegenerative, neuromuscular and cardiovascular diseases as well as cancer and ageing.

The role of Her2 and other oncogenes of the PI3K/AKT pathway in mitochondria.

Altered metabolism and resistance to cell death are typical hallmarks of cancer phenotype. Mitochondria are organelles central to cellular metabolism as well as to cell death induction. Hyperactivation of pro-survival and pro-proliferative pathways such as PI3K/AKT leads to cancer initiation, which affects mitochondria. Growing body of evidence indicates that oncogenes such as HER2, EGFR and RAS, as well as the downstream members of the PI3K/AKT signaling pathway, directly regulate mitochondria by translocating to the organelle. Here we discuss evidence of this scenario and consider mechanisms for direct regulation of mitochondrial function. Being in close proximity to mitochondrial bioenergetics machinery as well as to the regulators/executors of programed cell death, oncogenes in mitochondria may be ideally placed to perform this task. This represents a thus far under-explored area, which may be relevant to better understanding of cancer initiation, progression and treatment.

MicroRNA in Metabolic Re-Programming and Their Role in Tumorigenesis.

The process of metabolic re-programing is linked to the activation of oncogenes and/or suppression of tumour suppressor genes, which are regulated by microRNAs (miRNAs). The interplay between oncogenic transformation-driven metabolic re-programming and modulation of aberrant miRNAs further established their critical role in the initiation, promotion and progression of cancer by creating a tumorigenesis-prone microenvironment, thus orchestrating processes of evasion to apoptosis, angiogenesis and invasion/migration, as well metastasis. Given the involvement of miRNAs in tumour development and their global deregulation, they may be perceived as biomarkers in cancer of therapeutic relevance.

Structural re-arrangement and peroxidase activation of cytochrome c by anionic analogues of vitamin E, tocopherol succinate and tocopherol phosphate.

Cytochrome c is a multifunctional hemoprotein in the mitochondrial intermembrane space whereby its participation in electron shuttling between respiratory complexes III and IV is alternative to its role in apoptosis as a peroxidase activated by interaction with cardiolipin (CL), and resulting in selective CL peroxidation. The switch from electron transfer to peroxidase function requires partial unfolding of the protein upon binding of CL, whose specific features combine negative charges of the two phosphate groups with four hydrophobic fatty acid residues. Assuming that other endogenous small molecule ligands with a hydrophobic chain and a negatively charged functionality may activate cytochrome c into a peroxidase, we investigated two hydrophobic anionic analogues of vitamin E, α-tocopherol succinate (α-TOS) and α-tocopherol phosphate (α-TOP), as potential inducers of peroxidase activity of cytochrome c. NMR studies and computational modeling indicate that they interact with cytochrome c at similar sites previously proposed for CL. Absorption spectroscopy showed that both analogues effectively disrupt the Fe-S(Met(80)) bond associated with unfolding of cytochrome c. We found that α-TOS and α-TOP stimulate peroxidase activity of cytochrome c. Enhanced peroxidase activity was also observed in isolated rat liver mitochondria incubated with α-TOS and tBOOH. A mitochondria-targeted derivative of TOS, triphenylphosphonium-TOS (mito-VES), was more efficient in inducing H2O2-dependent apoptosis in mouse embryonic cytochrome c(+/+) cells than in cytochrome c(-/-) cells. Essential for execution of the apoptotic program peroxidase activation of cytochrome c by α-TOS may contribute to its known anti-cancer pharmacological activity.