bims-mibica Biomed News
on Mitochondrial bioenergetics in cancer
Issue of 2021‒02‒21
forty-nine papers selected by
Kelsey Fisher-Wellman
East Carolina University

  1. Mol Oncol. 2021 Feb 18.
      STAT3 is a transcription factor with roles in inflammation and tumorigenicity. A fraction of STAT3 localizes in mitochondria, where it augments tumorigenesis via regulation of mitochondrial functions, including modulation of respiration and redox status. We show a novel mechanism for mitochondrial STAT3 regulation of redox homeostasis in triple negative breast cancer cells. Loss of STAT3 diminished complex I dehydrogenase activity and impaired NAD+ regeneration, leading to impaired expression of glutathione biosynthetic genes and other antioxidant genes. Expressing mitochondrially-restricted STAT3 or replenishment of the cellular NAD pool restored antioxidant gene expression, as did complementation of the NADH dehydrogenase activity by expression of the STAT3-independent yeast dehydrogenase, NDI1. These NAD-regulated processes contributed to malignant phenotypes by promoting clonal cell growth and migration. Proximity interaction and protein pull-down assays identified three components of complex I that associated with mitochondrial STAT3, providing a potential mechanistic basis for how mitochondrial STAT3 affects complex I activity. Our data document a novel mechanism through which mitochondrial STAT3 indirectly controls antioxidant gene regulation through a retrograde NAD+ signal that is modulated by complex I dehydrogenase activity.
    Keywords:  Mitochondria; STAT3; breast cancer; glutathione; oxidative stress; reactive oxygen species
  2. Commun Biol. 2021 Feb 16. 4(1): 217
      During the cancerous transformation of normal hepatocytes into hepatocellular carcinoma (HCC), the enzyme catalyzing the first rate-limiting step of glycolysis, namely the glucokinase (GCK), is replaced by the higher affinity isoenzyme, hexokinase 2 (HK2). Here, we show that in HCC tumors the highest expression level of HK2 is inversely correlated to GCK expression, and is associated to poor prognosis for patient survival. To further explore functional consequences of the GCK-to-HK2 isoenzyme switch occurring during carcinogenesis, HK2 was knocked-out in the HCC cell line Huh7 and replaced by GCK, to generate the Huh7-GCK+/HK2- cell line. HK2 knockdown and GCK expression rewired central carbon metabolism, stimulated mitochondrial respiration and restored essential metabolic functions of normal hepatocytes such as lipogenesis, VLDL secretion, glycogen storage. It also reactivated innate immune responses and sensitivity to natural killer cells, showing that consequences of the HK switch extend beyond metabolic reprogramming.
  3. Cell Metab. 2021 Feb 10. pii: S1550-4131(21)00056-5. [Epub ahead of print]
      Obesity is a major risk factor for adverse outcomes in breast cancer; however, the underlying molecular mechanisms have not been elucidated. To investigate the role of crosstalk between mammary adipocytes and neoplastic cells in the tumor microenvironment (TME), we performed transcriptomic analysis of cancer cells and adjacent adipose tissue in a murine model of obesity-accelerated breast cancer and identified glycine amidinotransferase (Gatm) in adipocytes and Acsbg1 in cancer cells as required for obesity-driven tumor progression. Gatm is the rate-limiting enzyme in creatine biosynthesis, and deletion in adipocytes attenuated obesity-driven tumor growth. Similarly, genetic inhibition of creatine import into cancer cells reduced tumor growth in obesity. In parallel, breast cancer cells in obese animals upregulated the fatty acyl-CoA synthetase Acsbg1 to promote creatine-dependent tumor progression. These findings reveal key nodes in the crosstalk between adipocytes and cancer cells in the TME necessary for obesity-driven breast cancer progression.
    Keywords:  Acsbg1; Gatm; breast cancer; creatine; hypoxia; obesity
  4. Circulation. 2021 Feb 17.
      Background: Heart failure is a leading cause of death worldwide and is associated with the rising prevalence of obesity, hypertension and diabetes. O-GlcNAcylation is a post-translational modification of intracellular proteins and serves as a metabolic rheostat for cellular stress. The total levels of O-GlcNAcylation are determined by nutrient and metabolic flux, in addition to the net activity of two enzymes, O-GlcNAc transferase (OGT) and O-GlcNAcase (OGA). Failing myocardium is marked by increased O-GlcNAcylation, but it is unknown if excessive O-GlcNAcylation contributes to cardiomyopathy and heart failure. Methods: We developed two new transgenic mouse models with myocardial overexpression of OGT and OGA to control O-GlcNAcylation independent of pathological stress. Results: We found that OGT transgenic hearts showed increased O-GlcNAcylation, and developed severe dilated cardiomyopathy, ventricular arrhythmias and premature death. In contrast, OGA transgenic hearts had lower O-GlcNAcylation but identical cardiac function to wild type littermate controls. Additionally, OGA transgenic hearts were resistant to pathological stress induced by pressure overload with attenuated myocardial O-GlcNAcylation levels after stress and decreased pathological hypertrophy compared to wild type controls. Interbreeding OGT with OGA transgenic mice rescued cardiomyopathy and premature death, despite persistent elevation of myocardial OGT. Transcriptomic and functional studies revealed disrupted mitochondrial energetics with impairment of complex I activity in hearts from OGT transgenic mice. Complex I activity was rescued by OGA transgenic interbreeding, suggesting an important role for mitochondrial complex I in O-GlcNAc mediated cardiac pathology. Conclusions: Our data provide evidence that excessive O-GlcNAcylation causes cardiomyopathy, at least in part, due to defective energetics. Enhanced OGA activity is well tolerated and attenuation of O-GlcNAcylation is beneficial against pressure overload induced pathologic remodeling and heart failure. These findings suggest attenuation of excessive O-GlcNAcylation may represent a novel therapeutic approach for cardiomyopathy.
    Keywords:  Dilated cardiomyopathy; O-GlcNAcylation; mitochondrial energetics; mouse model
  5. Sci Adv. 2021 Feb;pii: eabf0717. [Epub ahead of print]7(8):
      Induction of the one-carbon cycle is an early hallmark of mitochondrial dysfunction and cancer metabolism. Vital intermediary steps are localized to mitochondria, but it remains unclear how one-carbon availability connects to mitochondrial function. Here, we show that the one-carbon metabolite and methyl group donor S-adenosylmethionine (SAM) is pivotal for energy metabolism. A gradual decline in mitochondrial SAM (mitoSAM) causes hierarchical defects in fly and mouse, comprising loss of mitoSAM-dependent metabolites and impaired assembly of the oxidative phosphorylation system. Complex I stability and iron-sulfur cluster biosynthesis are directly controlled by mitoSAM levels, while other protein targets are predominantly methylated outside of the organelle before import. The mitoSAM pool follows its cytosolic production, establishing mitochondria as responsive receivers of one-carbon units. Thus, we demonstrate that cellular methylation potential is required for energy metabolism, with direct relevance for pathophysiology, aging, and cancer.
  6. Cell Metab. 2021 Feb 09. pii: S1550-4131(21)00013-9. [Epub ahead of print]
      The architecture of cristae provides a spatial mitochondrial organization that contains functional respiratory complexes. Several protein components including OPA1 and MICOS complex subunits organize cristae structure, but upstream regulatory mechanisms are largely unknown. Here, in vivo and in vitro reconstitution experiments show that the endoplasmic reticulum (ER) kinase PERK promotes cristae formation by increasing TOM70-assisted mitochondrial import of MIC19, a critical subunit of the MICOS complex. Cold stress or β-adrenergic stimulation activates PERK that phosphorylates O-linked N-acetylglucosamine transferase (OGT). Phosphorylated OGT glycosylates TOM70 on Ser94, enhancing MIC19 protein import into mitochondria and promoting cristae formation and respiration. In addition, PERK-activated OGT O-GlcNAcylates and attenuates CK2α activity, which mediates TOM70 Ser94 phosphorylation and decreases MIC19 mitochondrial protein import. We have identified a cold-stress inter-organelle PERK-OGT-TOM70 axis that increases cell respiration through mitochondrial protein import and subsequent cristae formation. These studies have significant implications in cellular bioenergetics and adaptations to stress conditions.
    Keywords:  MIC19; PERK-OGT axis; TOM70; brown adipocytes; cold stress; cristae biogenesis; mitochondrial protein import; respiration
  7. Proc Natl Acad Sci U S A. 2021 Feb 23. pii: e2012469118. [Epub ahead of print]118(8):
      Emerging evidence suggests that intratumoral interferon (IFN) signaling can trigger targetable vulnerabilities. A hallmark of pancreatic ductal adenocarcinoma (PDAC) is its extensively reprogrammed metabolic network, in which nicotinamide adenine dinucleotide (NAD) and its reduced form, NADH, are critical cofactors. Here, we show that IFN signaling, present in a subset of PDAC tumors, substantially lowers NAD(H) levels through up-regulating the expression of NAD-consuming enzymes PARP9, PARP10, and PARP14. Their individual contributions to this mechanism in PDAC have not been previously delineated. Nicotinamide phosphoribosyltransferase (NAMPT) is the rate-limiting enzyme in the NAD salvage pathway, a dominant source of NAD in cancer cells. We found that IFN-induced NAD consumption increased dependence upon NAMPT for its role in recycling NAM to salvage NAD pools, thus sensitizing PDAC cells to pharmacologic NAMPT inhibition. Their combination decreased PDAC cell proliferation and invasion in vitro and suppressed orthotopic tumor growth and liver metastases in vivo.
    Keywords:  NAD; NAMPT; PARP; interferon; pancreatic cancer
  8. Nat Commun. 2021 Feb 19. 12(1): 1190
      DddA-derived cytosine base editors (DdCBEs), composed of the split interbacterial toxin DddAtox, transcription activator-like effector (TALE), and uracil glycosylase inhibitor (UGI), enable targeted C-to-T base conversions in mitochondrial DNA (mtDNA). Here, we demonstrate highly efficient mtDNA editing in mouse embryos using custom-designed DdCBEs. We target the mitochondrial gene, MT-ND5 (ND5), which encodes a subunit of NADH dehydrogenase that catalyzes NADH dehydration and electron transfer to ubiquinone, to obtain several mtDNA mutations, including m.G12918A associated with human mitochondrial diseases and m.C12336T that incorporates a premature stop codon, creating mitochondrial disease models in mice and demonstrating a potential for the treatment of mitochondrial disorders.
  9. Cell Rep. 2021 Jan 26. pii: S2211-1247(20)31665-X. [Epub ahead of print]34(4): 108676
      Hypoxia, low oxygen (O2), is a key feature of all solid cancers, including hepatocellular carcinoma (HCC). Genome-wide CRISPR-Cas9 knockout library screening is used to identify reliable therapeutic targets responsible for hypoxic survival in HCC. We find that protein-tyrosine phosphatase mitochondrial 1 (PTPMT1), an important enzyme for cardiolipin (CL) synthesis, is the most significant gene and ranks just after hypoxia-inducible factor (HIF)-1α and HIF-1β as crucial to hypoxic survival. CL constitutes the mitochondrial membrane and ensures the proper assembly of electron transport chain (ETC) complexes for efficient electron transfer in respiration. ETC becomes highly unstable during hypoxia. Knockout of PTPMT1 stops the maturation of CL and impairs the assembly of ETC complexes, leading to further electron leakage and ROS accumulation at ETC in hypoxia. Excitingly, HCC cells, especially under hypoxic conditions, show great sensitivity toward PTPMT1 inhibitor alexidine dihydrochloride (AD). This study unravels the protective roles of PTPMT1 in hypoxic survival and cancer development.
    Keywords:  CRISPR library screening; ROS; cardiolipin; electron transport chain; hypoxia; liver cancer; metabolism; mitochondria; oxidative stress
  10. Front Cardiovasc Med. 2020 ;7 604581
      Mitochondrial medicine is an exciting and rapidly evolving field. While the mitochondrial genome is small and differs from the nuclear genome in that it is circular and free of histones, it has been implicated in neurodegenerative diseases, type 2 diabetes, aging and cardiovascular disorders. Currently, there is a lack of efficient treatments for mitochondrial diseases. This has promoted the need for developing an appropriate platform to investigate and target the mitochondrial genome. However, developing these therapeutics requires a model system that enables rapid and effective studying of potential candidate therapeutics. In the past decade, induced pluripotent stem cells (iPSCs) have become a promising technology for applications in basic science and clinical trials, and have the potential to be transformative for mitochondrial drug development. Engineered iPSC-derived cardiomyocytes (iPSC-CM) offer a unique tool to model mitochondrial disorders. Additionally, these cellular models enable the discovery and testing of novel therapeutics and their impact on pathogenic mtDNA variants and dysfunctional mitochondria. Herein, we review recent advances in iPSC-CM models focused on mitochondrial dysfunction often causing cardiovascular diseases. The importance of mitochondrial disease systems biology coupled with genetically encoded NAD+/NADH sensors is addressed toward developing an in vitro translational approach to establish effective therapies.
    Keywords:  cardiomyocytes; drug discovery; human induced pluripotent stem cells; mitochondrial disease; regenerative medicine; sonar sensor
  11. EMBO Rep. 2021 Feb 15. e51635
      Mitochondria possess a small genome that codes for core subunits of the oxidative phosphorylation system and whose expression is essential for energy production. Information on the regulation and spatial organization of mitochondrial gene expression in the cellular context has been difficult to obtain. Here we devise an imaging approach to analyze mitochondrial translation within the context of single cells, by following the incorporation of clickable non-canonical amino acids. We apply this method to multiple cell types, including specialized cells such as cardiomyocytes and neurons, and monitor with spatial resolution mitochondrial translation in axons and dendrites. We also show that translation imaging allows to monitor mitochondrial protein expression in patient fibroblasts. Approaching mitochondrial translation with click chemistry opens new avenues to understand how mitochondrial biogenesis is integrated into the cellular context and can be used to assess mitochondrial gene expression in mitochondrial diseases.
    Keywords:  gene expression; hippocampal neuron; mitochondria; synapse; translation
  12. Front Cell Dev Biol. 2020 ;8 596819
      The fate and proliferative capacity of stem cells have been shown to strongly depend on their metabolic state. Mitochondria are the powerhouses of the cell being responsible for energy production via oxidative phosphorylation (OxPhos) as well as for several other metabolic pathways. Mitochondrial activity strongly depends on their structural organization, with their size and shape being regulated by mitochondrial fusion and fission, a process known as mitochondrial dynamics. However, the significance of mitochondrial dynamics in the regulation of stem cell metabolism and fate remains elusive. Here, we characterize the role of mitochondria morphology in female germ stem cells (GSCs) and in their more differentiated lineage. Mitochondria are particularly important in the female GSC lineage. Not only do they provide these cells with their energy requirements to generate the oocyte but they are also the only mitochondria pool to be inherited by the offspring. We show that the undifferentiated GSCs predominantly have fissed mitochondria, whereas more differentiated germ cells have more fused mitochondria. By reducing the levels of mitochondrial dynamics regulators, we show that both fused and fissed mitochondria are required for the maintenance of a stable GSC pool. Surprisingly, we found that disrupting mitochondrial dynamics in the germline also strongly affects nurse cells morphology, impairing egg chamber development and female fertility. Interestingly, reducing the levels of key enzymes in the Tricarboxylic Acid Cycle (TCA), known to cause OxPhos reduction, also affects GSC number. This defect in GSC self-renewal capacity indicates that at least basal levels of TCA/OxPhos are required in GSCs. Our findings show that mitochondrial dynamics is essential for female GSC maintenance and female fertility, and that mitochondria fusion and fission events are dynamically regulated during GSC differentiation, possibly to modulate their metabolic profile.
    Keywords:  Drosophila melanogaster; differentiation; fertility; germ stem cell; mitochondrial dynamics; oogenesis; oxidative phosphorylation
  13. Pharmacol Res. 2021 Feb 15. pii: S1043-6618(21)00079-7. [Epub ahead of print] 105495
      In mammalian cells enzymatic and non-enzymatic pathways produce H2S, a gaseous transmitter which recently emerged as promising therapeutic agent and modulator of mitochondrial bioenergetics. To explore this topic, the H2S donor NaHS, at micromolar concentrations, was tested on swine heart mitochondria. NaHS did not affect the F1FO-ATPase activated by the natural cofactor Mg2, but, when Mg2+ was replaced by Ca2+, a slight 15% enzyme inhibition at 100µM NaHS was shown. Conversely, both the NADH-O2 and succinate-O2 oxidoreductase activities were totally inhibited by 200μM NaHS with IC50 values of 61.6±4.1 and 16.5±4.6μM NaHS, respectively. Since the mitochondrial respiration was equally inhibited by NaHS at both first or second respiratory substrates sites, the H2S generation may prevent the electron transfer from complexes I and II to downhill respiratory chain complexes, probably because H2S competes with O2 in complex IV, thus reducing membrane potential as a consequence of the cytochrome c oxidase activity inhibition. The Complex IV blockage by H2S was consistent with the linear concentration-dependent NADH-O2 oxidoreductase inhibition and exponential succinate-O2 oxidoreductase inhibition by NaHS, whereas the coupling between substrate oxidation and phosphorylation was unaffected by NaHS. Even if H2S is known to cause sulfhydration of cysteine residues, thiol oxidizing (GSSG) or reducing (DTE) agents, did not affect the F1FO-ATPase activities and mitochondrial respiration, thus ruling out any involvement of post-translational modifications of thiols. The permeability transition pore, the lethal channel which forms when the F1FO-ATPase is stimulated by Ca2+, did not open in the presence of NaHS, which shows a similar effect to ruthenium red, thus suggesting a putative Ca2+ transport cycle inhibition.
    Keywords:  F(1)F(O)-ATPase; H(2)S; cofactors; mitochondria; mitochondrial respiration; permeability transition pore
  14. Am J Transl Res. 2021 ;13(2): 601-616
      Oxaliplatin (OXA), as a third-generation platinum anticancer drug, is a treatment drug for gastric cancer (GC). However, OXA resistance has become the main reason for OXA treatment failure. Serine beta-lactamase-like protein (LACTB), acts as a mitochondrial protein, can affect multiple cancer processes. Here, we aimed to investigate the function and mechanism of LACTB in OXA-resistant GC. After LACTB overexpression or autophagy activator (RAPA) treatment, cell proliferation, reactive oxygen species (ROS), apoptosis, mitochondrial dysfunction were evaluated through CCK-8 assay, Edu staining, flow cytometry and immunofluorescence assay. Moreover, DNA double-stranded damage and autophagy-related proteins were examined via western blot. We revealed that LACTB was downregulated in OXA-resistant MGC-803 cells, and overexpression of LACTB reduced the resistance of GC cells to OXA. Besides, our results uncovered that overexpression of LACTB induced apoptosis, reduced the mitochondrial membrane potential (MMP) and accelerated ROS accumulation in OXA-resistant MGC-803 (MGC-803/OXA) cells. Meanwhile, we verified that overexpression of LACTB decreased glucose uptake and ATP synthesis, induced mitochondria and DNA damages, and inhibited autophagy of MGC-803/OXA cells. Furthermore, our results certified that RAPA could weaken the function of LACTB on apoptosis and mitochondrial morphology and function in OXA-resistant MGC-803 cells with OXA treatment. Therefore, we demonstrated that LACTB could attenuate the resistance of MGC-803/OXA cells to OXA through autophagy-mediated mitochondrial morphological changes, mitochondrial dysfunction, and apoptosis, suggesting that LACTB, functions as a suppressor, is conducive to the therapy of OXA-resistant GC.
    Keywords:  Gastric cancer; apoptosis; autophagy; mitochondrial dysfunction; oxaliplatin; serine beta-lactamase-like protein
  15. Cancer Med. 2021 Feb 18.
      Autophagy removes damaged organelles to inhibit malignant transformation during tumor initiation. Once a cancer matures, it uses the autophagic pathway as an energy source. Optineurin (OPTN) is an autophagy adaptor protein that recruits microtubule-associated protein 1 light chain 3, an autophagosome marker, to the autophagosome. Despite studies of the relation between cancer progression and autophagy adaptor proteins, there are no reports to our knowledge of a correlation between hepatocellular carcinoma (HCC) and OPTN. We aimed here to investigate the effects of OPTN expression on HCC progression through autophagy. Immunohistochemistry was used to measure the OPTN expression in the tissues of 141 Japanese patients with HCC. The effects of OPTN expression on HCC progression and mitophagy were assessed using an OPTN knockout (KO) cell line in vitro. We used this KO cell line to establish and exploit a mouse model of HCC to determine the effects of OPTN expression on tumor progression. Immunohistochemical analysis showed that patients with elevated expression of OPTN experienced shorter overall survival (OS) and recurrence-free survival (RFS). OPTN KO cells proliferated relatively slower versus wild-type (WT) cells in vitro. Western blot analysis showed that mitophagy was suppressed in OPTN KO cells, and ATP synthesis and beta-oxidation were reduced. The mouse model of HCC showed that OPTN KO cells formed smaller tumors versus WT cells less 10 weeks after implantation. Overall, the present findings suggest that OPTN is a key mediator of mitophagy that contributes to HCC progression through mitochondrial energy production.
    Keywords:  adaptor protein; autophagy; beta-oxidation; liver; mitochondria
  16. Methods. 2021 Feb 12. pii: S1046-2023(21)00043-8. [Epub ahead of print]
      Mitochondria participate in series of metabolic processes and cellular events. It is widely known that only 13 proteins are encoded by the mammalian mitochondria genome. However, it is not acknowledged until recently that mitochondrial genomes encode hundreds of circular RNAs, named as mecciRNAs. Some of these mecciRNAs can serve as molecular chaperones to help folding nuclear-encoded proteins and facilitating their mitochondrial entrance. As a novel type of circular RNAs, functions and characteristics of mecciRNAs are waiting for further exploration and methods for mecciRNA studies need to be improved. Here, we describe detailed methods for mitochondrial RNA isolation and mecciRNA detection. In addition, we present effective mecciRNA overexpression and knockdown strategies for future functional studies of mecciRNAs.
  17. Sci Rep. 2021 Feb 18. 11(1): 4181
      The primary cilium is well-preserved in human differentiated thyroid cancers such as papillary and follicular carcinoma. Specific thyroid cancers such as Hürthle cell carcinoma, oncocytic variant of papillary thyroid carcinoma (PTC), and PTC with Hashimoto's thyroiditis show reduced biogenesis of primary cilia; these cancers are often associated the abnormalities in mitochondrial function. Here, we examined the association between primary cilia and the mitochondria-dependent apoptosis pathway. Tg-Cre;Ift88flox/flox mice (in which thyroid follicles lacked primary cilia) showed irregularly dilated follicles and increased apoptosis of thyrocytes. Defective ciliogenesis caused by deleting the IFT88 and KIF3A genes from thyroid cancer cell lines increased VDAC1 oligomerization following VDAC1 overexpression, thereby facilitating upregulation of mitochondria-dependent apoptosis. Furthermore, VDAC1 localized with the basal bodies of primary cilia in thyroid cancer cells. These results demonstrate that loss-of-function of primary cilia results in apoptogenic stimuli, which are responsible for mitochondrial-dependent apoptotic cell death in differentiated thyroid cancers. Therefore, regulating primary ciliogenesis might be a therapeutic approach to targeting differentiated thyroid cancers.
  18. Nat Commun. 2021 02 15. 12(1): 1041
      Growing evidence supports that pharmacological application of growth differentiation factor 15 (GDF15) suppresses appetite but also promotes sickness-like behaviors in rodents via GDNF family receptor α-like (GFRAL)-dependent mechanisms. Conversely, the endogenous regulation of GDF15 and its physiological effects on energy homeostasis and behavior remain elusive. Here we show, in four independent human studies that prolonged endurance exercise increases circulating GDF15 to levels otherwise only observed in pathophysiological conditions. This exercise-induced increase can be recapitulated in mice and is accompanied by increased Gdf15 expression in the liver, skeletal muscle, and heart muscle. However, whereas pharmacological GDF15 inhibits appetite and suppresses voluntary running activity via GFRAL, the physiological induction of GDF15 by exercise does not. In summary, exercise-induced circulating GDF15 correlates with the duration of endurance exercise. Yet, higher GDF15 levels after exercise are not sufficient to evoke canonical pharmacological GDF15 effects on appetite or responsible for diminishing exercise motivation.
  19. Nature. 2021 02;590(7846): 480-485
      Obesity increases the risk of mortality because of metabolic sequelae such as type 2 diabetes and cardiovascular disease1. Thermogenesis by adipocytes can counteract obesity and metabolic diseases2,3. In thermogenic fat, creatine liberates a molar excess of mitochondrial ADP-purportedly via a phosphorylation cycle4-to drive thermogenic respiration. However, the proteins that control this futile creatine cycle are unknown. Here we show that creatine kinase B (CKB) is indispensable for thermogenesis resulting from the futile creatine cycle, during which it traffics to mitochondria using an internal mitochondrial targeting sequence. CKB is powerfully induced by thermogenic stimuli in both mouse and human adipocytes. Adipocyte-selective inactivation of Ckb in mice diminishes thermogenic capacity, increases predisposition to obesity, and disrupts glucose homeostasis. CKB is therefore a key effector of the futile creatine cycle.
  20. Mol Genet Metab. 2021 Feb 11. pii: S1096-7192(21)00034-2. [Epub ahead of print]
      Osteopenia occurs in a subset of phenylalanine hydroxylase (PAH) deficient phenylketonuria (PKU) patients. While osteopenia is not fully penetrant in patients, the Pahenu2 classical PKU mouse is universally osteopenic, making it an ideal model of the phenotype. Pahenu2 Phe management, with a Phe-fee amino acid defined diet, does not improve bone density as histomorphometry metrics remain indistinguishable from untreated animals. Previously, we demonstrated Pahenu2 mesenchymal stem cells (MSCs) display impaired osteoblast differentiation. Oxidative stress is recognized in PKU patients and PKU animal models. Pahenu2 MSCs experience oxidative stress determined by intracellular superoxide over-representation. The deleterious impact of oxidative stress on mitochondria is recognized. Oximetry applied to Pahenu2 MSCs identified mitochondrial stress by increased basal respiration with concurrently reduced maximal respiration and respiratory reserve. Proton leak secondary to mitochondrial complex 1 dysfunction is a recognized superoxide source. Respirometry applied to Pahenu2 MSCs, in the course of osteoblast differentiation, identified a partial complex 1 deficit. Pahenu2 MSCs treated with the antioxidant resveratrol demonstrated increased mitochondrial mass by MitoTracker green labeling. In hyperphenylalaninemic conditions, resveratrol increased in situ alkaline phosphatase activity suggesting partial recovery of Pahenu2 MSCs osteoblast differentiation. Up-regulation of oxidative energy production is required for osteoblasts differentiation. Our data suggests impaired Pahenu2 MSC developmental competence involves an energy deficit. We posit energy support and oxidative stress reduction will enable Pahenu2 MSC differentiation in the osteoblast lineage to subsequently increase bone density.
    Keywords:  Osteopenia; Oxidative stress; Pah(enu2); Phenylketonuria; Respiratory complex 1
  21. J Clin Invest. 2021 Feb 15. pii: 141799. [Epub ahead of print]131(4):
      The relationship between adiposity and metabolic health is well established. However, very little is known about the fat depot, known as paracardial fat (pCF), located superior to and surrounding the heart. Here, we show that pCF remodels with aging and a high-fat diet and that the size and function of this depot are controlled by alcohol dehydrogenase 1 (ADH1), an enzyme that oxidizes retinol into retinaldehyde. Elderly individuals and individuals with obesity have low ADH1 expression in pCF, and in mice, genetic ablation of Adh1 is sufficient to drive pCF accumulation, dysfunction, and global impairments in metabolic flexibility. Metabolomics analysis revealed that pCF controlled the levels of circulating metabolites affecting fatty acid biosynthesis. Also, surgical removal of the pCF depot was sufficient to rescue the impairments in cardiometabolic flexibility and fitness observed in Adh1-deficient mice. Furthermore, treatment with retinaldehyde prevented pCF remodeling in these animals. Mechanistically, we found that the ADH1/retinaldehyde pathway works by driving PGC-1α nuclear translocation and promoting mitochondrial fusion and biogenesis in the pCF depot. Together, these data demonstrate that pCF is a critical regulator of cardiometabolic fitness and that retinaldehyde and its generating enzyme ADH1 act as critical regulators of adipocyte remodeling in the pCF depot.
    Keywords:  Adipose tissue; Cardiology; Cardiovascular disease; Metabolism
  22. Biochim Biophys Acta Bioenerg. 2021 Feb 15. pii: S0005-2728(21)00028-1. [Epub ahead of print] 148395
      Complexome Profiling (CP) combines size separation, by electrophoresis or other means, of native multimeric complexes with protein identification by mass spectrometry (MS). Peptide MS analysis of the multiple fractions in which the sample is separated, results in the creation of protein abundance profiles in function of molecular size, providing a visual output of the assembly status of a group of proteins of interest. Stable isotope labelling by amino acids in cell culture (SILAC) is an established quantitative proteomics technique that allows duplexing in the MS analysis as well as the comparison of relative protein abundances between the samples, which are processed and analysed together. Combining SILAC and CP permitted the direct comparison of migration and abundance of the proteins present in the mitochondrial respiratory chain complexes in two different samples. This analysis, however, introduced a level of complexity in data processing for which bioinformatic tools had to be developed in order to generate the normalised protein abundance profiles. The advantages and challenges of using of this type of analysis for the characterisation of two cell lines carrying pathological variants in MT-CO3 and MT-CYB is reviewed. An additional unpublished example of SILAC-CP of a cell line with an in-frame 18-bp deletion in MT-CYB is presented. In these cells, in contrast to other MT-CYB deficient models, a small proportion of complex III2 is formed and it is found associated with fully assembled complex I. This analysis also revealed a profuse accumulation of assembly intermediates containing complex III subunits UQCR10 and CYC1, as well as a profound early-stage complex IV assembly defect.
    Keywords:  Complex III; Complex IV; Mitochondrial Disease; Quantitative Proteomics; SILAC; mitochondrial DNA mutations
  23. J Comp Physiol B. 2021 Feb 16.
      Ruddy shelduck migrate from wintering grounds in lowland India and Myanmar to breeding grounds in central China and Mongolia, sustaining flight over the Himalayas, where oxygen availability is greatly reduced. We compared phenotypes of the pectoralis muscle and the ventricle of the heart from ruddy shelduck and common shelduck (a closely related low-altitude congener) that were raised in common conditions at sea level, predicting that oxidative capacity would be greater in ruddy shelduck to support high-altitude migration. Fibre-type composition of the pectoralis and the maximal activity of eight enzymes involved in mitochondrial energy metabolism in the pectoralis and heart, were compared between species. Few differences distinguished ruddy shelduck from common shelduck in the flight muscle, with the exception that ruddy shelduck had higher activities of complex II and higher ratios of complex IV (cytochrome c oxidase) and complex II when expressed relative to citrate synthase activity. There were no species differences in fibre-type composition, so these changes in enzyme activity may reflect an evolved modification in the functional properties of muscle mitochondria, potentially influencing mitochondrial respiratory capacity and/or oxygen affinity. Ruddy shelduck also had higher lactate dehydrogenase activity concurrent with lower pyruvate kinase and hexokinase activity in the left ventricle, which likely reflects an increased capacity for lactate oxidation by the heart. We conclude that changes in pathways of mitochondrial energy metabolism in the muscle and heart may contribute to the ability of ruddy shelduck to fly at high altitude.
    Keywords:  Enzyme; High altitude; Histology; Migrant; Muscle phenotype
  24. Nat Commun. 2021 02 16. 12(1): 1054
      In acute myeloid leukemia (AML), molecular heterogeneity across patients constitutes a major challenge for prognosis and therapy. AML with NPM1 mutation is a distinct genetic entity in the revised World Health Organization classification. However, differing patterns of co-mutation and response to therapy within this group necessitate further stratification. Here we report two distinct subtypes within NPM1 mutated AML patients, which we label as primitive and committed based on the respective presence or absence of a stem cell signature. Using gene expression (RNA-seq), epigenomic (ATAC-seq) and immunophenotyping (CyToF) analysis, we associate each subtype with specific molecular characteristics, disease differentiation state and patient survival. Using ex vivo drug sensitivity profiling, we show a differential drug response of the subtypes to specific kinase inhibitors, irrespective of the FLT3-ITD status. Differential drug responses of the primitive and committed subtype are validated in an independent AML cohort. Our results highlight heterogeneity among NPM1 mutated AML patient samples based on stemness and suggest that the addition of kinase inhibitors to the treatment of cases with the primitive signature, lacking FLT3-ITD, could have therapeutic benefit.
  25. Front Oncol. 2020 ;10 595524
      CISD2, a NEET protein that coordinates 2Fe-2S clusters through its CDGSH domain, is critical for normal development and iron homeostasis. CISD2 plays an important role in Fe-S cluster transfer and promotes cancer proliferation. However, its specific role in the development of non-small cell lung cancer (NSCLC) remains unclear. Bioinformatics of pan-cancer analysis from The Cancer Genome Atlas show that CISD2 has an aberrant expression in most types of human cancers. Moreover, CISD2 expression is associated with a higher hazard ratio and exhibits significantly poorer overall survival in lung adenocarcinoma (LUAD), uveal melanoma, head and neck squamous cell carcinoma, brain lower grade glioma, kidney chromophobe, and liver hepatocellular carcinoma. Further investigation revealed that CISD2 is highly expressed in LUAD and LUSC, which is associated with clinical pathological stages. In addition, survival data collected from GSE31210 and GSE13213, two datasets from the NCBI Gene Expression Omnibus, also confirmed that high CISD2 expression is associated with unfavorable survival in patients with LUAD. A cell-based assay indicated that the knockdown of CISD2 inhibited proliferation, invasion, and migration in A549 cells. Additionally, CISD2 knockdown accelerated the accumulation of cellular and mitochondrial reactive oxygen species, destroying the mitochondrial morphology and function. Moreover, CISD2 inhibition activated the iron starvation response, thus, accelerating iron accumulation in A549 cells. Pretreatment with DFO, the iron chelator, blocked mitochondrial dysfunction in CISD2-knockdown cells. Collectively, the present study provides novel insights into the regulatory role of CISD2 in NSCLC and presents a potential target to improve antitumor activity based on oxidative stress.
    Keywords:  CISD2; iron metabolism; iron-sulfur cluster; mitochondria; oxidative stress
  26. Methods Mol Biol. 2021 ;2224 47-60
      Like bacterial and cytoplasmic ribosomes, mitoribosomes are large ribonucleoprotein complexes with molecular weights in the range of several million Daltons. Traditionally, studying the assembly of such high molecular weight complexes is done using ultracentrifugation through linear density gradients, which remains the method of choice due to its versatility and superior resolving power in the high molecular weight range. Here, we present a protocol for the analysis of mitoribosomal assembly in heart mitochondrial extracts using linear density sucrose gradients that we have previously employed to characterize the essential role of different mitochondrial proteins in mitoribosomal biogenesis. This protocol details in a stepwise manner a typical mitoribosomal assembly analysis starting with isolation of mitochondria, preparation and ultracentrifugation of the gradients, fractionation and ending with SDS-PAGE, and immunoblotting of the gradient fractions. Even though we provide an example with heart mitochondria, this protocol can be directly applied to virtually all mouse tissues, as well as cultured cells, with little to no modifications.
    Keywords:  55S; Heart-specific knockout mice; Mitoribosome; Sedimentation analysis; Sucrose gradient; mt-LSU; mt-SSU
  27. Biochem Biophys Res Commun. 2021 Feb 11. pii: S0006-291X(21)00175-3. [Epub ahead of print]546 138-144
      Hepatic injury is common in patients who suffer from severe burns plus delayed resuscitation (B + DR). Stimulator of interferon genes (STING) is primarily expressed in Kupffer cells (KCs). We demonstrated that B + DR caused hepatic injury and oxidative stress. Reactive oxygen species (ROS) damage mitochondrial membranes in hepatocytes, leading to the release of mitochondrial DNA (mtDNA) into the hepatocyte cytosol and the circulation. The damaged hepatocytes then activate the mtDNA/STING pathway in KCs and trigger KCs polarization towards pro-inflammatory phenotype. SS-31 is a strong antioxidant that specifically concentrates in the inner mitochondrial membrane. SS-31 prevented hepatic injury by neutralizing ROS, inhibiting the release of mtDNA, protecting hepatocyte mitochondria, suppressing the activation of the mtDNA/STING pathway and inhibiting KCs polarization into pro-inflammatory phenotype.
    Keywords:  Hepatic injury; Kupffer cells; Mitochondria; Oxidative stress; STING; mtDNA
  28. Nano Lett. 2021 Feb 17.
      Immunogenic cell death (ICD), a manner of tumor cell death that can trigger antitumor immune responses, has received extensive attention as a potential synergistic modality for cancer immunotherapy. Although many calcium ion (Ca2+) nanomodulators have been developed for cancer therapy through mitochondrial Ca2+ overload, their ICD-inducing properties have not been explored. Herein, an acid-sensitive PEG-decorated calcium carbonate (CaCO3) nanoparticle incorporating curcumin (CUR; a Ca2+ enhancer) (PEGCaCUR) was prepared using a simple one-pot strategy. PEGCaCUR served as not only a Ca2+ nanomodulator inducing efficient mitochondrial Ca2+ overload but also an ICD inducer during improved synergistic cancer therapy. Combination of PEGCaCUR with ultrasound (US), PEGCaCUR+US, led to an enhanced ICD effect attributable to the enhanced mitochondrial Ca2+ overload, along with subsequent upregulation of reactive oxygen species levels. PEGCaCUR also facilitates photoacoustic/fluorescence dual-mode imaging, as well as effectively suppressing tumor growth and metastasis, indicating promising theranostic properties.
    Keywords:  Calcium nanomodulator; immunogenic cell death; immunotherapy; mitochondrial calcium ion overload; ultrasound
  29. Nat Commun. 2021 02 18. 12(1): 1135
      While >300 disease-causing variants have been identified in the mitochondrial DNA (mtDNA) polymerase γ, no mitochondrial phenotypes have been associated with POLRMT, the RNA polymerase responsible for transcription of the mitochondrial genome. Here, we characterise the clinical and molecular nature of POLRMT variants in eight individuals from seven unrelated families. Patients present with global developmental delay, hypotonia, short stature, and speech/intellectual disability in childhood; one subject displayed an indolent progressive external ophthalmoplegia phenotype. Massive parallel sequencing of all subjects identifies recessive and dominant variants in the POLRMT gene. Patient fibroblasts have a defect in mitochondrial mRNA synthesis, but no mtDNA deletions or copy number abnormalities. The in vitro characterisation of the recombinant POLRMT mutants reveals variable, but deleterious effects on mitochondrial transcription. Together, our in vivo and in vitro functional studies of POLRMT variants establish defective mitochondrial transcription as an important disease mechanism.
  30. iScience. 2021 Feb 19. 24(2): 102091
      Tumors comprise cancer cells and the associated stromal and immune/inflammatory cells, i.e., tumor microenvironment (TME). Here, we identify a metabolic signature of human and mouse model of gastric cancer and show that vagotomy in the mouse model reverses the metabolic reprogramming, reflected by metabolic switch from glutaminolysis to OXPHOS/glycolysis and normalization of the energy metabolism in cancer cells and TME. We next identify and validate SNAP25, mTOR, PDP1/α-KGDH, and glutaminolysis as drug targets and accordingly propose a therapeutic strategy to target the nerve-cancer metabolism. We demonstrate the efficacy of nerve-cancer metabolism therapy by intratumoral injection of BoNT-A (SNAP25 inhibitor) with systemic administration of RAD001 and CPI-613 but not cytotoxic drugs on overall survival in mice and show the feasibility in patients. These findings point to the importance of neural signaling in modulating the tumor metabolism and provide a rational basis for clinical translation of the potential strategy for gastric cancer.
    Keywords:  Cancer; Cancer Systems Biology
  31. Nat Commun. 2021 02 18. 12(1): 1134
      The BCL-2 family protein BAX has essential activity in mitochondrial regulation of cell death. While BAX activity ensures tissue homeostasis, when dysregulated it contributes to aberrant cell death in several diseases. During cellular stress BAX is transformed from an inactive cytosolic conformation to a toxic mitochondrial oligomer. Although the BAX transformation process is not well understood, drugs that interfere with this process are useful research tools and potential therapeutics. Here, we show that Eltrombopag,  an FDA-approved drug,  is a direct inhibitor of BAX. Eltrombopag binds the BAX trigger site distinctly from BAX activators, preventing them from triggering BAX conformational transformation and simultaneously promoting stabilization of the inactive BAX structure. Accordingly, Eltrombopag is capable of inhibiting BAX-mediated apoptosis induced by cytotoxic stimuli. Our data demonstrate structure-function insights into a mechanism of BAX inhibition and reveal a mechanism for Eltrombopag that may expand its use in diseases of uncontrolled cell death.
  32. Cell Rep. 2021 Feb 16. pii: S2211-1247(21)00063-2. [Epub ahead of print]34(7): 108750
      Inter-cellular heterogeneity in metabolic state has been proposed to influence many cancer phenotypes, including responses to targeted therapy. Here, we track the transitions and heritability of metabolic states in single PIK3CA mutant breast cancer cells, identify non-genetic glycolytic heterogeneity, and build on observations derived from methods reliant on bulk analyses. Using fluorescent biosensors in vitro and in tumors, we have identified distinct subpopulations of cells whose glycolytic and mitochondrial metabolism are regulated by combinations of phosphatidylinositol 3-kinase (PI3K) signaling, bromodomain activity, and cell crowding effects. The actin severing protein cofilin, as well as PI3K, regulates rapid changes in glucose metabolism, whereas treatment with the bromodomain inhibitor slowly abrogates a subpopulation of cells whose glycolytic activity is PI3K independent. We show how bromodomain function and PI3K signaling, along with actin remodeling, independently modulate glycolysis and how targeting these pathways affects distinct subpopulations of cancer cells.
    Keywords:  FRET imaging; PI3K signaling; breast cancer; cofilin; intra-tumor heterogeneity; intravital imaging; tumor metabolism
  33. Front Oncol. 2020 ;10 600980
      Background: The interplay between neoplastic cells and surrounding extracellular matrix (ECM) is one of the determinant elements for cancer growth. The remodeling of the ECM by cancer-associated fibroblasts (CAFs) shapes tumor microenvironment by depositing and digesting ECM proteins, hence promoting tumor growth and invasion. While for epithelial tumors CAFs are well characterized, little is known about the stroma composition of mesenchymal cancers, such as in rhabdomyosarcoma (RMS), the most common soft tissue sarcoma during childhood and adolescence. The aim of this work is to identify the importance of CAFs in specifying RMS microenvironment and the role of these stromal cells in RMS growth.Methods: We assessed in two dimensional (2D) and three dimensional (3D) systems the attraction between RMS cells and fibroblasts using epithelial colon cancer cell line as control. CAFs were studied in a xenogeneic mouse model of both tumor types and characterized in terms of fibroblast activation protein (FAP), mouse PDGFR expression, metalloproteases activation, and ECM gene and protein expression profiling.
    Results: In 2D model, the rate of interaction between stromal and malignant cells was significantly lower in RMS with respect to colon cancer. Particularly, in 3D system, RMS spheroids tended to dismantle the compact aggregate when grown on the layer of stromal cells. In vivo, despite the well-formed tumor mass, murine CAFs were found in low percentage in RMS xenogeneic samples.
    Conclusions: Our findings support the evidence that, differently from epithelial cancers, RMS cells are directly involved in their own ECM remodeling, and less dependent on CAFs support for cancer cell growth.
    Keywords:  cancer-associated fibroblasts; extracellular matrix proteins; rhabdomyosarcoma; stroma; tumor microenvironment
  34. Life Sci. 2021 Feb 16. pii: S0024-3205(21)00228-9. [Epub ahead of print] 119243
      High fat consumption leads to reactive oxygen species (ROS) which is associated with age-progressive neurological disorders. Cu/Zn superoxide dismutase (SOD1) is a critical enzyme against ROS. However, the relationship between SOD1 and the high-fat-induced ROS and neurodegeneration is poorly known. Here we showed that, upon treatment with a saturated fatty acid palmitic acid (PA), the SOD1 activity was decreased in mouse neuronal HT-22 cell line accompanied by elevation of ROS, but not in mouse microglial BV-2 cell line. We further showed that PA decreased the levels of copper chaperone for SOD1 (CCS) in HT-22 cells, which promoted the nuclear import of SOD1 and decreased its activity. We demonstrated that the reduction of CCS is involved in the PA-induced decrease of SOD1 activity and elevation of ROS. In addition, compared with the adult mice fed with a standard diet, the high-fat-diet adult mice presented an increase of plasma free fatty acids, reduction of hippocampal SOD1 activity and CCS, mitochondrial degeneration and long-term memory decline. Taken together, our findings suggest that the high-fat-induced lower CCS level is essential for SOD1 suppression which may be associated with neurodegeneration and cognitive decline.
    Keywords:  Copper chaperone for SOD1 (CCS); Cu/Zn superoxide dismutase (SOD1); High fat; Learning and memory; Mitochondrial degeneration; Reactive oxygen species (ROS)
  35. Hum Mol Genet. 2021 Feb 18. pii: ddab043. [Epub ahead of print]
      The human ATP synthase is an assembly of 29 subunits of 18 different types, of which only two (a and 8) are encoded in the mitochondrial genome. Subunit a, together with an oligomeric ring of c-subunit (c-ring), forms the proton pathway responsible for the transport of protons through the mitochondrial inner membrane, coupled to rotation of the c-ring and ATP synthesis. Neuromuscular diseases have been associated to a number of mutations in the gene encoding subunit a, ATP6. The most common, m.8993 T > G, leads to replacement of a strictly conserved leucine residue with arginine (aL156R). We previously showed that the equivalent mutation (aL173R) dramatically compromises respiratory growth of Saccharomyces cerevisiae and causes a 90% drop in the rate of mitochondrial ATP synthesis. Here we isolated revertants from the aL173R strain that show improved respiratory growth. Four first-site reversions at codon 173 (aL173M, aL173S, aL173K, and aL173W) and five second-site reversions at another codon (aR169M, aR169S, aA170P, aA170G, and aI216S) were identified. Based on the atomic structures of yeast ATP synthase and the biochemical properties of the revertant strains, we propose that the aL173R mutation is responsible for unfavorable electrostatic interactions that prevent the release of protons from the c-ring into a channel from which protons move from the c-ring to the mitochondrial matrix. The results provide further evidence that yeast aL173 (and thus human aL156) optimizes the exit of protons from ATP synthase, but is not essential despite its strict evolutionnary conservation.
  36. Mol Cell. 2021 Feb 18. pii: S1097-2765(21)00089-7. [Epub ahead of print]81(4): 642-644
      Luengo et al. (2020) demonstrate that pyruvate dehydrogenase (PDH) overactivation blunts NAD+ regeneration by overcharging the mitochondrial membrane potential and driving ATP synthesis beyond demand. Under these conditions, some cells prioritize aerobic glycolysis to meet the need for oxidized cofactors in biosynthetic metabolism.
  37. Cell Death Dis. 2021 Feb 16. 12(2): 189
      Oncogenic RAS is a critical driver for the initiation and progression of several types of cancers. However, effective therapeutic strategies by targeting RAS, in particular RASG12D and RASG12V, and associated downstream pathways have been so far unsuccessful. Treatment of oncogenic RAS-ravaged cancer patients remains a currently unmet clinical need. Consistent with a major role in cancer metabolism, oncogenic RAS activation elevates both reactive oxygen species (ROS)-generating NADPH oxidase (NOX) activity and ROS-scavenging glutathione biosynthesis. At a certain threshold, the heightened oxidative stress and antioxidant capability achieve a higher level of redox balance, on which cancer cells depend to gain a selective advantage on survival and proliferation. However, this prominent metabolic feature may irrevocably render cancer cells vulnerable to concurrent inhibition of both NOX activity and glutathione biosynthesis, which may be exploited as a novel therapeutic strategy. In this report, we test this hypothesis by treating the HRASG12V-transformed ovarian epithelial cells, mutant KRAS-harboring pancreatic and colon cancer cells of mouse and human origins, as well as cancer xenografts, with diphenyleneiodonium (DPI) and buthionine sulfoximine (BSO) combination, which inhibit NOX activity and glutathione biosynthesis, respectively. Our results demonstrate that concomitant targeting of NOX and glutathione biosynthesis induces a highly potent lethality to cancer cells harboring oncogenic RAS. Therefore, our studies provide a novel strategy against RAS-bearing cancers that warrants further mechanistic and translational investigation.
  38. Cancer Discov. 2021 Jan 27. pii: candisc.1211.2020. [Epub ahead of print]
      Metabolic reprogramming enables cancer cell growth, proliferation, and survival. This reprogramming is driven by the combined actions of oncogenic alterations in cancer cells and host cell factors acting on cancer cells in the tumor microenvironment. Cancer cell intrinsic mechanisms activate signal transduction components that either directly enhance metabolic enzyme activity or upregulate transcription factors that in turn increase expression of metabolic regulators. Extrinsic signaling mechanisms involve host-derived factors that further promote and amplify metabolic reprogramming in cancer cells. This review describes intrinsic and extrinsic mechanisms driving cancer metabolism in the tumor microenvironment and how such mechanisms may be targeted therapeutically.
  39. Signal Transduct Target Ther. 2021 Feb 19. 6(1): 70
      Long non-coding RNAs (lncRNAs) play key roles in colorectal carcinogenesis. Here, we aimed to identify the risk SNP-induced lncRNAs and to investigate their roles in colorectal carcinogenesis. First, we identified rs6695584 as the causative SNP in 1q41 locus. The A>G mutation of rs6695584 created a protein-binding motif of BATF, altered the enhancer activity, and subsequently activated lncSLCC1 expression. Further validation in two independent CRC cohorts confirmed the upregulation of lncSLCC1 in CRC tissues, and revealed that increased lncSLCC1 expression was associated with poor survival in CRC patients. Mechanistically, lncRNA-SLCC1 interacted with AHR and transcriptionally activated HK2 expression, the crucial enzyme in glucose metabolism, thereby driving the glycolysis pathway and accelerating CRC tumor growth. The functional assays revealed that lncSLCC1 induced glycolysis activation and tumor growth in CRC mediated by HK2. In addition, HK2 was upregulated in colorectal cancer tissues and positively correlated with lncSLCC1 expression and patient survival. Taken together, our findings reveal a risk SNP-mediated oncogene lncRNA-SLCC1 promotes CRC through activating the glycolysis pathway.
  40. Front Immunol. 2020 ;11 616367
      Solid tumors are often challenged by hypoxic and nutrient-deprived tumor microenvironments (TME) as tumors progress, due to limited perfusion and rapid nutrient consumption. While cancer cells can demonstrate the ability to survive in nutrient-deprived conditions through multiple intrinsic alterations, it is poorly understood how nutrient-deprived cancer cells co-opt the TME to promote cancer cell survival and tumor progression. In the present study, we found that glutamine deprivation markedly potentiated the expression of G-CSF and GM-CSF in mouse mammary cancer cells. The IRE1α-JNK pathway, which is activated by glutamine starvation, was found to be important for the upregulation of these cytokines. G-CSF and GM-CSF are well-known facilitators of myelopoiesis and mobilization of hematopoietic progenitor cells (HPC). Consistently, as tumors progressed, we found that several myeloid HPC compartments were gradually decreased in the bone marrow but were significantly increased in the spleen. Mechanistically, the HPC-maintaining capacity of the bone marrow was significantly impaired in tumor-bearing mice, with lower expression of HPC maintaining genes (i.e., CXCL12, SCF, ANGPT1, and VCAM1), and reduced levels of mesenchymal stem cells and CXCL12-producing cells. Furthermore, the mobilized HPCs that displayed the capacity for myelopoiesis were also found to accumulate in tumor tissue. Tumor-infiltrating HPCs were highly proliferative and served as important sources of immunosuppressive myeloid-derived suppressor cells (MDSCs) in the TME. Our work has identified an important role for glutamine starvation in regulating the expression of G-CSF and GM-CSF, and in facilitating the generation of immunosuppressive MDSCs in breast cancer.
    Keywords:  G-CSF; GM-CSF; MDSC; bone marrow; glutamine
  41. J Clin Invest. 2021 Feb 16. pii: 135963. [Epub ahead of print]
      Aberrant lipid metabolism promotes the development of skeletal muscle insulin resistance, but the exact identity of lipid-mediated mechanisms relevant to human obesity remains unclear. A comprehensive lipidomic analysis of primary myocytes from lean insulin-sensitive (LN) and obese insulin-resistant (OB) individuals revealed several species of lysophospholipids (lyso-PL) that were differentially-abundant. These changes coincided with greater expression of lysophosphatidylcholine acyltransferase 3 (LPCAT3), an enzyme involved in phospholipid transacylation (Lands cycle). Strikingly, mice with skeletal muscle-specific knockout of LPCAT3 (LPCAT3-MKO) exhibited greater muscle lyso-PC/PC, concomitant with improved skeletal muscle insulin sensitivity. Conversely, skeletal muscle-specific overexpression of LPCAT3 (LPCAT3-MKI) promoted glucose intolerance. The absence of LPCAT3 reduced phospholipid packing of cellular membranes and increased plasma membrane lipid clustering, suggesting that LPCAT3 affects insulin receptor phosphorylation by modulating plasma membrane lipid organization. In conclusion, obesity accelerates the skeletal muscle Lands cycle, whose consequence might induce the disruption of plasma membrane organization that suppresses muscle insulin action.
    Keywords:  Insulin signaling; Metabolism; Muscle Biology; Skeletal muscle
  42. Sci Rep. 2021 Feb 15. 11(1): 3813
      The maintenance of mitochondrial activity in hypothalamic neurons is determinant to the control of energy homeostasis in mammals. Disturbs in the mitochondrial proteostasis can trigger the mitonuclear imbalance and mitochondrial unfolded protein response (UPRmt) to guarantee the mitochondrial integrity and function. However, the role of mitonuclear imbalance and UPRmt in hypothalamic cells are unclear. Combining the transcriptomic analyses from BXD mice database and in vivo experiments, we demonstrated that physical training alters the mitochondrial proteostasis in the hypothalamus of C57BL/6J mice. This physical training elicited the mitonuclear protein imbalance, increasing the mtCO-1/Atp5a ratio, which was accompanied by high levels of UPRmt markers in the hypothalamus. Also, physical training increased the maximum mitochondrial respiratory capacity in the brain. Interestingly, the transcriptomic analysis across several strains of the isogenic BXD mice revealed that hypothalamic mitochondrial DNA-encoded genes were negatively correlated with body weight and several genes related to the orexigenic response. As expected, physical training reduced body weight and food intake. Interestingly, we found an abundance of mt-CO1, a mitochondrial DNA-encoded protein, in NPY-producing neurons in the lateral hypothalamus nucleus of exercised mice. Collectively, our data demonstrated that physical training altered the mitochondrial proteostasis and induced the mitonuclear protein imbalance and UPRmt in hypothalamic cells.
  43. Nat Rev Mol Cell Biol. 2021 Feb 16.
      Mitochondria are cellular organelles responsible for generation of chemical energy in the process called oxidative phosphorylation. They originate from a bacterial ancestor and maintain their own genome, which is expressed by designated, mitochondrial transcription and translation machineries that differ from those operating for nuclear gene expression. In particular, the mitochondrial protein synthesis machinery is structurally and functionally very different from that governing eukaryotic, cytosolic translation. Despite harbouring their own genetic information, mitochondria are far from being independent of the rest of the cell and, conversely, cellular fitness is closely linked to mitochondrial function. Mitochondria depend heavily on the import of nuclear-encoded proteins for gene expression and function, and hence engage in extensive inter-compartmental crosstalk to regulate their proteome. This connectivity allows mitochondria to adapt to changes in cellular conditions and also mediates responses to stress and mitochondrial dysfunction. With a focus on mammals and yeast, we review fundamental insights that have been made into the biogenesis, architecture and mechanisms of the mitochondrial translation apparatus in the past years owing to the emergence of numerous near-atomic structures and a considerable amount of biochemical work. Moreover, we discuss how cellular mitochondrial protein expression is regulated, including aspects of mRNA and tRNA maturation and stability, roles of auxiliary factors, such as translation regulators, that adapt mitochondrial translation rates, and the importance of inter-compartmental crosstalk with nuclear gene expression and cytosolic translation and how it enables integration of mitochondrial translation into the cellular context.
  44. J Clin Invest. 2021 Feb 15. pii: 140100. [Epub ahead of print]131(4):
      Rapidly proliferating tumor and immune cells need metabolic programs that support energy and biomass production. The amino acid glutamine is consumed by effector T cells and glutamine-addicted triple-negative breast cancer (TNBC) cells, suggesting that a metabolic competition for glutamine may exist within the tumor microenvironment, potentially serving as a therapeutic intervention strategy. Here, we report that there is an inverse correlation between glutamine metabolic genes and markers of T cell-mediated cytotoxicity in human basal-like breast cancer (BLBC) patient data sets, with increased glutamine metabolism and decreased T cell cytotoxicity associated with poor survival. We found that tumor cell-specific loss of glutaminase (GLS), a key enzyme for glutamine metabolism, improved antitumor T cell activation in both a spontaneous mouse TNBC model and orthotopic grafts. The glutamine transporter inhibitor V-9302 selectively blocked glutamine uptake by TNBC cells but not CD8+ T cells, driving synthesis of glutathione, a major cellular antioxidant, to improve CD8+ T cell effector function. We propose a "glutamine steal" scenario, in which cancer cells deprive tumor-infiltrating lymphocytes of needed glutamine, thus impairing antitumor immune responses. Therefore, tumor-selective targeting of glutamine metabolism may be a promising therapeutic strategy in TNBC.
    Keywords:  Amino acid metabolism; Breast cancer; Cancer immunotherapy; Oncology
  45. Nature. 2021 Feb 15.
      Limiting the metabolic competition in the tumor microenvironment (TME) may increase the effectiveness of immunotherapy. Because of its critical role in glucose metabolism of activated T cells, CD28 signaling has been proposed as a T-cell metabolic biosensor1. Conversely, CTLA-4 engagement has been shown to down-regulate T-cell glycolysis1. Here, we investigated the impact of CTLA-4 blockade on the metabolic fitness of intra-tumor T cells in relationship to the tumor glycolytic capacity. We found that CTLA-4 blockade promotes immune cell infiltration and metabolic fitness especially in glycolysis-low tumors. Accordingly, anti-CTLA-4 achieved better therapeutic outcomes in mice bearing glycolysis-defective tumors. Intriguingly, tumor-specific CD8+ T-cell responses correlated with phenotypic and functional destabilization of tumor-infiltrating regulatory T cells (Tregs) toward IFN-γ- and TNF-α-producing cells in glycolysis-defective tumors. By mimicking the highly and poorly glycolytic TME in vitro, we show that the effect of CTLA-4 blockade to promote Treg destabilization is dependent on Treg glycolysis and CD28 signaling. These findings indicate that decreasing tumor competition for glucose may facilitate the therapeutic activity of CTLA-4 blockade, thus supporting its combination with inhibitors of tumor glycolysis. Moreover, these results reveal a new mechanism through which anti-CTLA-4 interferes with Treg function in the presence of glucose.
  46. Proc Natl Acad Sci U S A. 2021 Feb 16. pii: e2020838118. [Epub ahead of print]118(7):
      We present a unifying theory to explain cancer recurrence, therapeutic resistance, and lethality. The basis of this theory is the formation of simultaneously polyploid and aneuploid cancer cells, polyaneuploid cancer cells (PACCs), that avoid the toxic effects of systemic therapy by entering a state of cell cycle arrest. The theory is independent of which of the classically associated oncogenic mutations have already occurred. PACCs have been generally disregarded as senescent or dying cells. Our theory states that therapeutic resistance is driven by PACC formation that is enabled by accessing a polyploid program that allows an aneuploid cancer cell to double its genomic content, followed by entry into a nondividing cell state to protect DNA integrity and ensure cell survival. Upon removal of stress, e.g., chemotherapy, PACCs undergo depolyploidization and generate resistant progeny that make up the bulk of cancer cells within a tumor.
    Keywords:  drug resistance; evolution; metastasis; tumor microenvironment; whole-genome doubling
  47. Elife. 2021 Feb 16. pii: e64690. [Epub ahead of print]10
      The mitochondrial carrier family protein SLC25A3 transports both copper and phosphate in mammals yet in Saccharomyces cerevisiae the transport of these substrates is partitioned across two paralogs: PIC2 and MIR1. To understand the ancestral state of copper and phosphate transport in mitochondria, we explored the evolutionary relationships of PIC2 and MIR1 orthologs across the eukaryotic tree of life. Phylogenetic analyses revealed that PIC2-like and MIR1-like orthologs are present in all major eukaryotic supergroups, indicating an ancient gene duplication created these paralogs. To link this phylogenetic signal to protein function, we used structural modelling and site-directed mutagenesis to identify residues involved in copper and phosphate transport. Based on these analyses, we generated a L175A variant of mouse SLC25A3 that retains the ability to transport copper but not phosphate. This work highlights the utility of using an evolutionary framework to uncover amino acids involved in substrate recognition by mitochondrial carrier family proteins.
    Keywords:  S. cerevisiae; biochemistry; chemical biology; evolutionary biology; mouse
  48. Cell Rep. 2021 Jan 26. pii: S2211-1247(20)31666-1. [Epub ahead of print]34(4): 108677
      Pioneering studies from the early 1980s suggested that bacterial peptidoglycan-derived muramyl peptides (MPs) could exert either stimulatory or immunosuppressive functions depending, in part, on chronicity of exposure. However, this Janus-faced property of MPs remains largely unexplored. Here, we demonstrate the immunosuppressive potential of Nod1, the bacterial sensor of diaminopimelic acid (DAP)-containing MPs. Using a model of self-limiting peritonitis, we show that systemic Nod1 activation promotes an autophagy-dependent reprogramming of macrophages toward an alternative phenotype. Moreover, Nod1 stimulation induces the expansion of myeloid-derived suppressor cells (MDSCs) and maintains their immunosuppressive potential via arginase-1 activity. Supporting the role of MDSCs and tumor-associated macrophages in cancer, we demonstrate that myeloid-intrinsic Nod1 expression sustains intra-tumoral arginase-1 levels to foster an immunosuppressive and tumor-permissive microenvironment during colorectal cancer (CRC) development. Our findings support the notion that bacterial products, via Nod1 detection, modulate the immunosuppressive activity of myeloid cells and fuel tumor progression in CRC.
    Keywords:  Atg16L1; MDSC; Nod1 receptor; TAM; TME; arginase-1; autophagy; colon cancer; immunosuppression; microbiota
  49. J Transl Med. 2021 Feb 16. 19(1): 71
      Skeletal muscle aging is associated with a decline in motor function and loss of muscle mass- a condition known as sarcopenia. The underlying mechanisms that drive this pathology are associated with a failure in energy generation in skeletal muscle, either from age-related decline in mitochondrial function, or from disuse. To an extent, lifelong exercise is efficacious in preserving the energetic properties of skeletal muscle and thus may delay the onset of sarcopenia. This review discusses the cellular and molecular changes in skeletal muscle mitochondria during the aging process and how different exercise modalities work to reverse these changes. A key factor that will be described is the efficiency of mitochondrial coupling-ATP production relative to O2 uptake in myocytes and how that efficiency is a main driver for age-associated decline in skeletal muscle function. With that, we postulate the most effective exercise modality and protocol for reversing the molecular hallmarks of skeletal muscle aging and staving off sarcopenia. Two other concepts pertinent to mitochondrial efficiency in exercise-trained skeletal muscle will be integrated in this review, including- mitophagy, the removal of dysfunctional mitochondrial via autophagy, as well as the implications of muscle fiber type changes with sarcopenia on mitochondrial function.
    Keywords:  Aging; Exercise; Mitochondria; Skeletal muscle