bims-mibica Biomed News
on Mitochondrial bioenergetics in cancer
Issue of 2023‒03‒12
forty papers selected by
Kelsey Fisher-Wellman
East Carolina University

  1. Elife. 2023 Mar 08. pii: e78654. [Epub ahead of print]12
      The oxidative tricarboxylic acid (TCA) cycle is a central mitochondrial pathway integrating catabolic conversions of NAD+ to NADH and anabolic production of aspartate, a key amino acid for cell proliferation. Several TCA cycle components are implicated in tumorigenesis, including loss of function mutations in subunits of succinate dehydrogenase (SDH), also known as complex II of the electron transport chain (ETC), but mechanistic understanding of how proliferating cells tolerate the metabolic defects of SDH loss is still lacking. Here, we identify that SDH supports human cell proliferation through aspartate synthesis but, unlike other ETC impairments, the effects of SDH inhibition are not ameliorated by electron acceptor supplementation. Interestingly, we find aspartate production and cell proliferation are restored to SDH-impaired cells by concomitant inhibition of ETC complex I (CI). We determine that the benefits of CI inhibition in this context depend on decreasing mitochondrial NAD+/NADH, which drives SDH-independent aspartate production through pyruvate carboxylation and reductive carboxylation of glutamine. We also find that genetic loss or restoration of SDH selects for cells with concordant CI activity, establishing distinct modalities of mitochondrial metabolism for maintaining aspartate synthesis. These data therefore identify a metabolically beneficial mechanism for CI loss in proliferating cells and reveal how compartmentalized redox changes can impact cellular fitness.
    Keywords:  biochemistry; cancer biology; chemical biology; human
  2. Elife. 2023 Mar 06. pii: e85494. [Epub ahead of print]12
      Recent studies reveal that lateral mitochondrial transfer, the movement of mitochondria from one cell to another, can affect cellular and tissue homeostasis1,2. Most of what we know about mitochondrial transfer stems from bulk cell studies and have led to the paradigm that functional transferred mitochondria restore bioenergetics and revitalize cellular functions to recipient cells with damaged or non-functional mitochondrial networks3. However, we show that mitochondrial transfer also occurs between cells with functioning endogenous mitochondrial networks, but the mechanisms underlying how transferred mitochondria can promote such sustained behavioral reprogramming remain unclear. We report that unexpectedly, transferred macrophage mitochondria are dysfunctional and accumulate reactive oxygen species in recipient cancer cells. We further discovered that reactive oxygen species accumulation activates ERK signaling, promoting cancer cell proliferation. Pro-tumorigenic macrophages exhibit fragmented mitochondrial networks, leading to higher rates of mitochondrial transfer to cancer cells. Finally, we observe that macrophage mitochondrial transfer promotes tumor cell proliferation in vivo. Collectively these results indicate that transferred macrophage mitochondria activate downstream signaling pathways in a ROS-dependent manner in cancer cells, and provide a model of how sustained behavioral reprogramming can be mediated by a relatively small amount of transferred mitochondria in vitro and in vivo.
    Keywords:  cancer biology; cell biology; human; mouse
  3. Chin J Nat Med. 2023 Feb;pii: S1875-5364(23)60391-7. [Epub ahead of print]21(2): 136-145
      Metabolic reprogramming, a newly recognized trait of tumor biology, is an intensively studied prospect for oncology medicines. For numerous tumors and cancer cell subpopulations, oxidative phosphorylation (OXPHOS) is essential for their biosynthetic and bioenergetic functions. Cancer cells with mutations in isocitrate dehydrogenase 1 (IDH1) exhibit differentiation arrest, epigenetic and transcriptional reprogramming, and sensitivity to mitochondrial OXPHOS inhibitors. In this study, we report that berberine, which is widely used in China to treat intestinal infections, acted solely at the mitochondrial electron transport chain (ETC) complex I, and that its association with IDH1 mutant inhibitor (IDH1mi) AG-120 decreased mitochondrial activity and enhanced antileukemic effect in vitro andin vivo. Our study gives a scientific rationale for the therapy of IDH1 mutant acute myeloid leukemia (AML) patients using combinatory mitochondrial targeted medicines, particularly those who are resistant to or relapsing from IDH1mi.
    Keywords:  Acute myeloid leukemia; Berberine; Complex I; Mutant IDH1; Oxidative phosphorylation
  4. Clin Transl Oncol. 2023 Mar 10.
      PURPOSE: Nowadays, the oxidative phosphorylation (OXPHOS) correlated with leukemogenesis and treatment response is extensive. Thus, exploration of novel approaches in disrupting OXPHOS in AML is urgently needed.MATERIALS AND METHODS: Bioinformatical analysis of TCGA AML dataset was performed to identify the molecular signaling of OXPHOS. The OXPHOS level was measured through a Seahorse XFe96 cell metabolic analyzer. Flow cytometry was applied to measure mitochondrial status. Real-time qPCR and western blot were used to analyze the expression of mitochondrial or inflammatory factors. MLL-AF9-induced leukemic mice were conducted to measure the anti-leukemia effect of chidamide.
    RESULTS: Here, we reported that AML patients with high OXPHOS level were in a poor prognosis, which was associated with high expression of HDAC1/3 (TCGA). Inhibition of HDAC1/3 by chidamide inhibited cell proliferation and induced apoptotic cell death in AML cells. Intriguingly, chidamide could disrupt mitochondrial OXPHOS as assessed by inducing mitochondrial superoxide and reducing oxygen consumption rate, as well as decreasing mitochondrial ATP production. We also observed that chidamide augmented HK1 expression, while glycolysis inhibitor 2-DG could reduce the elevation of HK1 and improve the sensitivity of AML cells exposed to chidamide. Furthermore, HDAC3 was correlated with hyperinflammatory status, while chidamide could downregulate the inflammatory signaling in AML. Notably, chidamide eradicated leukemic cells in vivo and prolonged the survival time of MLL-AF9-induced AML mice.
    CONCLUSION: Chidamide disrupted mitochondrial OXPHOS, promoted cell apoptosis and reduced inflammation in AML cells. These findings exhibited a novel mechanism that targeting OXPHOS would be a novel strategy for AML treatment.
    Keywords:  Acute myeloid leukemia; Chidamide; HDAC; Mitochondria; OXPHOS
  5. Geroscience. 2023 Mar 06.
      Dietary restriction (DR) increases lifespan in many organisms, but its underlying mechanisms are not fully understood. Mitochondria play a central role in metabolic regulation and are known to undergo changes in structure and function in response to DR. Mitochondrial membrane potential (Δψm) is the driving force for ATP production and mitochondrial outputs that integrate many cellular signals. One such signal regulated by Δψm is nutrient-status sensing. Here, we tested the hypothesis that DR promotes longevity through preserved Δψm during adulthood. Using the nematode Caenorhabditis elegans, we find that Δψm declines with age relatively early in the lifespan, and this decline is attenuated by DR. Pharmacologic depletion of Δψm blocked the longevity and health benefits of DR. Genetic perturbation of Δψm and mitochondrial ATP availability similarly prevented lifespan extension from DR. Taken together, this study provides further evidence that appropriate regulation of Δψm is a critical factor for health and longevity in response to DR.
    Keywords:  Aging; Bioenergetics; Calorie restriction; Fasting; Metabolism; Mitochondrial uncoupling
  6. Magn Reson Chem. 2023 Mar 07.
      Investigation of mitochondrial metabolism is gaining increased interest owing to the growing recognition of the role of mitochondria in health and numerous diseases. Studies of isolated mitochondria promise novel insights into the metabolism devoid of confounding effects from other cellular organelles such as cytoplasm. This study describes the isolation of mitochondria from mouse skeletal myoblast cells (C2C12) and the investigation of live mitochondrial metabolism in real time using isotope tracer-based NMR spectroscopy. [3-13 C1 ]pyruvate was used as the substrate to monitor the dynamic changes of the downstream metabolites in mitochondria. The results demonstrate an intriguing phenomenon, in which lactate is produced from pyruvate inside the mitochondria and the results were confirmed by treating mitochondria with an inhibitor of mitochondrial pyruvate carrier (UK5099). Lactate is associated with health and numerous diseases including cancer and, to date, it is known to occur only in the cytoplasm. The insight that lactate is also produced inside mitochondria opens avenues for exploring new pathways of lactate metabolism. Further, experiments performed using inhibitors of the mitochondrial respiratory chain, FCCP and rotenone, show that [2-13 C1 ]acetyl coenzyme A, which is produced from [3-13 C1 ]pyruvate and acts as a primary substrate for the tricarboxylic acid cycle in mitochondria, exhibits a remarkable sensitivity to the inhibitors. These results offer a direct approach to visualize mitochondrial respiration through altered levels of the associated metabolites.
    Keywords:  13C; 1H; NMR; [2-13C1]acetyl coenzyme A; [3-13C1]lactate; [3-13C1]pyruvate; live mitochondria; metabolism
  7. Nutr Cancer. 2023 Mar 07. 1-10
      Cisplatin is a common chemotherapeutic drug for treating ovarian cancer, but its clinical efficacy is hampered by intrinsic and acquired resistance. Previous studies had shown inhibiting oxidative phosphorylation overcomes cisplatin resistance in ovarian cancer. Studies reveal that bedaquiline, a clinically available antimicrobial drug, inhibits cancer via targeting mitochondria. This study systematically assessed the efficacy of bedaquiline in ovarian cancer and its underlying mechanism. Using a panel of ovarian cancer cell lines and normal ovary cells, we demonstrated bedaquiline is selective for anti-ovarian cancer activities. Furthermore, the sensitivity varied among different ovarian cancer cell lines regardless of their sensitivity to cisplatin. Bedaquiline inhibited growth, survival and migration, through decreasing levels of ATP synthase subunit, complex V activity, mitochondrial respiration and ATP. We further found that ovarian cancer displayed increased levels of ATP, oxygen consumption rate (OCR), complex V activity and ATP synthase subunits compared to normal counterpart. Combination index analysis showed that bedaquiline and cisplatin is synergistic. Bedaquiline remarkably enhanced the efficacy of cisplatin in inhibiting ovarian cancer growth in mice. Our study provides evidence to repurpose bedaquiline for ovarian cancer treatment and suggests that ATP synthase is a selective target to overcome cisplatin resistance in ovarian cancer.
  8. Cancer Res. 2023 Mar 09. pii: CAN-22-1999. [Epub ahead of print]
      Induction of ferroptosis, a recently defined form of nonapoptotic cell death caused by iron-dependent lipid peroxidation, has emerged as an anti-cancer strategy. Erastin is a ferroptosis activator that promotes cell death that not only depends on the depletion of cellular cysteine but also relies on mitochondrial oxidative metabolism of glutamine. Here, we demonstrate that ASS1, a key enzyme involved in the urea cycle, plays a crucial role in ferroptosis resistance. Loss of ASS1 increased the sensitivity of non-small cell lung cancer (NSCLC) cells to erastin in vitro and decreased tumor growth in vivo. Metabolomics analysis with stable isotope-labeled glutamine showed that ASS1 promotes reductive carboxylation of cytosolic glutamine and compromises the oxidative TCA cycle from glutamine anaplerosis, reducing mitochondrial-derived lipid reactive oxygen species. Moreover, transcriptome sequencing showed that ASS1 activates the mTORC1-SREBP1-SCD5 axis to promote de novo monounsaturated fatty acid synthesis by utilizing acetyl-CoA derived from the glutamine reductive pathway. Treating ASS1-deficient NSCLC cells with erastin combined with arginine deprivation significantly enhanced cell death compared to either treatment alone. Collectively, these results reveal a previously unknown regulatory role of ASS1 in ferroptosis resistance and provide a potential therapeutic target for ASS1-deficient NSCLC.
  9. Eur J Med Chem. 2023 Feb 24. pii: S0223-5234(23)00185-X. [Epub ahead of print]251 115219
      Mitochondrial complex I (CI) as a critical multifunctional respiratory complex of electron transport chain (ETC) in mitochondrial oxidative phosphorylation has been identified as vital and essence in ATP production, biosynthesis and redox balance. Recent progress in targeting CI has provided both insight and inspiration for oncotherapy, highlighting that the development of CI-targeting inhibitors is a promising therapeutic approach to fight cancer. Natural products possessing of ample scaffold diversity and structural complexity are the majority source of CI inhibitors, although low specificity and safety hinder their extensive application. Along with the gradual deepening in understanding of CI structure and function, significant progress has been achieved in exploiting novel and selective small molecules targeting CI. Among them, IACS-010759 had been approved by FDA for phase I trial in advanced cancers. Moreover, drug repurposing represents an effective and prospective strategy for CI inhibitor discovery. In this review, we mainly elaborate the biological function of CI in tumor progression, summarize the CI inhibitors reported in recent years and discuss the further perspectives for CI inhibitor application, expecting this work may provide insights into innovative discovery of CI-targeting drugs for cancer treatment.
    Keywords:  Cancer therapy; Inhibitors; Mitochondrial complex I
  10. Nature. 2023 Mar 08.
      Metabolic rewiring underlies the effector functions of macrophages1-3, but the mechanisms involved remain incompletely defined. Here, using unbiased metabolomics and stable isotope-assisted tracing, we show that an inflammatory aspartate-argininosuccinate shunt is induced following lipopolysaccharide stimulation. The shunt, supported by increased argininosuccinate synthase (ASS1) expression, also leads to increased cytosolic fumarate levels and fumarate-mediated protein succination. Pharmacological inhibition and genetic ablation of the tricarboxylic acid cycle enzyme fumarate hydratase (FH) further increases intracellular fumarate levels. Mitochondrial respiration is also suppressed and mitochondrial membrane potential increased. RNA sequencing and proteomics analyses demonstrate that there are strong inflammatory effects resulting from FH inhibition. Notably, acute FH inhibition suppresses interleukin-10 expression, which leads to increased tumour necrosis factor secretion, an effect recapitulated by fumarate esters. Moreover, FH inhibition, but not fumarate esters, increases interferon-β production through mechanisms that are driven by mitochondrial RNA (mtRNA) release and activation of the RNA sensors TLR7, RIG-I and MDA5. This effect is recapitulated endogenously when FH is suppressed following prolonged lipopolysaccharide stimulation. Furthermore, cells from patients with systemic lupus erythematosus also exhibit FH suppression, which indicates a potential pathogenic role for this process in human disease. We therefore identify a protective role for FH in maintaining appropriate macrophage cytokine and interferon responses.
  11. Biophys J. 2023 Mar 09. pii: S0006-3495(23)00164-9. [Epub ahead of print]
      Mitochondrial inner membrane potentials in cardiomyocytes may oscillate in cycles of depolarization/repolarization when the mitochondrial network is exposed to metabolic or oxidative stress. The frequencies of such oscillations are dynamically changing while clusters of weakly coupled mitochondrial oscillators adjust to a common phase and frequency. Across the cardiac myocyte, the averaged signal of the mitochondrial population follows self-similar or fractal dynamics; however, fractal properties of individual mitochondrial oscillators have not yet been examined. We show that the largest synchronously oscillating cluster exhibits a fractal dimension,D, that is indicative of self-similar behavior with D=1.27±0.11, in contrast to the remaining network mitochondria whose fractal dimension is close to that of Brownian noise,D=1.58±0.10 . We further demonstrate that fractal behavior is correlated with local coupling mechanisms, while it is only weakly linked to measures of functional connections between mitochondria. Our findings suggest that individual mitochondrial fractal dimensions may serve as a simple measure of local mitochondrial coupling.
    Keywords:  cardiac myocyte; fractal dimension; mitochondrial oscillator; wavelets
  12. Nature. 2023 Mar 08.
      Mutations in fumarate hydratase (FH) cause hereditary leiomyomatosis and renal cell carcinoma1. Loss of FH in the kidney elicits several oncogenic signalling cascades through the accumulation of the oncometabolite fumarate2. However, although the long-term consequences of FH loss have been described, the acute response has not so far been investigated. Here we generated an inducible mouse model to study the chronology of FH loss in the kidney. We show that loss of FH leads to early alterations of mitochondrial morphology and the release of mitochondrial DNA (mtDNA) into the cytosol, where it triggers the activation of the cyclic GMP-AMP synthase (cGAS)-stimulator of interferon genes (STING)-TANK-binding kinase 1 (TBK1) pathway and stimulates an inflammatory response that is also partially dependent on retinoic-acid-inducible gene I (RIG-I). Mechanistically, we show that this phenotype is mediated by fumarate and occurs selectively through mitochondrial-derived vesicles in a manner that depends on sorting nexin 9 (SNX9). These results reveal that increased levels of intracellular fumarate induce a remodelling of the mitochondrial network and the generation of mitochondrial-derived vesicles, which allows the release of mtDNAin the cytosol and subsequent activation of the innate immune response.
  13. Nature. 2023 Mar 08.
    Keywords:  Cell biology; Immunology; Metabolism
  14. MicroPubl Biol. 2023 ;2023
      The mitochondrial genome (mtDNA) is packaged into discrete protein-DNA complexes called nucleoids. mtDNA packaging factor TFAM (mitochondrial transcription factor-A) promotes nucleoid compaction and is required for mtDNA replication. Here, we investigate how changing TFAM levels affects mtDNA in the Caenorhabditis elegans germ line. We show that increasing germline TFAM activity boosts mtDNA number and significantly increases the relative proportion of a selfish mtDNA mutant, uaDf5 . We conclude that TFAM levels must be tightly controlled to ensure appropriate mtDNA composition in the germ line.
  15. Cells. 2023 Feb 22. pii: 695. [Epub ahead of print]12(5):
      Adenosine 5' triphosphate (ATP) is the energy currency of life, which is produced in mitochondria (~90%) and cytosol (less than 10%). Real-time effects of metabolic changes on cellular ATP dynamics remain indeterminate. Here we report the design and validation of a genetically encoded fluorescent ATP indicator that allows for real-time, simultaneous visualization of cytosolic and mitochondrial ATP in cultured cells. This dual-ATP indicator, called smacATPi (simultaneous mitochondrial and cytosolic ATP indicator), combines previously described individual cytosolic and mitochondrial ATP indicators. The use of smacATPi can help answer biological questions regarding ATP contents and dynamics in living cells. As expected, 2-deoxyglucose (2-DG, a glycolytic inhibitor) led to substantially decreased cytosolic ATP, and oligomycin (a complex V inhibitor) markedly decreased mitochondrial ATP in cultured HEK293T cells transfected with smacATPi. With the use of smacATPi, we can also observe that 2-DG treatment modestly attenuates mitochondrial ATP and oligomycin reduces cytosolic ATP, indicating the subsequent changes of compartmental ATP. To evaluate the role of ATP/ADP carrier (AAC) in ATP trafficking, we treated HEK293T cells with an AAC inhibitor, Atractyloside (ATR). ATR treatment attenuated cytosolic and mitochondrial ATP in normoxia, suggesting AAC inhibition reduces ADP import from the cytosol to mitochondria and ATP export from mitochondria to cytosol. In HEK293T cells subjected to hypoxia, ATR treatment increased mitochondrial ATP along with decreased cytosolic ATP, implicating that ACC inhibition during hypoxia sustains mitochondrial ATP but may not inhibit the reversed ATP import from the cytosol. Furthermore, both mitochondrial and cytosolic signals decrease when ATR is given in conjunction with 2-DG in hypoxia. Thus, real-time visualization of spatiotemporal ATP dynamics using smacATPi provides novel insights into how cytosolic and mitochondrial ATP signals respond to metabolic changes, providing a better understanding of cellular metabolism in health and disease.
    Keywords:  ATP; biosensor; fluorescence; metabolism; mitochondria
  16. Elife. 2023 Mar 07. pii: e68047. [Epub ahead of print]12
      Malonyl-CoA-acyl carrier protein transacylase (MCAT) is an enzyme involved in mitochondrial fatty acid synthesis (mtFAS) and catalyzes the transfer of the malonyl moiety of malonyl-CoA to the mitochondrial acyl carrier protein (ACP). Previously, we showed that loss-of-function of mtFAS genes, including Mcat, is associated with severe loss of electron transport chain (ETC) complexes in mouse immortalized skeletal myoblasts (Nowinski et al., 2020). Here, we report a proband presenting with hypotonia, failure to thrive, nystagmus, and abnormal brain MRI findings. Using whole exome sequencing, we identified biallelic variants in MCAT. Protein levels for NDUFB8 and COXII, subunits of complex I and IV respectively, were markedly reduced in lymphoblasts and fibroblasts, as well as SDHB for complex II in fibroblasts. ETC enzyme activities were decreased in parallel. Re-expression of wild-type MCAT rescued the phenotype in patient fibroblasts. This is the first report of a patient with MCAT pathogenic variants and combined oxidative phosphorylation deficiency.
    Keywords:  MCAT; genetics; genomics; human; mitochondria; mitochondrial disease
  17. Science. 2023 Mar 10. 379(6636): 996-1003
      Metabolic networks are interconnected and influence diverse cellular processes. The protein-metabolite interactions that mediate these networks are frequently low affinity and challenging to systematically discover. We developed mass spectrometry integrated with equilibrium dialysis for the discovery of allostery systematically (MIDAS) to identify such interactions. Analysis of 33 enzymes from human carbohydrate metabolism identified 830 protein-metabolite interactions, including known regulators, substrates, and products as well as previously unreported interactions. We functionally validated a subset of interactions, including the isoform-specific inhibition of lactate dehydrogenase by long-chain acyl-coenzyme A. Cell treatment with fatty acids caused a loss of pyruvate-lactate interconversion dependent on lactate dehydrogenase isoform expression. These protein-metabolite interactions may contribute to the dynamic, tissue-specific metabolic flexibility that enables growth and survival in an ever-changing nutrient environment.
  18. Cell Chem Biol. 2023 Mar 01. pii: S2451-9456(23)00055-7. [Epub ahead of print]
      Reactive oxygen species (ROS) can modulate protein function through cysteine oxidation. Identifying protein targets of ROS can provide insight into uncharacterized ROS-regulated pathways. Several redox-proteomic workflows, such as oxidative isotope-coded affinity tags (OxICAT), exist to identify sites of cysteine oxidation. However, determining ROS targets localized within subcellular compartments and ROS hotspots remains challenging with existing workflows. Here, we present a chemoproteomic platform, PL-OxICAT, which combines proximity labeling (PL) with OxICAT to monitor localized cysteine oxidation events. We show that TurboID-based PL-OxICAT can monitor cysteine oxidation events within subcellular compartments such as the mitochondrial matrix and intermembrane space. Furthermore, we use ascorbate peroxidase (APEX)-based PL-OxICAT to monitor oxidation events within ROS hotspots by using endogenous ROS as the source of peroxide for APEX activation. Together, these platforms further hone our ability to monitor cysteine oxidation events within specific subcellular locations and ROS hotspots and provide a deeper understanding of the protein targets of endogenous and exogenous ROS.
    Keywords:  APEX; OxICAT; TurboID; cysteine oxidation; proximity labeling; reactive oxygen species
  19. Mol Metab. 2023 Mar 04. pii: S2212-8778(23)00035-2. [Epub ahead of print] 101701
      OBJECTIVE: Emerging evidence suggest the existence of constant basal lipolysis and re-esterification of a substantial fraction of thus liberated fatty acids. In stimulated lipolysis, the re-esterification is proposed to be a protective mechanism against lipotoxicity; however, the role of the lipolysis coupled to re-esterification under basal conditions has not been deciphered.METHODS: We used adipocytes (in vitro differentiated brown and white adipocytes derived from a cell line or primary SVF culture) to study the effect of inhibition of re-esterification by pharmacological DGAT1 and DGAT2 inhibitors alone or in combination. We then evaluated cellular energetics, lipolysis flux, and lipidomic parameters along with mitochondrial properties and fuel utilization.
    RESULTS: In adipocytes, DGAT1 and 2 mediated re-esterification is a moderator of fatty acid oxidation. Combined inhibition of both DGATs (D1+2i) increases oxygen consumption, which is largely due to enhanced mitochondrial respiration by lipolysis-derived fatty acids (FAs). Acute D1+2i selectively affects mitochondrial respiration without affecting the transcriptional homeostasis of genes relevant to mitochondrial health and lipid metabolism. D1+2i enhances the mitochondrial import of pyruvate and activates AMP Kinase to counteract CPT1 antagonism, thus facilitating the mitochondrial import of fatty acyl-CoA.
    CONCLUSIONS: These data implicate the process of re-esterification in the regulation of mitochondrial FA usage and uncover a mechanism of FAO regulation via crosstalk with FA re-esterification.
    Keywords:  Adipose tissue; DGAT1; DGAT2; Fatty acid oxidation; Lipidomics; Lipolysis; Re-esterification
  20. FEBS J. 2023 Mar 04.
      Radiation resistance is the leading cause of radiotherapy failure in patients with cancer. Enhanced DNA damage repair is the main reason for cancer cells to develop resistance to radiation. Autophagy has been widely reported to be linked to increased genome stability and radiation resistance. Mitochondria are highly involved in cell response to radiotherapy. However, the autophagy subtype mitophagy has not been studied in terms of genome stability. We have previously demonstrated that mitochondrial dysfunction is the cause of radiation resistance in tumor cells. We found in this study that SIRT3 was highly expressed in colorectal cancer cells with mitochondrial dysfunction, leading to PINK1/Parkin-mediated mitophagy. Excessive activation of mitophagy enhanced DNA damage repair, therefore promoting the resistance of tumor cells to radiation. Mechanistically, mitophagy resulted in decreased RING1b expression, which led to a reduction in the ubiquitination of histone H2A at K119, thereby enhancing the repair of DNA damage caused by radiation. Additionally, high expression of SIRT3 was related to a poor tumor regression grade in rectal cancer patients treated with neoadjuvant radiotherapy. These findings suggest that restoring mitochondrial function could be an effective method for increasing the radiosensitivity of patients with colorectal cancer.
    Keywords:  Cancer cells; DNA damage repair; Mitochondrial dysfunction; Radiation; SIRT3
  21. Mol Cell. 2023 Feb 28. pii: S1097-2765(23)00115-6. [Epub ahead of print]
      Mitochondria are not only central organelles in metabolism and energy conversion but are also platforms for cellular signaling cascades. Classically, the shape and ultrastructure of mitochondria were depicted as static. The discovery of morphological transitions during cell death and of conserved genes controlling mitochondrial fusion and fission contributed to establishing the concept that mitochondrial morphology and ultrastructure are dynamically regulated by mitochondria-shaping proteins. These finely tuned, dynamic changes in mitochondrial shape can in turn control mitochondrial function, and their alterations in human diseases suggest that this space can be explored for drug discovery. Here, we review the basic tenets and molecular mechanisms of mitochondrial morphology and ultrastructure, describing how they can coordinately define mitochondrial function.
    Keywords:  cristae; cristae remodeling; fusion fission; mitochondria
  22. Front Cell Dev Biol. 2023 ;11 1124164
      Acute lymphoblastic leukemia (aLL) is a malignant cancer in the blood and bone marrow characterized by rapid expansion of lymphoblasts. It is a common pediatric cancer and the principal basis of cancer death in children. Previously, we reported that L-asparaginase, a key component of acute lymphoblastic leukemia chemotherapy, causes IP3R-mediated ER Ca2+ release, which contributes to a fatal rise in [Ca2+]cyt, eliciting aLL cell apoptosis via upregulation of the Ca2+-regulated caspase pathway (Blood, 133, 2222-2232). However, the cellular events leading to the rise in [Ca2+]cyt following L-asparaginase-induced ER Ca2+ release remain obscure. Here, we show that in acute lymphoblastic leukemia cells, L-asparaginase causes mitochondrial permeability transition pore (mPTP) formation that is dependent on IP3R-mediated ER Ca2+ release. This is substantiated by the lack of L-asparaginase-induced ER Ca2+ release and loss of mitochondrial permeability transition pore formation in cells depleted of HAP1, a key component of the functional IP3R/HAP1/Htt ER Ca2+ channel. L-asparaginase induces ER Ca2+ transfer into mitochondria, which evokes an increase in reactive oxygen species (ROS) level. L-asparaginase-induced rise in mitochondrial Ca2+ and reactive oxygen species production cause mitochondrial permeability transition pore formation that then leads to an increase in [Ca2+]cyt. Such rise in [Ca2+]cyt is inhibited by Ruthenium red (RuR), an inhibitor of the mitochondrial calcium uniporter (MCU) that is required for mitochondrial Ca2+ uptake, and cyclosporine A (CsA), an mitochondrial permeability transition pore inhibitor. Blocking ER-mitochondria Ca2+ transfer, mitochondrial ROS production, and/or mitochondrial permeability transition pore formation inhibit L-asparaginase-induced apoptosis. Taken together, these findings fill in the gaps in our understanding of the Ca2+-mediated mechanisms behind L-asparaginase-induced apoptosis in acute lymphoblastic leukemia cells.
    Keywords:  L-asparaginase; acute lymphoblastic leukemia; blood-related disorders; chemotherapy; leukemia
  23. Cancers (Basel). 2023 Feb 21. pii: 1370. [Epub ahead of print]15(5):
      Osteosarcoma is considered the most common bone tumor affecting children and young adults. The standard of care is chemotherapy; however, the onset of drug resistance still jeopardizes osteosarcoma patients, thus making it necessary to conduct a thorough investigation of the possible mechanisms behind this phenomenon. In the last decades, metabolic rewiring of cancer cells has been proposed as a cause of chemotherapy resistance. Our aim was to compare the mitochondrial phenotype of sensitive osteosarcoma cells (HOS and MG-63) versus their clones when continuously exposed to doxorubicin (resistant cells) and identify alterations exploitable for pharmacological approaches to overcome chemotherapy resistance. Compared with sensitive cells, doxorubicin-resistant clones showed sustained viability with less oxygen-dependent metabolisms, and significantly reduced mitochondrial membrane potential, mitochondrial mass, and ROS production. In addition, we found reduced expression of TFAM gene generally associated with mitochondrial biogenesis. Finally, combined treatment of resistant osteosarcoma cells with doxorubicin and quercetin, a known inducer of mitochondrial biogenesis, re-sensitizes the doxorubicin effect in resistant cells. Despite further investigations being needed, these results pave the way for the use of mitochondrial inducers as a promising strategy to re-sensitize doxorubicin cytotoxicity in patients who do not respond to therapy or reduce doxorubicin side effects.
    Keywords:  cancer; chemotherapy; doxorubicin; drug resistance; metabolism; mitochondria; osteosarcoma; targeting mitochondrial alterations
  24. Cell Metab. 2023 Mar 06. pii: S1550-4131(23)00049-9. [Epub ahead of print]
      The metabolic state represents a major hurdle for an effective adoptive T cell therapy (ACT). Indeed, specific lipids can harm CD8+ T cell (CTL) mitochondrial integrity, leading to defective antitumor responses. However, the extent to which lipids can affect the CTL functions and fate remains unexplored. Here, we show that linoleic acid (LA) is a major positive regulator of CTL activity by improving metabolic fitness, preventing exhaustion, and stimulating a memory-like phenotype with superior effector functions. We report that LA treatment enhances the formation of ER-mitochondria contacts (MERC), which in turn promotes calcium (Ca2+) signaling, mitochondrial energetics, and CTL effector functions. As a direct consequence, the antitumor potency of LA-instructed CD8 T cells is superior in vitro and in vivo. We thus propose LA treatment as an ACT potentiator in tumor therapy.
    Keywords:  CD8 T cells; adoptive T cell therapy; linoleic acid; lipid metabolism; metabolic fitness
  25. J Transl Med. 2023 Mar 10. 21(1): 189
      BACKGROUND: The inner membrane mitochondrial protein (IMMT) is a central unit of the mitochondrial contact site and cristae organizing system (MICOS). While researchers continue to demonstrate the physiological function of IMMT in regulating mitochondrial dynamics and preserving mitochondrial structural integrity, the roles of IMMT in clinicopathology, the tumor immune microenvironment (TIME), and precision oncology in breast cancer (BC) remain unclear.METHODS: Multi-omics analysis was used here to evaluate the diagnostic and prognostic value of IMMT. Web applications aimed at analyzing the whole tumor tissue, single cells, and spatial transcriptomics were used to examine the relationship of IMMT with TIME. Gene set enrichment analysis (GSEA) was employed to determine the primary biological impact of IMMT. Experimental verification using siRNA knockdown and clinical specimens of BC patients confirmed the mechanisms behind IMMT on BC cells and the clinical significance, respectively. Potent drugs were identified by accessing the data repositories of CRISPR-based drug screenings.
    RESULTS: High IMMT expression served as an independent diagnostic biomarker, correlated with advanced clinical status, and indicated a poor relapse-free survival (RFS) rate for patients with BC. Although, the contents of Th1, Th2, MSC, macrophages, basophil, CD4 + T cell and B cell, and TMB levels counteracted the prognostic significance. Single-cell level and whole-tissue level analyses revealed that high IMMT was associated with an immunosuppressive TIME. GSEA identified IMMT perturbation as involved in cell cycle progression and mitochondrial antioxidant defenses. Experimental knockdown of IMMT impeded the migration and viability of BC cells, arrested the cell cycle, disturbed mitochondrial function, and increased the ROS level and lipid peroxidation. The clinical values of IMMT were amenable to ethnic Chinese BC patients, and can be extrapolated to some other cancer types. Furthermore, we discovered that pyridostatin acted as a potent drug candidate in BC cells harboring an elevated IMMT expression.
    CONCLUSION: This study combined a multi-omics survey with experimental verification to reveal the novel clinical significance of IMMT in BC, demonstrating its role in TIME, cancer cell growth and mitochondrial fitness, and identified pyridostatin as a promising drug candidate for the development of precision medicine.
    Keywords:  Breast cancer; Diagnostic biomarker; Inner membrane mitochondrial protein; Precision medicine; Prognosis; Tumor immune microenvironment
  26. EMBO J. 2023 Mar 10. e113033
      Mitophagy is a fundamental quality control mechanism of mitochondria. Its regulatory mechanisms and pathological implications remain poorly understood. Here, via a mitochondria-targeted genetic screen, we found that knockout (KO) of FBXL4, a mitochondrial disease gene, hyperactivates mitophagy at basal conditions. Subsequent counter screen revealed that FBXL4-KO hyperactivates mitophagy via two mitophagy receptors BNIP3 and NIX. We determined that FBXL4 functions as an integral outer-membrane protein that forms an SCF-FBXL4 ubiquitin E3 ligase complex. SCF-FBXL4 ubiquitinates BNIP3 and NIX to target them for degradation. Pathogenic FBXL4 mutations disrupt SCF-FBXL4 assembly and impair substrate degradation. Fbxl4-/- mice exhibit elevated BNIP3 and NIX proteins, hyperactive mitophagy, and perinatal lethality. Importantly, knockout of either Bnip3 or Nix rescues metabolic derangements and viability of the Fbxl4-/- mice. Together, beyond identifying SCF-FBXL4 as a novel mitochondrial ubiquitin E3 ligase restraining basal mitophagy, our results reveal hyperactivated mitophagy as a cause of mitochondrial disease and suggest therapeutic strategies.
    Keywords:  BNIP3/NIX; FBXL4; mitochondrial disease; mitophagy; ubiquitin-proteasome pathway
  27. Blood. 2023 Mar 08. pii: blood.2022018711. [Epub ahead of print]
      Mitochondrial damage-associated molecular patterns (mtDAMPs) include proteins, lipids, metabolites and DNA and have various context specific immunoregulatory functions. Cell-free mitochondrial DNA (mtDNA) is recognised via pattern recognition receptors and is a potent activator of the innate immune system. Cell-free mtDNA is elevated in the circulation of trauma and cancer patients, however the functional consequences of elevated mtDNA are largely undefined. Multiple myeloma (MM) relies upon cellular interactions within the bone marrow (BM) microenvironment for survival and progression. Here, using in-vivo models, we describe the role of MM cell derived mtDAMPs in the pro-tumoral BM microenvironment, and the mechanism and functional consequence of mtDAMPs in myeloma disease progression. Initially, we identified elevated levels of mtDNA in the peripheral blood serum of MM patients compared to healthy controls. Using the MM1S cells engrafted into NSG mice we established that elevated mtDNA was derived from MM cells. We further show that BM macrophages sense and respond to mtDAMPs through the STING pathway and inhibition of this pathway reduces MM tumor-burden in the KaLwRij-5TGM1 mouse model. Moreover, we found that MM derived mtDAMPs induced upregulation of chemokine signatures in BM macrophages and inhibition of this signature resulted in egress of MM cells from the BM. Here, we demonstrate that malignant plasma cells release mtDNA, a form of mtDAMPs, into the myeloma BM microenvironment, which in turn activates macrophages via STING signalling. We establish the functional role of these mtDAMP-activated macrophages in promoting disease progression and retaining MM cells in the pro-tumoral BM microenvironment.
  28. Cancers (Basel). 2023 Feb 23. pii: 1417. [Epub ahead of print]15(5):
      Lactic acidosis, a hallmark of solid tumour microenvironment, originates from lactate hyperproduction and its co-secretion with protons by cancer cells displaying the Warburg effect. Long considered a side effect of cancer metabolism, lactic acidosis is now known to play a major role in tumour physiology, aggressiveness and treatment efficiency. Growing evidence shows that it promotes cancer cell resistance to glucose deprivation, a common feature of tumours. Here we review the current understanding of how extracellular lactate and acidosis, acting as a combination of enzymatic inhibitors, signal, and nutrient, switch cancer cell metabolism from the Warburg effect to an oxidative metabolic phenotype, which allows cancer cells to withstand glucose deprivation, and makes lactic acidosis a promising anticancer target. We also discuss how the evidence about lactic acidosis' effect could be integrated in the understanding of the whole-tumour metabolism and what perspectives it opens up for future research.
    Keywords:  Warburg effect; glucose deprivation; lactic acidosis; metabolic symbiosis; tumour heterogeneity
  29. Mol Cell. 2023 Mar 06. pii: S1097-2765(23)00116-8. [Epub ahead of print]
      Oxygen is toxic across all three domains of life. Yet, the underlying molecular mechanisms remain largely unknown. Here, we systematically investigate the major cellular pathways affected by excess molecular oxygen. We find that hyperoxia destabilizes a specific subset of Fe-S cluster (ISC)-containing proteins, resulting in impaired diphthamide synthesis, purine metabolism, nucleotide excision repair, and electron transport chain (ETC) function. Our findings translate to primary human lung cells and a mouse model of pulmonary oxygen toxicity. We demonstrate that the ETC is the most vulnerable to damage, resulting in decreased mitochondrial oxygen consumption. This leads to further tissue hyperoxia and cyclic damage of the additional ISC-containing pathways. In support of this model, primary ETC dysfunction in the Ndufs4 KO mouse model causes lung tissue hyperoxia and dramatically increases sensitivity to hyperoxia-mediated ISC damage. This work has important implications for hyperoxia pathologies, including bronchopulmonary dysplasia, ischemia-reperfusion injury, aging, and mitochondrial disorders.
    Keywords:  DNA damage; Fe-S clusters; hyperoxia; lung injury; mitochondria; oxygen; purine synthesis; redox; translation fidelity
  30. Nat Aging. 2023 Feb;3(2): 157-161
      Mitochondrial dysfunction plays a central role in aging but the exact biological causes are still being determined. Here, we show that optogenetically increasing mitochondrial membrane potential during adulthood using a light-activated proton pump improves age-associated phenotypes and extends lifespan in C. elegans. Our findings provide direct causal evidence that rescuing the age-related decline in mitochondrial membrane potential is sufficient to slow the rate of aging and extend healthspan and lifespan.
  31. Cancer Res Commun. 2022 Mar;2(3): 182-201
      Deregulated expression of MYC family oncogenes occurs frequently in human cancer and is often associated with aggressive disease and poor prognosis. While MYC is a highly warranted target, it has been considered "undruggable," and no specific anti-MYC drugs are available in the clinic. We recently identified molecules named MYCMIs that inhibit the interaction between MYC and its essential partner MAX. Here we show that one of these molecules, MYCMI-7, efficiently and selectively inhibits MYC:MAX and MYCN:MAX interactions in cells, binds directly to recombinant MYC, and reduces MYC-driven transcription. In addition, MYCMI-7 induces degradation of MYC and MYCN proteins. MYCMI-7 potently induces growth arrest/apoptosis in tumor cells in a MYC/MYCN-dependent manner and downregulates the MYC pathway on a global level as determined by RNA sequencing. Sensitivity to MYCMI-7 correlates with MYC expression in a panel of 60 tumor cell lines and MYCMI-7 shows high efficacy toward a collection of patient-derived primary glioblastoma and acute myeloid leukemia (AML) ex vivo cultures. Importantly, a variety of normal cells become G1 arrested without signs of apoptosis upon MYCMI-7 treatment. Finally, in mouse tumor models of MYC-driven AML, breast cancer, and MYCN-amplified neuroblastoma, treatment with MYCMI-7 downregulates MYC/MYCN, inhibits tumor growth, and prolongs survival through apoptosis with few side effects. In conclusion, MYCMI-7 is a potent and selective MYC inhibitor that is highly relevant for the development into clinically useful drugs for the treatment of MYC-driven cancer.Significance: Our findings demonstrate that the small-molecule MYCMI-7 binds MYC and inhibits interaction between MYC and MAX, thereby hampering MYC-driven tumor cell growth in culture and in vivo while sparing normal cells.
  32. Cell Discov. 2023 Mar 07. 9(1): 26
      Metabolic reprogramming is a hallmark of cancer. However, it is not well known how metabolism affects cancer progression. We identified that metabolic enzyme acyl-CoA oxidase 1 (ACOX1) suppresses colorectal cancer (CRC) progression by regulating palmitic acid (PA) reprogramming. ACOX1 is highly downregulated in CRC, which predicts poor clinical outcome in CRC patients. Functionally, ACOX1 depletion promotes CRC cell proliferation in vitro and colorectal tumorigenesis in mouse models, whereas ACOX1 overexpression inhibits patient-derived xenograft growth. Mechanistically, DUSP14 dephosphorylates ACOX1 at serine 26, promoting its polyubiquitination and proteasomal degradation, thereby leading to an increase of the ACOX1 substrate PA. Accumulated PA promotes β-catenin cysteine 466 palmitoylation, which inhibits CK1- and GSK3-directed phosphorylation of β-catenin and subsequent β-Trcp-mediated proteasomal degradation. In return, stabilized β-catenin directly represses ACOX1 transcription and indirectly activates DUSP14 transcription by upregulating c-Myc, a typical target of β-catenin. Finally, we confirmed that the DUSP14-ACOX1-PA-β-catenin axis is dysregulated in clinical CRC samples. Together, these results identify ACOX1 as a tumor suppressor, the downregulation of which increases PA-mediated β-catenin palmitoylation and stabilization and hyperactivates β-catenin signaling thus promoting CRC progression. Particularly, targeting β-catenin palmitoylation by 2-bromopalmitate (2-BP) can efficiently inhibit β-catenin-dependent tumor growth in vivo, and pharmacological inhibition of DUSP14-ACOX1-β-catenin axis by Nu-7441 reduced the viability of CRC cells. Our results reveal an unexpected role of PA reprogramming induced by dephosphorylation of ACOX1 in activating β-catenin signaling and promoting cancer progression, and propose the inhibition of the dephosphorylation of ACOX1 by DUSP14 or β-catenin palmitoylation as a viable option for CRC treatment.
  33. Comput Struct Biotechnol J. 2023 ;21 1606-1620
      Short-chain fatty acids (SCFAs) exhibit anticancer activity in cellular and animal models of colon cancer. Acetate, propionate, and butyrate are the three major SCFAs produced from dietary fiber by gut microbiota fermentation and have beneficial effects on human health. Most previous studies on the antitumor mechanisms of SCFAs have focused on specific metabolites or genes involved in antitumor pathways, such as reactive oxygen species (ROS) biosynthesis. In this study, we performed a systematic and unbiased analysis of the effects of acetate, propionate, and butyrate on ROS levels and metabolic and transcriptomic signatures at physiological concentrations in human colorectal adenocarcinoma cells. We observed significantly elevated levels of ROS in the treated cells. Furthermore, significantly regulated signatures were involved in overlapping pathways at metabolic and transcriptomic levels, including ROS response and metabolism, fatty acid transport and metabolism, glucose response and metabolism, mitochondrial transport and respiratory chain complex, one-carbon metabolism, amino acid transport and metabolism, and glutaminolysis, which are directly or indirectly linked to ROS production. Additionally, metabolic and transcriptomic regulation occurred in a SCFAs types-dependent manner, with an increasing degree from acetate to propionate and then to butyrate. This study provides a comprehensive analysis of how SCFAs induce ROS production and modulate metabolic and transcriptomic levels in colon cancer cells, which is vital for understanding the mechanisms of the effects of SCFAs on antitumor activity in colon cancer.
    Keywords:  1H–13C HMBC, 1H–13C Heteronuclear Multiple Bond Correlation Spectroscopy; 1H–13C HSQC, 1H–13C Heteronuclear Single Quantum Coherence Spectroscopy; 1H–1H COSY, 1H–1H Correlation Spectroscopy; 1H–1H TOCSY, 1H–1H Total Correlation Spectroscopy; ADP, Adenosine diphosphate; AMP, Adenosine monophosphate; ATP, Adenosine triphosphate; Ace, Acetate; Ach, Acetylcholine; Ala, Alanine; CRC, Colorectal Cancer; Caco-2, Human Colon Adenocarcinoma; Cho, Choline; CoA, Coenzyme A; Cre, Creatine; DCFH-DA, Dichloro-Dihydro-Fluorescein Diacetate; DEGs, Differentially Expressed Genes; DMEM, Dulbecco's Modified Eagle Medium; DMG, Dimethylglycine; DNA, Deoxyribonucleic Acid; EP, Eppendorf; FA, Formate; FDR, False Discovery Rate; Fru, Fructose; Fum, Fumaric acid; GLS, Glutaminase; GSEA, Gene Set Enrichment Analysis; GSH, Glutathione; Gal-1-P, Galactose-1-phosphate; Glc, Glucose; Gln, Glutamine; Glu, Glutamate; Gly, Glycine; HCT116, Human Colorectal Carcinoma Cell Line; HEK, Human Embryonic Kidney cells; HT29, Human Colorectal Adenocarcinoma Cell Line with Epithelial Morphology; His, Histidine; Ile, Isoleucine; J-Res, J-resolved Spectroscopy; LDH, Lactate Dehydrogenase; Lac, Lactate; Leu, Leucine; Lys, Lysine; MCF-7, Human Breast Cancer Cell Line with Estrogen; MCT, Monocarboxylate Transporters; Met, Methionine; MetS, Metabolic Syndrome; Mitochondrial function; NAD+, Nicotinamide adenine dinucleotide; NAG, N-Acetyl-L-Glutamine; NMR, Nuclear Magnetic Resonance; NMR-based Metabolomics; NOESY, Nuclear Overhauser Effect Spectroscopy; O-PLS-DA, Orthogonal Projection to the Latent Structures Discriminant Analysis; PA, Pantothenate; PC, Phosphocholine; PCA, Principal Component Analysis; PDC, Pyruvate Decarboxylase; PDK, Pyruvate Dehydrogenase Kinase; PKC, Protein Kinase C; PPP, Pentose Phosphate Pathway; Phe, Phenylalanine; Pyr, Pyruvate; RNA, Ribonucleic Acid; ROS, Reactive Oxygen Species; RPKM, Reads per Kilobase of Transcript per Million Reads Mapped; Reactive oxygen species; SCFAs, Short Chain Fatty Acids; SLC, Solute-Carrier Genes; Short-chain fatty acids; Suc, Succinate; T2DM, Type 2 Diabetes; TCA, Tricarboxylic Acid; Tau, Taurine; Thr, Threonine; Transcriptomics; Tyr, Tyrosine; UDP, Uridine 5′-diphosphate; UDP-GLC, UDP Glucose; UDPG, UDP Glucuronate; UDPGs, UDP Glucose and UDP Glucuronate; UMP, Uridine 5′-monophosphate; Val, Valine; WST-1, Water-Soluble Tetrazolium salts; dDNP, dissolution Dynamic Nuclear Polarization; qRT-PCR, Real-Time Quantitative Reverse Transcription Polymerase Chain Reaction; α-KIV, α-Keto-isovalerate; α-KMV, α-keto-β-methyl-valerate
  34. Cancer Discov. 2023 Mar 09. pii: CD-22-0939. [Epub ahead of print]
      The BCL-2 inhibitor Venetoclax (VEN) in combination with Azacitidine (5-AZA) is currently transforming Acute Myeloid Leukemia (AML) therapy. However, there is a lack of clinically relevant biomarkers that predict response to 5-AZA/VEN. Here, we integrated transcriptomic, proteomic, functional and clinical data to identify predictors of 5-AZA/VEN response. Although cultured monocytic AML cells displayed upfront resistance, monocytic differentiation was not clinically predictive in our patient cohort. We identified leukemic stem cells (LSC) as primary targets of 5-AZA/VEN whose elimination determined therapy outcome. LSCs of 5-AZA/VEN refractory patients displayed perturbed apoptotic dependencies. We developed and validated a flow cytometry-based "Mediators-of-Apoptosis-Combinatorial-Score" (MAC-Score) linking the ratio of protein expression of BCL-2, BCL-xL, and MCL-1 in LSCs. MAC-Scoring predicts initial response with a positive predictive-value of >97% associated to increased event-free survival. In summary, combinatorial levels of BCL-2-family members in AML-LSCs are a key denominator of response and MAC-Scoring reliably predicts patient response to 5-AZA/VEN.
  35. Stem Cell Res Ther. 2023 Mar 08. 14(1): 36
      Bone marrow samples from discarded femoral heads are often used as healthy controls in studies investigating the in vitro characteristics of cells from patients with hematologic malignancies. Since patient samples are usually derived from iliac crest aspirates, this carries the risk that the properties of the cells from both sources might be different due to the site and method of harvesting. Comparing BM cells from iliac crest aspirates and femoral heads from age-matched healthy donors, we show that, while mesenchymal stromal cells have indistinguishable properties between both sources, hematopoietic stem and progenitor cells (HSPC) from femoral heads show a considerable proliferative advantage in vitro. These data therefore suggest that experiments comparing leukemic cells from the iliac crest to healthy HSPC obtained from femoral heads should be interpreted with caution.
    Keywords:  Femoral head; Healthy control; Hematopoietic stem cells; Iliac crest; Mesenchymal stromal cells
  36. Sci Rep. 2023 Mar 07. 13(1): 3839
      The mitochondrial ATP synthase, an enzyme that synthesizes ATP and is involved in the formation of the mitochondrial mega-channel and permeability transition, is a multi-subunit complex. In S. cerevisiae, the uncharacterized protein Mco10 was previously found to be associated with ATP synthase and referred as a new 'subunit l'. However, recent cryo-EM structures could not ascertain Mco10 with the enzyme making questionable its role as a structural subunit. The N-terminal part of Mco10 is very similar to k/Atp19 subunit, which along with subunits g/Atp20 and e/Atp21 plays a major role in stabilization of the ATP synthase dimers. In our effort to confidently define the small protein interactome of ATP synthase we found Mco10. We herein investigate the impact of Mco10 on ATP synthase functioning. Biochemical analysis reveal in spite of similarity in sequence and evolutionary lineage, that Mco10 and Atp19 differ significantly in function. The Mco10 is an auxiliary ATP synthase subunit that only functions in permeability transition.
  37. Nat Commun. 2023 Mar 08. 14(1): 1285
      Acute myeloid leukemia may be characterized by a fraction of leukemia stem cells (LSCs) that sustain disease propagation eventually leading to relapse. Yet, the contribution of LSCs to early therapy resistance and AML regeneration remains controversial. We prospectively identify LSCs in AML patients and xenografts by single-cell RNA sequencing coupled with functional validation by a microRNA-126 reporter enriching for LSCs. Through nucleophosmin 1 (NPM1) mutation calling or chromosomal monosomy detection in single-cell transcriptomes, we discriminate LSCs from regenerating hematopoiesis, and assess their longitudinal response to chemotherapy. Chemotherapy induced a generalized inflammatory and senescence-associated response. Moreover, we observe heterogeneity within progenitor AML cells, some of which proliferate and differentiate with expression of oxidative-phosphorylation (OxPhos) signatures, while others are OxPhos (low) miR-126 (high) and display enforced stemness and quiescence features. miR-126 (high) LSCs are enriched at diagnosis in chemotherapy-refractory AML and at relapse, and their transcriptional signature robustly stratifies patients for survival in large AML cohorts.
  38. Ann Maxillofac Surg. 2022 Jul-Dec;12(2):12(2): 144-150
      Introduction: Cytochrome C1 (CYC1) is an important subunit of mitochondrial complex III and plays a vital role in oxidative phosphorylation (OXPHOS) and reactive oxygen species generation. Overexpression of the CYC1 gene has been implicated in cancer development and its prognosis previously, but unexplored in head-and-neck squamous cell carcinomas (HNSCC), especially oral squamous cell carcinoma (OSCC).Materials and Methods: CYC1 m-RNA expression and gene alterations were assessed using the Cancer Genome Atlas dataset in HNSCC and validated in OSCC tissues using real-time polymerase chain reaction (RT-PCR). The protein-protein interaction (PPI) network and functional enrichment pathways were also analysed.
    Results: A thorough analysis of the TCGA (The Cancer Genome Atlas) database revealed that CYC1 was overexpressed in the HNSCC cases and the increased expression correlated with several parameters which involve the prediction of advanced diseases such as histopathological grade, tumour-node-metastasis staging, and nodal metastases (P < 0.05). The expression of CYC1 was validated using RT-PCR showing significant upregulation (P < 0.05) in OSCC tissue samples compared to the normal tissue counterparts. PPI network and functional analysis show the prominent role of CYC1 in OXPHOS, especially in electron transport chain III complex regulation.
    Discussion: The study revealed that CYC1 is highly expressed in HNSCC, and is validated in the OSCC patient tissue samples compared to the normal counterparts and associated with advanced disease stages and grade of the tumour. CYC1 could be a novel promising therapeutic and prognostic marker in HNSCC, especially in OSCC.
    Keywords:  Cytochrome C1; mRNA expression; oral squamous cell carcinoma; prognostic value; the cancer genome atlas database
  39. FEBS Open Bio. 2023 Mar 06.
      Mitofusin-2 (MFN2) is a transmembrane GTPase that regulates mitochondrial fusion and thereby modulates mitochondrial function. However, the role of MFN2 in lung adenocarcinoma remains controversial. Here, we investigated the effect of MFN2 regulation on mitochondria in lung adenocarcinoma. We found that MFN2 deficiency resulted in decreased UCP4 expression and mitochondrial dysfunction in A549 and H1975 cells. UCP4 overexpression restored ATP and intracellular calcium concentration, but not mtDNA copy number, mitochondrial membrane potential or reactive oxygen species level. Furthermore, mass spectrometry analysis identified 460 overlapping proteins after independent overexpression of MFN2 and UCP4; these proteins were significantly enriched in the cytoskeleton, energy production and calponin homology (CH) domains. Moreover, the calcium signaling pathway was confirmed to be enriched in KEGG pathway analysis. We also found by protein-protein interaction network analysis that PINK1 may be a key regulator of MFN2- and UCP4-mediated calcium homeostasis. Furthermore, PINK1 increased MFN2/UCP4-mediated intracellular Ca2+ concentration in A549 and H1975 cells. Finally, we demonstrated that low expression levels of MFN2 and UCP4 in lung adenocarcinoma are associated with poor clinical prognosis. In conclusion, our data suggest not only a potential role of MFN2 and UCP4 in co-regulating calcium homeostasis in lung adenocarcinoma, but also their potential use as therapeutic targets in lung cancer.
    Keywords:  Calcium homeostasis; Lung adenocarcinoma; MFN2; UCP4
  40. Oncogene. 2023 Mar 06.
      Oncometabolites, such as D/L-2-hydroxyglutarate (2HG), have directly been implicated in carcinogenesis; however, the underlying molecular mechanisms remain poorly understood. Here, we showed that the levels of the L-enantiomer of 2HG (L2HG) were specifically increased in colorectal cancer (CRC) tissues and cell lines compared with the D-enantiomer of 2HG (D2HG). In addition, L2HG increased the expression of ATF4 and its target genes by activating the mTOR pathway, which subsequently provided amino acids and improved the survival of CRC cells under serum deprivation. Downregulating the expression of L-2-hydroxyglutarate dehydrogenase (L2HGDH) and oxoglutarate dehydrogenase (OGDH) increased L2HG levels in CRC, thereby activating mTOR-ATF4 signaling. Furthermore, L2HGDH overexpression reduced L2HG-mediated mTOR-ATF4 signaling under hypoxia, whereas L2HGDH knockdown promoted tumor growth and amino acid metabolism in vivo. Together, these results indicate that L2HG ameliorates nutritional stress by activating the mTOR-ATF4 axis and thus could be a potential therapeutic target for CRC.