bims-camemi Biomed News
on Mitochondrial metabolism in cancer
Issue of 2022–09–18
forty-six papers selected by
Christian Frezza, Universität zu Köln
  1. Metabolic reprogramming in the OPA1-deficient cells.
  2. OMA1-mediated integrated stress response protects against ferroptosis in mitochondrial cardiomyopathy.
  3. Mitochondrial autophagy in the sleeping brain.
  4. Sex differences in brain tumor glutamine metabolism reveal sex-specific vulnerabilities to treatment.
  5. Brain-enriched RagB isoforms regulate the dynamics of mTORC1 activity through GATOR1 inhibition.
  6. How An Assembly Factor Enhances Covalent FAD Attachment to the Flavoprotein Subunit of Complex II.
  7. ISCA2 inhibition decreases HIF and induces ferroptosis in clear cell renal carcinoma.
  8. A happy cell stays home: When metabolic stress creates epigenetic advantages in the tumor microenvironment.
  9. Making the leap from structure to mechanism: are the open states of mammalian complex I identified by cryoEM resting states or catalytic intermediates?
  10. The intestine is a major contributor to circulating succinate in mice.
  11. Cytoskeletal vimentin regulates cell size and autophagy through mTORC1 signaling.
  12. The roles of intratumour heterogeneity in the biology and treatment of pancreatic ductal adenocarcinoma.
  13. A stochastic basis for metabolic phenotypes.
  14. Physiological media advance cell culture experiments.
  15. A Rag GTPase dimer code defines the regulation of mTORC1 by amino acids.
  16. Structure and mechanism of human cystine exporter cystinosin.
  17. Inhibition of UBA6 by inosine augments tumour immunogenicity and responses.
  18. mTOR-regulated mitochondrial metabolism limits mycobacterium-induced cytotoxicity.
  19. Eprenetapopt triggers ferroptosis, inhibits NFS1 cysteine desulfurase, and synergizes with serine and glycine dietary restriction.
  20. Nutrient-sensitive protein O-GlcNAcylation shapes daily biological rhythms.
  21. Lamin A/C impairments cause mitochondrial dysfunction by attenuating PGC1α and the NAMPT-NAD+ pathway.
  22. Structural basis for FLCN RagC GAP activation in MiT-TFE substrate-selective mTORC1 regulation.
  23. Solute exchange through gap junctions lessens the adverse effects of inactivating mutations in metabolite-handling genes.
  24. Exploration of metabolic responses towards hypoxia mimetic DMOG in cancer cells by using untargeted metabolomics.
  25. Cuproptosis identifies respiratory subtype of renal cancer that confers favorable prognosis.
  26. Independent phenotypic plasticity axes define distinct obesity sub-types.
  27. Isoform-resolved mRNA profiling of ribosome load defines interplay of HIF and mTOR dysregulation in kidney cancer.
  28. Gain-of-function genetic screens in human cells identify SLC transporters overcoming environmental nutrient restrictions.
  29. Glucose-driven TOR-FIE-PRC2 signalling controls plant development.
  30. A universal coupling mechanism of respiratory complex I.
  31. Mettl3-dependent m6A modification attenuates the brain stress response in Drosophila.
  32. Pathogenic variants of sphingomyelin synthase SMS2 disrupt lipid landscapes in the secretory pathway.
  33. ER Stress-Induced Sphingosine-1-Phosphate Lyase Phosphorylation Potentiates the Mitochondrial Unfolded Protein Response.
  34. Pathogenic putrescine.
  35. Tissue-restricted inhibition of mTOR using chemical genetics.
  36. Kinetic compartmentalization by unnatural reaction for itaconate production.
  37. Age alters the oncogenic trajectory toward luminal mammary tumors that activate unfolded proteins responses.
  38. Quantification of Intracellular Citrate Concentrations with Genetically Encoded Biosensors.
  39. Cell Competition Shapes Metastatic Latency and Relapse.
  40. Stem cell-derived synthetic embryos self-assemble by exploiting cadherin codes and cortical tension.
  41. Control of cell state transitions.
  42. Clonal hematopoiesis and differential outcomes after immune checkpoint blockade.
  43. ER as master regulator of membrane trafficking and organelle function.
  44. Ketogenic diet uncovers differential metabolic plasticity of brain cells.
  45. Unraveling a mystery: Why human cells require cholesterol.
  46. Adipocyte mesenchymal transition contributes to mammary tumor progression.
  1. Cell Mol Life Sci. 2022 Sep 14. 79(10): 517
    Metabolic reprogramming in the OPA1-deficient cells.
    Wenting Dai, Zhichao Wang, Qiong A Wang, David Chan, Lei Jiang.
      OPA1, a dynamin-related GTPase mutated in autosomal dominant optic atrophy, is essential for the fusion of the inner mitochondrial membrane. Although OPA1 deficiency leads to impaired mitochondrial morphology, the role of OPA1 in central carbon metabolism remains unclear. Here, we aim to explore the functional role and metabolic mechanism of OPA1 in cell fitness beyond the control of mitochondrial fusion. We applied [U-13C]glucose and [U-13C]glutamine isotope tracing techniques to OPA1-knockout (OPA1-KO) mouse embryonic fibroblasts (MEFs) compared to OPA1 wild-type (OPA1-WT) controls. Furthermore, the resulting tracing data were integrated by metabolic flux analysis to understand the underlying metabolic mechanism through which OPA1 deficiency reprograms cellular metabolism. OPA1-deficient MEFs were depleted of intracellular citrate, which was consistent with the decreased oxygen consumption rate in these cells with mitochondrial fission that is not balanced by mitochondrial fusion. Whereas oxidative glucose metabolism was impaired, OPA1-deficient cells activated glutamine-dependent reductive carboxylation and subsequently relied on this reductive metabolism to produce cytosolic citrate as a predominant acetyl-CoA source for de novo fatty acid synthesis. Prevention of cytosolic glutamine reductive carboxylation by GSK321, an inhibitor of isocitrate dehydrogenase 1 (IDH1), largely repressed lipid synthesis and blocked cell proliferation in OPA1-deficient MEFs. Our data support that, when glucose oxidation failed to support lipogenesis and proliferation in cells with unbalanced mitochondrial fission, OPA1 deficiency stimulated metabolic anaplerosis into glutamine-dependent reductive carboxylation in an IDH1-mediated manner.
    Keywords:  Cell growth; Citrate; De novo lipogenesis; OPA1 dysfunction; Oxidative metabolism; Reductive carboxylation
    DOI:  https://doi.org/10.1007/s00018-022-04542-5
  2. Cell Metab. 2022 Sep 08. pii: S1550-4131(22)00360-6. [Epub ahead of print]
    OMA1-mediated integrated stress response protects against ferroptosis in mitochondrial cardiomyopathy.
    Sofia Ahola, Pablo Rivera Mejías, Steffen Hermans, Srikanth Chandragiri, Patrick Giavalisco, Hendrik Nolte, Thomas Langer.
      Cardiomyopathy and heart failure are common manifestations in mitochondrial disease caused by deficiencies in the oxidative phosphorylation (OXPHOS) system of mitochondria. Here, we demonstrate that the cardiac-specific loss of the assembly factor Cox10 of the cytochrome c oxidase causes mitochondrial cardiomyopathy in mice, which is associated with OXPHOS deficiency, lysosomal defects, and an aberrant mitochondrial morphology. Activation of the mitochondrial peptidase Oma1 in Cox10-/- mice results in mitochondrial fragmentation and induction of the integrated stress response (ISR) along the Oma1-Dele1-Atf4 signaling axis. Ablation of Oma1 or Dele1 in Cox10-/- mice aggravates cardiomyopathy. ISR inhibition impairs the cardiac glutathione metabolism, limits the selenium-dependent accumulation of the glutathione peroxidase Gpx4, and increases lipid peroxidation in the heart, ultimately culminating in ferroptosis. Our results demonstrate a protective role of the Oma1-Dele1-mediated ISR in mitochondrial cardiomyopathy and link ferroptosis to OXPHOS deficiency and mitochondrial disease.
    Keywords:  Atf4; Dele1; Gpx4; Oma1; cardiomyopathy; ferroptosis; glutathione; integrated stress response; mitochondria; selenium
    DOI:  https://doi.org/10.1016/j.cmet.2022.08.017
  3. Front Cell Dev Biol. 2022 ;10 956394
    Mitochondrial autophagy in the sleeping brain.
    Sofia Mauri, Mariavittoria Favaro, Greta Bernardo, Gabriella M Mazzotta, Elena Ziviani.
      A significant percentage of the mitochondrial mass is replaced on a daily basis via mechanisms of mitochondrial quality control. Through mitophagy (a selective type of autophagy that promotes mitochondrial proteostasis) cells keep a healthy pool of mitochondria, and prevent oxidative stress and inflammation. Furthermore, mitophagy helps adapting to the metabolic demand of the cells, which changes on a daily basis. Core components of the mitophagy process are PINK1 and Parkin, which mutations are linked to Parkinson's Disease. The crucial role of PINK1/Parkin pathway during stress-induced mitophagy has been extensively studied in vitro in different cell types. However, recent advances in the field allowed discovering that mitophagy seems to be only slightly affected in PINK1 KO mice and flies, putting into question the physiological relevance of this pathway in vivo in the whole organism. Indeed, several cell-specific PINK1/Parkin-independent mitophagy pathways have been recently discovered, which appear to be activated under physiological conditions such as those that promote mitochondrial proteome remodeling during differentiation or in response to specific physiological stimuli. In this Mini Review we want to summarize the recent advances in the field, and add another level of complexity by focusing attention on a potentially important aspect of mitophagy regulation: the implication of the circadian clock. Recent works showed that the circadian clock controls many aspects of mitochondrial physiology, including mitochondrial morphology and dynamic, respiratory activity, and ATP synthesis. Furthermore, one of the essential functions of sleep, which is controlled by the clock, is the clearance of toxic metabolic compounds from the brain, including ROS, via mechanisms of proteostasis. Very little is known about a potential role of the clock in the quality control mechanisms that maintain the mitochondrial repertoire healthy during sleep/wake cycles. More importantly, it remains completely unexplored whether (dys)function of mitochondrial proteostasis feedbacks to the circadian clockwork.
    Keywords:  Parkinson’s disease; animal models; circadian rhythms; mitophagy; proteostasis
    DOI:  https://doi.org/10.3389/fcell.2022.956394
  4. Med (N Y). 2022 Sep 08. pii: S2666-6340(22)00365-8. [Epub ahead of print]
    Sex differences in brain tumor glutamine metabolism reveal sex-specific vulnerabilities to treatment.
    Jasmin Sponagel, Jill K Jones, Cheryl Frankfater, Shanshan Zhang, Olivia Tung, Kevin Cho, Kelsey L Tinkum, Hannah Gass, Elena Nunez, Douglas R Spitz, Prakash Chinnaiyan, Jacob Schaefer, Gary J Patti, Maya S Graham, Audrey Mauguen, Milan Grkovski, Mark P Dunphy, Simone Krebs, Jingqin Luo, Joshua B Rubin, Joseph E Ippolito.
       BACKGROUND: Brain cancer incidence and mortality rates are greater in males. Understanding the molecular mechanisms that underlie those sex differences could improve treatment strategies. Although sex differences in normal metabolism are well described, it is currently unknown whether they persist in cancerous tissue.
    METHODS: Using positron emission tomography (PET) imaging and mass spectrometry, we assessed sex differences in glioma metabolism in samples from affected individuals. We assessed the role of glutamine metabolism in male and female murine transformed astrocytes using isotope labeling, metabolic rescue experiments, and pharmacological and genetic perturbations to modulate pathway activity.
    FINDINGS: We found that male glioblastoma surgical specimens are enriched for amino acid metabolites, including glutamine. Fluoroglutamine PET imaging analyses showed that gliomas in affected male individuals exhibit significantly higher glutamine uptake. These sex differences were well modeled in murine transformed astrocytes, in which male cells imported and metabolized more glutamine and were more sensitive to glutaminase 1 (GLS1) inhibition. The sensitivity to GLS1 inhibition in males was driven by their dependence on glutamine-derived glutamate for α-ketoglutarate synthesis and tricarboxylic acid (TCA) cycle replenishment. Females were resistant to GLS1 inhibition through greater pyruvate carboxylase (PC)-mediated TCA cycle replenishment, and knockdown of PC sensitized females to GLS1 inhibition.
    CONCLUSION: Our results show that clinically important sex differences exist in targetable elements of metabolism. Recognition of sex-biased metabolism may improve treatments through further laboratory and clinical research.
    FUNDING: This work was supported by NIH grants, Joshua's Great Things, the Siteman Investment Program, and the Barnard Research Fund.
    Keywords:  TCA cycle; Translation to patients; cancer metabolism; glioma; glutaminase; glutamine metabolism; glutathione; pyruvate carboxylase; sex differences; solid-state NMR; α-ketoglutarate
    DOI:  https://doi.org/10.1016/j.medj.2022.08.005
  5. Nat Cell Biol. 2022 Sep;24(9): 1407-1421
    Brain-enriched RagB isoforms regulate the dynamics of mTORC1 activity through GATOR1 inhibition.
    Gianluca Figlia, Sandra Müller, Anna M Hagenston, Susanne Kleber, Mykola Roiuk, Jan-Philipp Quast, Nora Ten Bosch, Damian Carvajal Ibañez, Daniela Mauceri, Ana Martin-Villalba, Aurelio A Teleman.
      Mechanistic target of rapamycin complex 1 (mTORC1) senses nutrient availability to appropriately regulate cellular anabolism and catabolism. During nutrient restriction, different organs in an animal do not respond equally, with vital organs being relatively spared. This raises the possibility that mTORC1 is differentially regulated in different cell types, yet little is known about this mechanistically. The Rag GTPases, RagA or RagB bound to RagC or RagD, tether mTORC1 in a nutrient-dependent manner to lysosomes where mTORC1 becomes activated. Although the RagA and B paralogues were assumed to be functionally equivalent, we find here that the RagB isoforms, which are highly expressed in neurons, impart mTORC1 with resistance to nutrient starvation by inhibiting the RagA/B GTPase-activating protein GATOR1. We further show that high expression of RagB isoforms is observed in some tumours, revealing an alternative strategy by which cancer cells can retain elevated mTORC1 upon low nutrient availability.
    DOI:  https://doi.org/10.1038/s41556-022-00977-x
  6. J Biol Chem. 2022 Sep 08. pii: S0021-9258(22)00915-2. [Epub ahead of print] 102472
    How An Assembly Factor Enhances Covalent FAD Attachment to the Flavoprotein Subunit of Complex II.
    Elena Maklashina, T M Iverson, Gary Cecchini.
      The membrane-bound complex II family of proteins is composed of enzymes that catalyze succinate and fumarate interconversion coupled with reduction or oxidation of quinones within the membrane domain. The majority of complex II enzymes are protein heterotetramers with the different subunits harboring a variety of redox centers. These redox centers are used to transfer electrons between the site of succinate-fumarate oxidation/reduction and the membrane domain harboring the quinone. A covalently bound FAD cofactor is present in the flavoprotein subunit, and the covalent flavin linkage is absolutely required to enable the enzyme to oxidize succinate. Assembly of the covalent flavin linkage in eukaryotic cells and many bacteria requires additional protein assembly factors. Here, we provide mechanistic details for how the assembly factors work to enhance covalent flavinylation. Both prokaryotic SdhE and mammalian SDHAF2 enhance FAD binding to their respective apo-protein of complex II. These assembly factors also increase the affinity for dicarboxylates to the apo-protein-non-covalent FAD complex and stabilize the pre-assembly complex. These findings are corroborated by previous investigations of the roles of SdhE in enhancing covalent flavinylation in both bacterial succinate dehydrogenase and fumarate reductase flavoprotein subunits and of SDHAF2 in performing the same function for the human mitochondrial succinate dehydrogenase flavoprotein. In conclusion, we provide further insight into assembly factor involvement in building the complex II flavoprotein subunit active site required for succinate oxidation.
    Keywords:  Succinate dehydrogenase; complex II; covalent flavins; fumarate reductase; mitochondrial assembly factors
    DOI:  https://doi.org/10.1016/j.jbc.2022.102472
  7. Oncogene. 2022 Sep 12.
    ISCA2 inhibition decreases HIF and induces ferroptosis in clear cell renal carcinoma.
    Yangsook Song Green, Maria C Ferreira Dos Santos, Daniel G Fuja, Ethan C Reichert, Alexandre R Campos, Sophie J Cowman, Karen Acuña Pilarte, Jessica Kohan, Sheryl R Tripp, Elizabeth A Leibold, Deepika Sirohi, Neeraj Agarwal, Xiaohui Liu, Mei Yee Koh.
      Clear cell renal cell carcinoma (ccRCC), the most common form of kidney cancer, is typically initiated by inactivation of the von Hippel Lindau (VHL) gene, which results in the constitutive activation of the hypoxia inducible factors, HIF-1α and HIF-2α. Using a high throughput screen, we identify novel compounds that decrease HIF-1/2α levels and induce ferroptosis by targeting Iron Sulfur Cluster Assembly 2 (ISCA2), a component of the late mitochondrial Iron Sulfur Cluster (L-ISC) assembly complex. ISCA2 inhibition either pharmacologically or using siRNA decreases HIF-2α protein levels by blocking iron-responsive element (IRE)-dependent translation, and at higher concentrations, also decreases HIF-1α translation through unknown mechanisms. Additionally, ISCA2 inhibition triggers the iron starvation response, resulting in iron/metals overload and death via ferroptosis. ISCA2 levels are decreased in ccRCC compared to normal kidney, and decreased ISCA2 levels are associated with pVHL loss and with sensitivity to ferroptosis induced by ISCA2 inhibition. Strikingly, pharmacological inhibition of ISCA2 using an orally available ISCA2 inhibitor significantly reduced ccRCC xenograft growth in vivo, decreased HIF-α levels and increased lipid peroxidation, suggesting increased ferroptosis in vivo. Thus, the targeting of ISCA2 may be a promising therapeutic strategy to inhibit HIF-1/2α and to induce ferroptosis in pVHL deficient cells.
    DOI:  https://doi.org/10.1038/s41388-022-02460-1
  8. Front Oncol. 2022 ;12 962928
    A happy cell stays home: When metabolic stress creates epigenetic advantages in the tumor microenvironment.
    Eric A Hanse, Mei Kong.
      A paradox of fast-proliferating tumor cells is that they deplete extracellular nutrients that often results in a nutrient poor microenvironment in vivo. Having a better understanding of the adaptation mechanisms cells exhibit in response to metabolic stress will open new therapeutic windows targeting the tumor's extreme nutrient microenvironment. Glutamine is one of the most depleted amino acids in the tumor core and here, we provide insight into how important glutamine and its downstream by-product, α-ketoglutarate (αKG), are to communicating information about the nutrient environment. This communication is key in the cell's ability to foster adaptation. We highlight the epigenetic changes brought on when αKG concentrations are altered in cancer and discuss how depriving cells of glutamine may lead to cancer cell de-differentiation and the ability to grow and thrive in foreign environments. When we starve cells, they adapt to survive. Those survival "skills" allow them to go out looking for other places to live and metastasize. We further examine current challenges to modelling the metabolic tumor microenvironment in the laboratory and discuss strategies that consider current findings to target the tumor's poor nutrient microenvironment.
    Keywords:  alpha ketoglutarate; epigenetics; glutamine; glutaminolysis-inhibition; metabolism; tumor; tumor microenvironment
    DOI:  https://doi.org/10.3389/fonc.2022.962928
  9. Curr Opin Struct Biol. 2022 Sep 07. pii: S0959-440X(22)00126-9. [Epub ahead of print]77 102447
    Making the leap from structure to mechanism: are the open states of mammalian complex I identified by cryoEM resting states or catalytic intermediates?
    Injae Chung, Daniel N Grba, John J Wright, Judy Hirst.
      Respiratory complex I (NADH:ubiquinone oxidoreductase) is a multi-subunit, energy-transducing mitochondrial enzyme that is essential for oxidative phosphorylation and regulating NAD+/NADH pools. Despite recent advances in structural knowledge and a long history of biochemical analyses, the mechanism of redox-coupled proton translocation by complex I remains unknown. Due to its ability to separate molecules in a mixed population into distinct classes, single-particle electron cryomicroscopy has enabled identification and characterisation of different complex I conformations. However, deciding on their catalytic and/or regulatory properties to underpin mechanistic hypotheses, especially without detailed biochemical characterisation of the structural samples, has proven challenging. In this review we explore different mechanistic interpretations of the closed and open states identified in cryoEM analyses of mammalian complex I.
    DOI:  https://doi.org/10.1016/j.sbi.2022.102447
  10. FASEB J. 2022 10;36(10): e22546
    The intestine is a major contributor to circulating succinate in mice.
    Wenxin Tong, Sarah A Hannou, You Wang, Inna Astapova, Ashot Sargsyan, Ruby Monn, Venkataramana Thiriveedi, Diana Li, Jessica R McCann, John F Rawls, Jatin Roper, Guo-Fang Zhang, Mark A Herman.
      The tricarboxylic acid (TCA) cycle is the epicenter of cellular aerobic metabolism. TCA cycle intermediates facilitate energy production and provide anabolic precursors, but also function as intra- and extracellular metabolic signals regulating pleiotropic biological processes. Despite the importance of circulating TCA cycle metabolites as signaling molecules, the source of circulating TCA cycle intermediates remains uncertain. We observe that in mice, the concentration of TCA cycle intermediates in the portal blood exceeds that in tail blood indicating that the gut is a major contributor to circulating TCA cycle metabolites. With a focus on succinate as a representative of a TCA cycle intermediate with signaling activities and using a combination of gut microbiota depletion mouse models and isotopomer tracing, we demonstrate that intestinal microbiota is not a major contributor to circulating succinate. Moreover, we demonstrate that endogenous succinate production is markedly higher than intestinal succinate absorption in normal physiological conditions. Altogether, these results indicate that endogenous succinate production within the intestinal tissue is a major physiological source of circulating succinate. These results provide a foundation for an investigation into the role of the intestine in regulating circulating TCA cycle metabolites and their potential signaling effects on health and disease.
    Keywords:  TCA cycle intermediates; circulating biomarkers; intestine; succinate
    DOI:  https://doi.org/10.1096/fj.202200135RR
  11. PLoS Biol. 2022 Sep;20(9): e3001737
    Cytoskeletal vimentin regulates cell size and autophagy through mTORC1 signaling.
    Ponnuswamy Mohanasundaram, Leila S Coelho-Rato, Mayank Kumar Modi, Marta Urbanska, Franziska Lautenschläger, Fang Cheng, John E Eriksson.
      The nutrient-activated mTORC1 (mechanistic target of rapamycin kinase complex 1) signaling pathway determines cell size by controlling mRNA translation, ribosome biogenesis, protein synthesis, and autophagy. Here, we show that vimentin, a cytoskeletal intermediate filament protein that we have known to be important for wound healing and cancer progression, determines cell size through mTORC1 signaling, an effect that is also manifested at the organism level in mice. This vimentin-mediated regulation is manifested at all levels of mTOR downstream target activation and protein synthesis. We found that vimentin maintains normal cell size by supporting mTORC1 translocation and activation by regulating the activity of amino acid sensing Rag GTPase. We also show that vimentin inhibits the autophagic flux in the absence of growth factors and/or critical nutrients, demonstrating growth factor-independent inhibition of autophagy at the level of mTORC1. Our findings establish that vimentin couples cell size and autophagy through modulating Rag GTPase activity of the mTORC1 signaling pathway.
    DOI:  https://doi.org/10.1371/journal.pbio.3001737
  12. Oncogene. 2022 Sep 10.
    The roles of intratumour heterogeneity in the biology and treatment of pancreatic ductal adenocarcinoma.
    Theodore Evan, Victoria Min-Yi Wang, Axel Behrens.
      Intratumour heterogeneity (ITH) has become an important focus of cancer research in recent years. ITH describes the cellular variation that enables tumour evolution, including tumour progression, metastasis and resistance to treatment. The selection and expansion of genetically distinct treatment-resistant cancer cell clones provides one explanation for treatment failure. However, tumour cell variation need not be genetically encoded. In pancreatic ductal adenocarcinoma (PDAC) in particular, the complex tumour microenvironment as well as crosstalk between tumour and stromal cells result in exceptionally variable tumour cell phenotypes that are also highly adaptable. In this review we discuss four different types of phenotypic heterogeneity within PDAC, from morphological to metabolic heterogeneity. We suggest that these different types of ITH are not independent, but, rather, can inform one another. Lastly, we highlight recent findings that suggest how therapeutic efforts may halt PDAC progression by constraining cellular heterogeneity.
    DOI:  https://doi.org/10.1038/s41388-022-02448-x
  13. Nat Metab. 2022 Sep 12.
    A stochastic basis for metabolic phenotypes.
    Michelle L Holland, Vardhman K Rakyan.
      
    DOI:  https://doi.org/10.1038/s42255-022-00630-9
  14. Trends Biochem Sci. 2022 Sep 13. pii: S0968-0004(22)00232-8. [Epub ahead of print]
    Physiological media advance cell culture experiments.
    Martin Fischer.
      The metabolism plays a fundamental role in cellular signaling pathways, but commonly used cell culture media do not reflect physiological metabolite concentrations. The metabolic control hub mammalian target of rapamycin complex 1 (mTORC1) kinase is an illuminating example that it is about time to advance our cell culture to become more physiological and relevant.
    Keywords:  RFX7; cancer research; cell culture media; mTOR; metabolism; p53
    DOI:  https://doi.org/10.1016/j.tibs.2022.08.007
  15. Nat Cell Biol. 2022 Sep;24(9): 1394-1406
    A Rag GTPase dimer code defines the regulation of mTORC1 by amino acids.
    Peter Gollwitzer, Nina Grützmacher, Sabine Wilhelm, Daniel Kümmel, Constantinos Demetriades.
      Amino acid availability controls mTORC1 activity via a heterodimeric Rag GTPase complex that functions as a scaffold at the lysosomal surface, bringing together mTORC1 with its activators and effectors. Mammalian cells express four Rag proteins (RagA-D) that form dimers composed of RagA/B bound to RagC/D. Traditionally, the Rag paralogue pairs (RagA/B and RagC/D) are referred to as functionally redundant, with the four dimer combinations used interchangeably in most studies. Here, by using genetically modified cell lines that express single Rag heterodimers, we uncover a Rag dimer code that determines how amino acids regulate mTORC1. First, RagC/D differentially define the substrate specificity downstream of mTORC1, with RagD promoting phosphorylation of its lysosomal substrates TFEB/TFE3, while both Rags are involved in the phosphorylation of non-lysosomal substrates such as S6K. Mechanistically, RagD recruits mTORC1 more potently to lysosomes through increased affinity to the anchoring LAMTOR complex. Furthermore, RagA/B specify the signalling response to amino acid removal, with RagB-expressing cells maintaining lysosomal and active mTORC1 even upon starvation. Overall, our findings reveal key qualitative differences between Rag paralogues in the regulation of mTORC1, and underscore Rag gene duplication and diversification as a potentially impactful event in mammalian evolution.
    DOI:  https://doi.org/10.1038/s41556-022-00976-y
  16. Cell. 2022 Sep 09. pii: S0092-8674(22)01114-X. [Epub ahead of print]
    Structure and mechanism of human cystine exporter cystinosin.
    Xue Guo, Philip Schmiege, Tufa E Assafa, Rong Wang, Yan Xu, Linda Donnelly, Michael Fine, Xiaodan Ni, Jiansen Jiang, Glenn Millhauser, Liang Feng, Xiaochun Li.
      Lysosomal amino acid efflux by proton-driven transporters is essential for lysosomal homeostasis, amino acid recycling, mTOR signaling, and maintaining lysosomal pH. To unravel the mechanisms of these transporters, we focus on cystinosin, a prototypical lysosomal amino acid transporter that exports cystine to the cytosol, where its reduction to cysteine supplies this limiting amino acid for diverse fundamental processes and controlling nutrient adaptation. Cystinosin mutations cause cystinosis, a devastating lysosomal storage disease. Here, we present structures of human cystinosin in lumen-open, cytosol-open, and cystine-bound states, which uncover the cystine recognition mechanism and capture the key conformational states of the transport cycle. Our structures, along with functional studies and double electron-electron resonance spectroscopic investigations, reveal the molecular basis for the transporter's conformational transitions and protonation switch, show conformation-dependent Ragulator-Rag complex engagement, and demonstrate an unexpected activation mechanism. These findings provide molecular insights into lysosomal amino acid efflux and a potential therapeutic strategy.
    Keywords:  DEER; Keywords; Ragulator-Rag complex; X-ray crystallography; cryo-EM; cystinosin; cystinosis; fast adaptation; lysosomal storage disease; lysosomal transporter; membrane protein dynamics
    DOI:  https://doi.org/10.1016/j.cell.2022.08.020
  17. Nat Commun. 2022 Sep 15. 13(1): 5413
    Inhibition of UBA6 by inosine augments tumour immunogenicity and responses.
    Lei Zhang, Li Jiang, Liang Yu, Qin Li, Xiangjun Tian, Jingquan He, Ling Zeng, Yuqin Yang, Chaoran Wang, Yuhan Wei, Xiaoyue Jiang, Jing Li, Xiaolu Ge, Qisheng Gu, Jikun Li, Di Wu, Anthony J Sadler, Di Yu, Dakang Xu, Yue Gao, Xiangliang Yuan, Baokun He.
      Anti-cancer immunity and response to immune therapy is influenced by the metabolic states of the tumours. Immune checkpoint blockade therapy (ICB) is known to involve metabolic adaptation, however, the mechanism is not fully known. Here we show, by metabolic profiling of plasma samples from melanoma-bearing mice undergoing anti-PD1 and anti-CTLA4 combination therapy, that higher levels of purine metabolites, including inosine, mark ICB sensitivity. Metabolic profiles of ICB-treated human cancers confirm the association between inosine levels and ICB sensitivity. In mouse models, inosine supplementation sensitizes tumours to ICB, even if they are intrinsically ICB resistant, by enhancing T cell-mediated cytotoxicity and hence generating an immunologically hotter microenvironment. We find that inosine directly inhibits UBA6 in tumour cells, and lower level of UBA6 makes the tumour more immunogenic and this is reflected in favourable outcome following ICB therapy in human melanomas. Transplanted mouse melanoma and breast cancer cells with genetic ablation of Uba6 show higher sensitivity to ICB than wild type tumours. Thus, we provide evidence of an inosine-regulated UBA6-dependent pathway governing tumour-intrinsic immunogenicity and hence sensitivity to immune checkpoint inhibition, which might provide targets to overcome ICB resistance.
    DOI:  https://doi.org/10.1038/s41467-022-33116-z
  18. Cell. 2022 Sep 08. pii: S0092-8674(22)01112-6. [Epub ahead of print]
    mTOR-regulated mitochondrial metabolism limits mycobacterium-induced cytotoxicity.
    Antonio J Pagán, Lauren J Lee, Joy Edwards-Hicks, Cecilia B Moens, David M Tobin, Elisabeth M Busch-Nentwich, Erika L Pearce, Lalita Ramakrishnan.
      Necrosis of macrophages in the granuloma, the hallmark immunological structure of tuberculosis, is a major pathogenic event that increases host susceptibility. Through a zebrafish forward genetic screen, we identified the mTOR kinase, a master regulator of metabolism, as an early host resistance factor in tuberculosis. We found that mTOR complex 1 protects macrophages from mycobacterium-induced death by enabling infection-induced increases in mitochondrial energy metabolism fueled by glycolysis. These metabolic adaptations are required to prevent mitochondrial damage and death caused by the secreted mycobacterial virulence determinant ESAT-6. Thus, the host can effectively counter this early critical mycobacterial virulence mechanism simply by regulating energy metabolism, thereby allowing pathogen-specific immune mechanisms time to develop. Our findings may explain why Mycobacterium tuberculosis, albeit humanity's most lethal pathogen, is successful in only a minority of infected individuals.
    Keywords:  ESAT-6 mitotoxicity; Mycobacterium marinum; Mycobacterium tuberculosis; granuloma necrosis; mTOR; macrophage death; mitochondrial metabolism; oxidative phosphorylation; tuberculosis; zebrafish TB model
    DOI:  https://doi.org/10.1016/j.cell.2022.08.018
  19. Sci Adv. 2022 Sep 16. 8(37): eabm9427
    Eprenetapopt triggers ferroptosis, inhibits NFS1 cysteine desulfurase, and synergizes with serine and glycine dietary restriction.
    Kenji M Fujihara, Bonnie Z Zhang, Thomas D Jackson, Moses O Ogunkola, Brunda Nijagal, Julia V Milne, David A Sallman, Ching-Seng Ang, Iva Nikolic, Conor J Kearney, Simon J Hogg, Carlos S Cabalag, Vivien R Sutton, Sally Watt, Asuka T Fujihara, Joseph A Trapani, Kaylene J Simpson, Diana Stojanovski, Silke Leimkühler, Sue Haupt, Wayne A Phillips, Nicholas J Clemons.
      The mechanism of action of eprenetapopt (APR-246, PRIMA-1MET) as an anticancer agent remains unresolved, although the clinical development of eprenetapopt focuses on its reported mechanism of action as a mutant-p53 reactivator. Using unbiased approaches, this study demonstrates that eprenetapopt depletes cellular antioxidant glutathione levels by increasing its turnover, triggering a nonapoptotic, iron-dependent form of cell death known as ferroptosis. Deficiency in genes responsible for supplying cancer cells with the substrates for de novo glutathione synthesis (SLC7A11, SHMT2, and MTHFD1L), as well as the enzymes required to synthesize glutathione (GCLC and GCLM), augments the activity of eprenetapopt. Eprenetapopt also inhibits iron-sulfur cluster biogenesis by limiting the cysteine desulfurase activity of NFS1, which potentiates ferroptosis and may restrict cellular proliferation. The combination of eprenetapopt with dietary serine and glycine restriction synergizes to inhibit esophageal xenograft tumor growth. These findings reframe the canonical view of eprenetapopt from a mutant-p53 reactivator to a ferroptosis inducer.
    DOI:  https://doi.org/10.1126/sciadv.abm9427
  20. Open Biol. 2022 Sep;12(9): 220215
    Nutrient-sensitive protein O-GlcNAcylation shapes daily biological rhythms.
    Xianhui Liu, Joanna C Chiu.
      O-linked-N-acetylglucosaminylation (O-GlcNAcylation) is a nutrient-sensitive protein modification that alters the structure and function of a wide range of proteins involved in diverse cellular processes. Similar to phosphorylation, another protein modification that targets serine and threonine residues, O-GlcNAcylation occupancy on cellular proteins exhibits daily rhythmicity and has been shown to play critical roles in regulating daily rhythms in biology by modifying circadian clock proteins and downstream effectors. We recently reported that daily rhythm in global O-GlcNAcylation observed in Drosophila tissues is regulated via the integration of circadian and metabolic signals. Significantly, mistimed feeding, which disrupts coordination of these signals, is sufficient to dampen daily O-GlcNAcylation rhythm and is predicted to negatively impact animal biological rhythms and health span. In this review, we provide an overview of published and potential mechanisms by which metabolic and circadian signals regulate hexosamine biosynthetic pathway metabolites and enzymes, as well as O-GlcNAc processing enzymes to shape daily O-GlcNAcylation rhythms. We also discuss the significance of functional interactions between O-GlcNAcylation and other post-translational modifications in regulating biological rhythms. Finally, we highlight organ/tissue-specific cellular processes and molecular pathways that could be modulated by rhythmic O-GlcNAcylation to regulate time-of-day-specific biology.
    Keywords:  O-GlcNAcylation rhythm; biological rhythms; circadian input; hexosamine biosynthetic pathway; metabolic input; phosphorylation
    DOI:  https://doi.org/10.1098/rsob.220215
  21. Nucleic Acids Res. 2022 Sep 13. pii: gkac741. [Epub ahead of print]
    Lamin A/C impairments cause mitochondrial dysfunction by attenuating PGC1α and the NAMPT-NAD+ pathway.
    Scott Maynard, Arnaldur Hall, Panagiotis Galanos, Salvatore Rizza, Tatsuro Yamamoto, Helena Hagner Gram, Sebastian H N Munk, Muhammad Shoaib, Claus Storgaard Sørensen, Vilhelm A Bohr, Mads Lerdrup, Apolinar Maya-Mendoza, Jiri Bartek.
      Mutations in the lamin A/C gene (LMNA) cause laminopathies such as the premature aging Hutchinson Gilford progeria syndrome (HGPS) and altered lamin A/C levels are found in diverse malignancies. The underlying lamin-associated mechanisms remain poorly understood. Here we report that lamin A/C-null mouse embryo fibroblasts (Lmna-/- MEFs) and human progerin-expressing HGPS fibroblasts both display reduced NAD+ levels, unstable mitochondrial DNA and attenuated bioenergetics. This mitochondrial dysfunction is associated with reduced chromatin recruitment (Lmna-/- MEFs) or low levels (HGPS) of PGC1α, the key transcription factor for mitochondrial homeostasis. Lmna-/- MEFs showed reduced expression of the NAD+-biosynthesis enzyme NAMPT and attenuated activity of the NAD+-dependent deacetylase SIRT1. We find high PARylation in lamin A/C-aberrant cells, further decreasing the NAD+ pool and consistent with impaired DNA base excision repair in both cell models, a condition that fuels DNA damage-induced PARylation under oxidative stress. Further, ATAC-sequencing revealed a substantially altered chromatin landscape in Lmna-/- MEFs, including aberrantly reduced accessibility at the Nampt gene promoter. Thus, we identified a new role of lamin A/C as a key modulator of mitochondrial function through impairments of PGC1α and the NAMPT-NAD+ pathway, with broader implications for the aging process.
    DOI:  https://doi.org/10.1093/nar/gkac741
  22. Sci Adv. 2022 Sep 16. 8(37): eadd2926
    Structural basis for FLCN RagC GAP activation in MiT-TFE substrate-selective mTORC1 regulation.
    Rachel M Jansen, Roberta Peruzzo, Simon A Fromm, Adam L Yokom, Roberto Zoncu, James H Hurley.
      The mechanistic target of rapamycin complex 1 (mTORC1) regulates cell growth and catabolism in response to nutrients through phosphorylation of key substrates. The tumor suppressor folliculin (FLCN) is a RagC/D guanosine triphosphatase (GTPase)-activating protein (GAP) that regulates mTORC1 phosphorylation of MiT-TFE transcription factors, controlling lysosome biogenesis and autophagy. We determined the cryo-electron microscopy structure of the active FLCN complex (AFC) containing FLCN, FNIP2, the N-terminal tail of SLC38A9, the RagAGDP:RagCGDP.BeFx- GTPase dimer, and the Ragulator scaffold. Relative to the inactive lysosomal FLCN complex structure, FLCN reorients by 90°, breaks contact with RagA, and makes previously unseen contacts with RagC that position its Arg164 finger for catalysis. Disruption of the AFC-specific interfaces of FLCN and FNIP2 with RagC eliminated GAP activity and led to nuclear retention of TFE3, with no effect on mTORC1 substrates S6K or 4E-BP1. The structure provides a basis for regulation of an mTORC1 substrate-specific pathway and a roadmap to discover MiT-TFE family selective mTORC1 antagonists.
    DOI:  https://doi.org/10.1126/sciadv.add2926
  23. Elife. 2022 Sep 15. pii: e78425. [Epub ahead of print]11
    Solute exchange through gap junctions lessens the adverse effects of inactivating mutations in metabolite-handling genes.
    Stefania Monterisi, Johanna Michl, Alzbeta Hulikova, Jana Koth, Esther M Bridges, Amaryllis E Hill, Gulnar Abdullayeva, Walter F Bodmer, Pawel Swietach.
      Growth of cancer cells in vitro can be attenuated by genetically inactivating selected metabolic pathways. However, loss-of-function mutations in metabolic pathways are not negatively selected in human cancers, indicating that these genes are not essential in vivo. We hypothesize that spontaneous mutations in 'metabolic genes' will not necessarily produce functional defects because mutation-bearing cells may be rescued by metabolite exchange with neighboring wild-type cells via gap junctions. Using fluorescent substances to probe inter-cellular diffusion, we show that colorectal cancer (CRC) cells are coupled by gap junctions assembled from connexins, particularly Cx26. Cells with genetically inactivated components of pH regulation (SLC9A1), glycolysis (ALDOA), or mitochondrial respiration (NDUFS1) could be rescued through access to functional proteins in co-cultured wild-type cells. The effect of diffusive coupling was also observed in co-culture xenografts. Rescue was largely dependent on solute exchange via Cx26 channels, a uniformly and constitutively expressed isoform in CRCs. Due to diffusive coupling, the emergent phenotype is less heterogenous than its genotype, and thus an individual cell should not be considered as the unit under selection, at least for metabolite-handling processes. Our findings can explain why certain loss-of-function mutations in genes ascribed as 'essential' do not influence the growth of human cancers.
    Keywords:  biochemistry; cancer biology; chemical biology
    DOI:  https://doi.org/10.7554/eLife.78425
  24. Saudi J Biol Sci. 2022 Oct;29(10): 103426
    Exploration of metabolic responses towards hypoxia mimetic DMOG in cancer cells by using untargeted metabolomics.
    Mohammad Imran Khan.
      Hypoxia is considered as one of the most crucial elements of tumor microenvironment. The hypoxia inducible transcription factors (HIF-1/2) are used by the cancer cells to adapt hypoxic microenvironment through regulating the expression of various target genes, including metabolic enzymes. Dimethyloxalylglycine (DMOG), a hypoxic mimetic used for HIF stabilisation in cell and animal models, also demonstrates multiple metabolic effects. In past, it was shown that in cancer cells, DMOG treatment alters mitochondrial ATP production, glycolysis, respiration etc. However, a global landscape of metabolic level alteration in cancer cells during DMOG treatment is still not established. In the current work, the metabolic landscape of cancer cells during DMOG treatment is explored by using untargeted metabolomics approach. Results showed that DMOG treatment primarily alters the one carbon and lipid metabolism. The levels of one-carbon metabolism related metabolites like serine, ornithine, and homomethionine levels significantly altered during DMOG treatment. Further, DMOG treatment reduces the global fatty acyls like palmitic acids, stearic acids, and arachidonic acid levels in cancer cell lines. Additionally, we found an alteration in glycolytic metabolites known to be regulated by hypoxia in cancer cell lines. Collectively, the results provided novel insights into the metabolic impact of DMOG on cancer cells and showed that the use of DMOG to induce hypoxia yields similar metabolic features relative to physiological hypoxia.
    Keywords:  Dimethyloxalylglycine (DMOG); Fatty acyls; Hypoxia; Lipidomics; Untargeted Metabolomics
    DOI:  https://doi.org/10.1016/j.sjbs.2022.103426
  25. Apoptosis. 2022 Sep 14.
    Cuproptosis identifies respiratory subtype of renal cancer that confers favorable prognosis.
    Kunping Li, Linyi Tan, Yuqing Li, Yinfeng Lyu, Xinyi Zheng, Haowen Jiang, Xiaohua Zhang, Hui Wen, Chenchen Feng.
      Cuproptosis is a newly discovered cell death induced by excessive copper in mitochondria distinct from any known forms of apoptosis. Role of cuproptosis has not been well-reported in cancer, especially in clear-cell renal cell carcinoma (ccRCC). We comprehensively interrogated cuproptotic gene signature in ccRCC by reproducing multi-omics datasets and found cuproptosis was decreased in ccRCC compared with normal kidney. Cuproptosis identified a subgroup with significantly better prognosis. Functional annotation supported increased tricarboxylic acid cycle activity and decreased hypoxia signaling corroborated by metabolomics. Cuproptotic tumors showed decreased angiogenesis but were sensitive to Sunitinib and Sorafenib. Cuproptotic level in ccRCC cell lines showed robust negative correlation with copper ionophore Elesclomol. All findings support a respiratory subtype of ccRCC identified by cuproptosis.
    Keywords:  Clear cell renal cell carcinoma; Cuproptosis; Mitochondrial respiration
    DOI:  https://doi.org/10.1007/s10495-022-01769-2
  26. Nat Metab. 2022 Sep 12.
    Independent phenotypic plasticity axes define distinct obesity sub-types.
    PERMUTE
      Studies in genetically 'identical' individuals indicate that as much as 50% of complex trait variation cannot be traced to genetics or to the environment. The mechanisms that generate this 'unexplained' phenotypic variation (UPV) remain largely unknown. Here, we identify neuronatin (NNAT) as a conserved factor that buffers against UPV. We find that Nnat deficiency in isogenic mice triggers the emergence of a bi-stable polyphenism, where littermates emerge into adulthood either 'normal' or 'overgrown'. Mechanistically, this is mediated by an insulin-dependent overgrowth that arises from histone deacetylase (HDAC)-dependent β-cell hyperproliferation. A multi-dimensional analysis of monozygotic twin discordance reveals the existence of two patterns of human UPV, one of which (Type B) phenocopies the NNAT-buffered polyphenism identified in mice. Specifically, Type-B monozygotic co-twins exhibit coordinated increases in fat and lean mass across the body; decreased NNAT expression; increased HDAC-responsive gene signatures; and clinical outcomes linked to insulinemia. Critically, the Type-B UPV signature stratifies both childhood and adult cohorts into four metabolic states, including two phenotypically and molecularly distinct types of obesity.
    DOI:  https://doi.org/10.1038/s42255-022-00629-2
  27. Nat Struct Mol Biol. 2022 Sep 12.
    Isoform-resolved mRNA profiling of ribosome load defines interplay of HIF and mTOR dysregulation in kidney cancer.
    Yoichiro Sugimoto, Peter J Ratcliffe.
      Hypoxia inducible factor (HIF) and mammalian target of rapamycin (mTOR) pathways orchestrate responses to oxygen and nutrient availability. These pathways are frequently dysregulated in cancer, but their interplay is poorly understood, in part because of difficulties in simultaneous measurement of global and mRNA-specific translation. Here, we describe a workflow for measurement of ribosome load of mRNAs resolved by their transcription start sites (TSSs). Its application to kidney cancer cells reveals extensive translational reprogramming by mTOR, strongly affecting many metabolic enzymes and pathways. By contrast, global effects of HIF on translation are limited, and we do not observe reported translational activation by HIF2A. In contrast, HIF-dependent alterations in TSS usage are associated with robust changes in translational efficiency in a subset of genes. Analyses of the interplay of HIF and mTOR reveal that specific classes of HIF1A and HIF2A transcriptional target gene manifest different sensitivity to mTOR, in a manner that supports combined use of HIF2A and mTOR inhibitors in treatment of kidney cancer.
    DOI:  https://doi.org/10.1038/s41594-022-00819-2
  28. Life Sci Alliance. 2022 Nov;pii: e202201404. [Epub ahead of print]5(11):
    Gain-of-function genetic screens in human cells identify SLC transporters overcoming environmental nutrient restrictions.
    Manuele Rebsamen, Enrico Girardi, Vitaly Sedlyarov, Stefania Scorzoni, Konstantinos Papakostas, Manuela Vollert, Justyna Konecka, Bettina Guertl, Kristaps Klavins, Tabea Wiedmer, Giulio Superti-Furga.
      Solute carrier (SLC) transporters control fluxes of nutrients and metabolites across membranes and thereby represent a critical interface between the microenvironment and cellular and subcellular metabolism. Because of substantial functional overlap, the interplay and relative contributions of SLCs in response to environmental stresses remain poorly elucidated. To infer functional relationships between SLCs and metabolites, we developed a strategy to identify SLCs able to sustain cell viability and proliferation under growth-limiting concentrations of essential nutrients. One-by-one depletion of 13 amino acids required for cell proliferation enabled gain-of-function genetic screens using a SLC-focused CRISPR/Cas9-based transcriptional activation approach to uncover transporters relieving cells from growth-limiting metabolic bottlenecks. Among the transporters identified, we characterized the cationic amino acid transporter SLC7A3 as a gene that, when up-regulated, overcame low availability of arginine and lysine by increasing their uptake, whereas SLC7A5 was able to sustain cellular fitness upon deprivation of several neutral amino acids. Moreover, we identified metabolic compensation mediated by the glutamate/aspartate transporters SLC1A2 and SLC1A3 under glutamine-limiting conditions. Overall, this gain-of-function approach using human cells uncovered functional transporter-nutrient relationships and revealed that transport activity up-regulation may be sufficient to overcome environmental metabolic restrictions.
    DOI:  https://doi.org/10.26508/lsa.202201404
  29. Nature. 2022 Sep 14.
    Glucose-driven TOR-FIE-PRC2 signalling controls plant development.
    Ruiqiang Ye, Meiyue Wang, Hao Du, Shweta Chhajed, Jin Koh, Kun-Hsiang Liu, Jinwoo Shin, Yue Wu, Lin Shi, Lin Xu, Sixue Chen, Yijing Zhang, Jen Sheen.
      Nutrients and energy have emerged as central modulators of developmental programmes in plants and animals1-3. The evolutionarily conserved target of rapamycin (TOR) kinase is a master integrator of nutrient and energy signalling that controls growth. Despite its key regulatory roles in translation, proliferation, metabolism and autophagy2-5, little is known about how TOR shapes developmental transitions and differentiation. Here we show that glucose-activated TOR kinase controls genome-wide histone H3 trimethylation at K27 (H3K27me3) in Arabidopsis thaliana, which regulates cell fate and development6-10. We identify FERTILIZATION-INDEPENDENT ENDOSPERM (FIE), an indispensable component of Polycomb repressive complex 2 (PRC2), which catalyses H3K27me3 (refs. 6-8,10-12), as a TOR target. Direct phosphorylation by TOR promotes the dynamic translocation of FIE from the cytoplasm to the nucleus. Mutation of the phosphorylation site on FIE abrogates the global H3K27me3 landscape, reprogrammes the transcriptome and disrupts organogenesis in plants. Moreover, glucose-TOR-FIE-PRC2 signalling modulates vernalization-induced floral transition. We propose that this signalling axis serves as a nutritional checkpoint leading to epigenetic silencing of key transcription factor genes that specify stem cell destiny in shoot and root meristems and control leaf, flower and silique patterning, branching and vegetative-to-reproduction transition. Our findings reveal a fundamental mechanism of nutrient signalling in direct epigenome reprogramming, with broad relevance for the developmental control of multicellular organisms.
    DOI:  https://doi.org/10.1038/s41586-022-05171-5
  30. Nature. 2022 Sep 14.
    A universal coupling mechanism of respiratory complex I.
    Vladyslav Kravchuk, Olga Petrova, Domen Kampjut, Anna Wojciechowska-Bason, Zara Breese, Leonid Sazanov.
      Complex I is the first enzyme in the respiratory chain, which is responsible for energy production in mitochondria and bacteria1. Complex I couples the transfer of two electrons from NADH to quinone and the translocation of four protons across the membrane2, but the coupling mechanism remains contentious. Here we present cryo-electron microscopy structures of Escherichia coli complex I (EcCI) in different redox states, including catalytic turnover. EcCI exists mostly in the open state, in which the quinone cavity is exposed to the cytosol, allowing access for water molecules, which enable quinone movements. Unlike the mammalian paralogues3, EcCI can convert to the closed state only during turnover, showing that closed and open states are genuine turnover intermediates. The open-to-closed transition results in the tightly engulfed quinone cavity being connected to the central axis of the membrane arm, a source of substrate protons. Consistently, the proportion of the closed state increases with increasing pH. We propose a detailed but straightforward and robust mechanism comprising a 'domino effect' series of proton transfers and electrostatic interactions: the forward wave ('dominoes stacking') primes the pump, and the reverse wave ('dominoes falling') results in the ejection of all pumped protons from the distal subunit NuoL. This mechanism explains why protons exit exclusively from the NuoL subunit and is supported by our mutagenesis data. We contend that this is a universal coupling mechanism of complex I and related enzymes.
    DOI:  https://doi.org/10.1038/s41586-022-05199-7
  31. Nat Commun. 2022 Sep 14. 13(1): 5387
    Mettl3-dependent m6A modification attenuates the brain stress response in Drosophila.
    Alexandra E Perlegos, Emily J Shields, Hui Shen, Kathy Fange Liu, Nancy M Bonini.
      N6-methyladenosine (m6A), the most prevalent internal modification on eukaryotic mRNA, plays an essential role in various stress responses. The brain is uniquely vulnerable to cellular stress, thus defining how m6A sculpts the brain's susceptibility may provide insight to brain aging and disease-related stress. Here we investigate the impact of m6A mRNA methylation in the adult Drosophila brain with stress. We show that m6A is enriched in the adult brain and increases with heat stress. Through m6A-immunoprecipitation sequencing, we show 5'UTR Mettl3-dependent m6A is enriched in transcripts of neuronal processes and signaling pathways that increase upon stress. Mettl3 knockdown results in increased levels of m6A targets and confers resilience to stress. We find loss of Mettl3 results in decreased levels of nuclear m6A reader Ythdc1, and knockdown of Ythdc1 also leads to stress resilience. Overall, our data suggest that m6A modification in Drosophila dampens the brain's biological response to stress.
    DOI:  https://doi.org/10.1038/s41467-022-33085-3
  32. Elife. 2022 Sep 14. pii: e79278. [Epub ahead of print]11
    Pathogenic variants of sphingomyelin synthase SMS2 disrupt lipid landscapes in the secretory pathway.
    Tolulope Sokoya, Jan Parolek, Mads Møller Foged, Dmytro I Danylchuk, Manuel Bozan, Bingshati Sarkar, Angelika Hilderink, Michael Philippi, Lorenzo D Botto, Paulien A Terhal, Outi Mäkitie, Jacob Piehler, Yeongho Kim, Christopher G Burd, Andrey S Klymchenko, Kenji Maeda, Joost C M Holthuis.
      Sphingomyelin is a dominant sphingolipid in mammalian cells. Its production in the trans-Golgi traps cholesterol synthesized in the ER to promote formation of a sphingomyelin/sterol gradient along the secretory pathway. This gradient marks a fundamental transition in physical membrane properties that help specify organelle identify and function. We previously identified mutations in sphingomyelin synthase SMS2 that cause osteoporosis and skeletal dysplasia. Here we show that SMS2 variants linked to the most severe bone phenotypes retain full enzymatic activity but fail to leave the ER owing to a defective autonomous ER export signal. Cells harboring pathogenic SMS2 variants accumulate sphingomyelin in the ER and display a disrupted transbilayer sphingomyelin asymmetry. These aberrant sphingomyelin distributions also occur in patient-derived fibroblasts and are accompanied by imbalances in cholesterol organization, glycerophospholipid profiles and lipid order in the secretory pathway. We postulate that pathogenic SMS2 variants undermine the capacity of osteogenic cells to uphold nonrandom lipid distributions that are critical for their bone forming activity.
    Keywords:  biochemistry; cell biology; chemical biology; human
    DOI:  https://doi.org/10.7554/eLife.79278
  33. J Lipid Res. 2022 Sep 10. pii: S0022-2275(22)00112-2. [Epub ahead of print] 100279
    ER Stress-Induced Sphingosine-1-Phosphate Lyase Phosphorylation Potentiates the Mitochondrial Unfolded Protein Response.
    Asli D Yildirim, Mevlut Citir, Asli E Dogan, Zehra Veli, Zehra Yildirim, Ozlem Tufanli, Alexis Traynor Kaplan, Carsten Schultz, Ebru Erbay.
      The unfolded protein response (UPR) is an elaborate signaling network that evolved to maintain proteostasis in the endoplasmic reticulum (ER) and mitochondria (mt). These organelles are functionally and physically associated and consequently, their stress responses are often intertwined. It is unclear how these two adaptive stress responses are coordinated during ER stress. The inositol-requiring enzyme-1 (IRE1), a central ER stress sensor and proximal regulator of the UPRER, harbors dual kinase and endoribonuclease (RNase) activities. IRE1 RNase activity initiates the transcriptional layer of the UPRER, but IRE1's kinase substrate(s) and their functions are largely unknown. Here, we discovered that sphingosine 1-phosphate (S1P) lyase (SPL), the enzyme that degrades S1P, is a substrate for the mammalian IRE1 kinase. Our data show that IRE1-dependent SPL phosphorylation inhibits SPL's enzymatic activity, resulting in increased intracellular S1P levels. S1P has previously been shown to induce the activation of mitochondrial UPR (UPRmt) in nematodes. We determined that IRE1 kinase-dependent S1P induction during ER stress potentiates UPRmt signaling in mammalian cells. Phosphorylation of eukaryotic translation initiation factor 2α (eif2α) is recognized as a critical molecular event for UPRmt activation in mammalian cells. Our data further demonstrate that inhibition of the IRE1-SPL axis abrogates the activation of two eif2α kinases, namely double-stranded RNA-activated protein kinase (PKR) and PKR-like ER kinase (PERK) upon ER stress. These findings show that the IRE1-SPL axis plays a central role in coordinating the adaptive responses of both organelles to ER stress in mammalian cells.
    Keywords:  Adaptive Stress Response; Endoplasmic Reticulum; Endoribonuclease; Eukaryotic Translation Initiation Factor 2α; IRE1-SPL axis; Inositol-Requiring Enzyme-1; Kinase; Mitochondria; Proteostasis; Signaling Networks
    DOI:  https://doi.org/10.1016/j.jlr.2022.100279
  34. Sci Signal. 2022 Sep 13. 15(751): eade8161
    Pathogenic putrescine.
    Wei Wong.
      Urea metabolism in astrocytes that produce putrescine leads to memory impairment in Alzheimer's disease.
    DOI:  https://doi.org/10.1126/scisignal.ade8161
  35. Proc Natl Acad Sci U S A. 2022 Sep 20. 119(38): e2204083119
    Tissue-restricted inhibition of mTOR using chemical genetics.
    Douglas R Wassarman, Kondalarao Bankapalli, Leo J Pallanck, Kevan M Shokat.
      Mammalian target of rapamycin (mTOR) is a highly conserved eukaryotic protein kinase that coordinates cell growth and metabolism, and plays a critical role in cancer, immunity, and aging. It remains unclear how mTOR signaling in individual tissues contributes to whole-organism processes because mTOR inhibitors, like the natural product rapamycin, are administered systemically and target multiple tissues simultaneously. We developed a chemical-genetic system, termed selecTOR, that restricts the activity of a rapamycin analog to specific cell populations through targeted expression of a mutant FKBP12 protein. This analog has reduced affinity for its obligate binding partner FKBP12, which reduces its ability to inhibit mTOR in wild-type cells and tissues. Expression of the mutant FKBP12, which contains an expanded binding pocket, rescues the activity of this rapamycin analog. Using this system, we show that selective mTOR inhibition can be achieved in Saccharomyces cerevisiae and human cells, and we validate the utility of our system in an intact metazoan model organism by identifying the tissues responsible for a rapamycin-induced developmental delay in Drosophila.
    Keywords:  Drosophila; kinase inhibitor; mTOR; rapamycin; tissue specific
    DOI:  https://doi.org/10.1073/pnas.2204083119
  36. Nat Commun. 2022 Sep 12. 13(1): 5353
    Kinetic compartmentalization by unnatural reaction for itaconate production.
    Dae-Yeol Ye, Myung Hyun Noh, Jo Hyun Moon, Alfonsina Milito, Minsun Kim, Jeong Wook Lee, Jae-Seong Yang, Gyoo Yeol Jung.
      Physical compartmentalization of metabolism using membranous organelles in eukaryotes is helpful for chemical biosynthesis to ensure the availability of substrates from competitive metabolic reactions. Bacterial hosts lack such a membranous system, which is one of the major limitations for efficient metabolic engineering. Here, we employ kinetic compartmentalization with the introduction of an unnatural enzymatic reaction by an engineered enzyme as an alternative strategy to enable substrate availability from competitive reactions through kinetic isolation of metabolic pathways. As a proof of concept, we kinetically isolate the itaconate synthetic pathway from the tricarboxylic acid cycle in Escherichia coli, which is natively separated by mitochondrial membranes in Aspergillus terreus. Specifically, 2-methylcitrate dehydratase is engineered to alternatively catalyze citrate and kinetically secure cis-aconitate for efficient production using a high-throughput screening system. Itaconate production can be significantly improved with kinetic compartmentalization and its strategy has the potential to be widely applicable.
    DOI:  https://doi.org/10.1038/s41467-022-33033-1
  37. Aging Cell. 2022 Sep 15. e13665
    Age alters the oncogenic trajectory toward luminal mammary tumors that activate unfolded proteins responses.
    Edmund Charles Jenkins, Mrittika Chattopadhyay, Maria Gomez, Denis Torre, Avi Ma'ayan, Miguel Torres-Martin, Daniela Sia, Doris Germain.
      A major limitation in the use of mouse models in breast cancer research is that most mice develop estrogen receptor-alpha (ERα)-negative mammary tumors, while in humans, the majority of breast cancers are ERα-positive. Therefore, developing mouse models that best mimic the disease in humans is of fundamental need. Here, using an inducible MMTV-rtTA/TetO-NeuNT mouse model, we show that despite being driven by the same oncogene, mammary tumors in young mice are ERα-negative, while they are ERα-positive in aged mice. To further elucidate the mechanisms for this observation, we performed RNAseq analysis and identified genes that are uniquely expressed in aged female-derived mammary tumors. We found these genes to be involved in the activation of the ERα axis of the mitochondrial UPR and the ERα-mediated regulation of XBP-1s, a gene involved in the endoplasmic reticulum UPR. Collectively, our results indicate that aging alters the oncogenic trajectory towards the ERα-positive subtype of breast cancers, and that mammary tumors in aged mice are characterized by the upregulation of multiple UPR stress responses regulated by the ERα.
    Keywords:  ER stress; XBP-1; aged mammary gland; aging; endoplasmic reticulum; estrogen receptor-alpha; mitochondrial UPR; unfolded protein response
    DOI:  https://doi.org/10.1111/acel.13665
  38. Methods Mol Biol. 2023 ;2564 247-258
    Quantification of Intracellular Citrate Concentrations with Genetically Encoded Biosensors.
    Yufeng Zhao, Kei Takahashi-Yamashiro, Yi Shen, Robert E Campbell.
      Citrate is a central intracellular metabolite with roles in a variety of normal and aberrant biological processes. The methods for quantifying citrate concentration in cells can enable the study of the molecular mechanisms of citrate-related biological processes and diseases. Compared to existing analytical methods such as enzymatic assays and mass spectrometry, genetically encoded biosensors based on fluorescent proteins (FPs) offer the advantage of noninvasively tracking intracellular ion and small molecule dynamics with high spatial-temporal resolution. These biosensors are less toxic than chemosensors and can be targeted to specific organelles for subcellular imaging. Here we present a protocol for quantification of cytosolic and mitochondrial citrate in mammalian cells with recently reported genetically encoded biosensors for citrate.
    Keywords:  Biosensor; Citrate; Fluorescent protein; Mammalian cell; Microscopy
    DOI:  https://doi.org/10.1007/978-1-0716-2667-2_12
  39. Cancer Discov. 2022 Sep 13. pii: CD-22-0236. [Epub ahead of print]
    Cell Competition Shapes Metastatic Latency and Relapse.
    Kangsan Kim, Huocong Huang, Pravat Kumar Parida, Lan He, Mauricio Marquez-Palencia, Tanner C Reese, Payal Kapur, James Brugarolas, Rolf A Brekken, Srinivas Malladi.
      Cell competition, a fitness sensing process is essential for tissue homeostasis. Employing cancer metastatic latency models, we show that cell competition results in displacement of latent metastatic (Lat-M) cells from the primary tumor. Lat-M cells resist anoikis and survive as residual metastatic disease. Remodelled extracellular matrix facilitates Lat-M cell displacement and survival in circulation. Disrupting cell competition dynamics by depleting SPARC reduced displacement from orthotopic tumors and attenuated metastases. In contrast, depletion of SPARC post-extravasation in lung resident Lat-M cells increased metastatic outgrowth. Furthermore, multi-regional transcriptomic analyses of matched primary tumors and metachronous metastases from kidney cancer patients identified tumor subclones with latent metastatic traits. Kidney cancer enriched for these latent metastatic traits had rapid onset of metachronous metastases and significantly reduced disease-free survival. Thus, an unexpected consequence of cell competition is displacement of cells with latent metastatic potential, thereby shaping metastatic latency and relapse.
    DOI:  https://doi.org/10.1158/2159-8290.CD-22-0236
  40. Nat Cell Biol. 2022 Sep;24(9): 1341-1349
    Stem cell-derived synthetic embryos self-assemble by exploiting cadherin codes and cortical tension.
    Min Bao, Jake Cornwall-Scoones, Estefania Sanchez-Vasquez, Dong-Yuan Chen, Joachim De Jonghe, Shahriar Shadkhoo, Florian Hollfelder, Matt Thomson, David M Glover, Magdalena Zernicka-Goetz.
      Mammalian embryos sequentially differentiate into trophectoderm and an inner cell mass, the latter of which differentiates into primitive endoderm and epiblast. Trophoblast stem (TS), extraembryonic endoderm (XEN) and embryonic stem (ES) cells derived from these three lineages can self-assemble into synthetic embryos, but the mechanisms remain unknown. Here, we show that a stem cell-specific cadherin code drives synthetic embryogenesis. The XEN cell cadherin code enables XEN cell sorting into a layer below ES cells, recapitulating the sorting of epiblast and primitive endoderm before implantation. The TS cell cadherin code enables TS cell sorting above ES cells, resembling extraembryonic ectoderm clustering above epiblast following implantation. Whereas differential cadherin expression drives initial cell sorting, cortical tension consolidates tissue organization. By optimizing cadherin code expression in different stem cell lines, we tripled the frequency of correctly formed synthetic embryos. Thus, by exploiting cadherin codes from different stages of development, lineage-specific stem cells bypass the preimplantation structure to directly assemble a postimplantation embryo.
    DOI:  https://doi.org/10.1038/s41556-022-00984-y
  41. Nature. 2022 Sep 14.
    Control of cell state transitions.
    Oleksii S Rukhlenko, Melinda Halasz, Nora Rauch, Vadim Zhernovkov, Thomas Prince, Kieran Wynne, Stephanie Maher, Eugene Kashdan, Kenneth MacLeod, Neil O Carragher, Walter Kolch, Boris N Kholodenko.
      Understanding cell state transitions and purposefully controlling them is a longstanding challenge in biology. Here we present cell state transition assessment and regulation (cSTAR), an approach for mapping cell states, modelling transitions between them and predicting targeted interventions to convert cell fate decisions. cSTAR uses omics data as input, classifies cell states, and develops a workflow that transforms the input data into mechanistic models that identify a core signalling network, which controls cell fate transitions by influencing whole-cell networks. By integrating signalling and phenotypic data, cSTAR models how cells manoeuvre in Waddington's landscape1 and make decisions about which cell fate to adopt. Notably, cSTAR devises interventions to control the movement of cells in Waddington's landscape. Testing cSTAR in a cellular model of differentiation and proliferation shows a high correlation between quantitative predictions and experimental data. Applying cSTAR to different types of perturbation and omics datasets, including single-cell data, demonstrates its flexibility and scalability and provides new biological insights. The ability of cSTAR to identify targeted perturbations that interconvert cell fates will enable designer approaches for manipulating cellular development pathways and mechanistically underpinned therapeutic interventions.
    DOI:  https://doi.org/10.1038/s41586-022-05194-y
  42. Cancer Cell. 2022 Sep 09. pii: S1535-6108(22)00392-0. [Epub ahead of print]
    Clonal hematopoiesis and differential outcomes after immune checkpoint blockade.
    David Hsiehchen, Hannah J Sfreddo, Karena Zhao, Catherine Y Han, Luc G T Morris.
      
    DOI:  https://doi.org/10.1016/j.ccell.2022.08.024
  43. J Cell Biol. 2022 Oct 03. pii: e202205135. [Epub ahead of print]221(10):
    ER as master regulator of membrane trafficking and organelle function.
    Eva Maria Wenzel, Liv Anker Elfmark, Harald Stenmark, Camilla Raiborg.
      The endoplasmic reticulum (ER), which occupies a large portion of the cytoplasm, is the cell's main site for the biosynthesis of lipids and carbohydrate conjugates, and it is essential for folding, assembly, and biosynthetic transport of secreted proteins and integral membrane proteins. The discovery of abundant membrane contact sites (MCSs) between the ER and other membrane compartments has revealed that, in addition to its biosynthetic and secretory functions, the ER plays key roles in the regulation of organelle dynamics and functions. In this review, we will discuss how the ER regulates endosomes, lysosomes, autophagosomes, mitochondria, peroxisomes, and the Golgi apparatus via MCSs. Such regulation occurs via lipid and Ca2+ transfer and also via control of in trans dephosphorylation reactions and organelle motility, positioning, fusion, and fission. The diverse controls of other organelles via MCSs manifest the ER as master regulator of organelle biology.
    DOI:  https://doi.org/10.1083/jcb.202205135
  44. Sci Adv. 2022 Sep 16. 8(37): eabo7639
    Ketogenic diet uncovers differential metabolic plasticity of brain cells.
    Tim Düking, Lena Spieth, Stefan A Berghoff, Lars Piepkorn, Annika M Schmidke, Miso Mitkovski, Nirmal Kannaiyan, Leon Hosang, Patricia Scholz, Ali H Shaib, Lennart V Schneider, Dörte Hesse, Torben Ruhwedel, Ting Sun, Lisa Linhoff, Andrea Trevisiol, Susanne Köhler, Adrian Marti Pastor, Thomas Misgeld, Michael Sereda, Imam Hassouna, Moritz J Rossner, Francesca Odoardi, Till Ischebeck, Livia de Hoz, Johannes Hirrlinger, Olaf Jahn, Gesine Saher.
      To maintain homeostasis, the body, including the brain, reprograms its metabolism in response to altered nutrition or disease. However, the consequences of these challenges for the energy metabolism of the different brain cell types remain unknown. Here, we generated a proteome atlas of the major central nervous system (CNS) cell types from young and adult mice, after feeding the therapeutically relevant low-carbohydrate, high-fat ketogenic diet (KD) and during neuroinflammation. Under steady-state conditions, CNS cell types prefer distinct modes of energy metabolism. Unexpectedly, the comparison with KD revealed distinct cell type-specific strategies to manage the altered availability of energy metabolites. Astrocytes and neurons but not oligodendrocytes demonstrated metabolic plasticity. Moreover, inflammatory demyelinating disease changed the neuronal metabolic signature in a similar direction as KD. Together, these findings highlight the importance of the metabolic cross-talk between CNS cells and between the periphery and the brain to manage altered nutrition and neurological disease.
    DOI:  https://doi.org/10.1126/sciadv.abo7639
  45. Sci Adv. 2022 Sep 16. 8(37): eade5927
    Unraveling a mystery: Why human cells require cholesterol.
    Philip L Yeagle.
      Understanding how cholesterol binds to mammalian cells offers critical insights into the waxy substance's role in protein modulation and cell function.
    DOI:  https://doi.org/10.1126/sciadv.ade5927
  46. Cell Rep. 2022 Sep 13. pii: S2211-1247(22)01194-9. [Epub ahead of print]40(11): 111362
    Adipocyte mesenchymal transition contributes to mammary tumor progression.
    Qingzhang Zhu, Yi Zhu, Chelsea Hepler, Qianbin Zhang, Jiyoung Park, Christy Gliniak, Gervaise H Henry, Clair Crewe, Dawei Bu, Zhuzhen Zhang, Shangang Zhao, Thomas Morley, Na Li, Dae-Seok Kim, Douglas Strand, Yingfeng Deng, Jacob J Robino, Oleg Varlamov, Ruth Gordillo, Mikhail G Kolonin, Christine M Kusminski, Rana K Gupta, Philipp E Scherer.
      Obesity is associated with increased cancer incidence and progression. However, the relationship between adiposity and cancer remains poorly understood at the mechanistic level. Here, we report that adipocytes from tumor-invasive mammary fat undergo de-differentiation to fibroblast-like precursor cells during tumor progression and integrate into the tumor microenvironment. Single-cell sequencing reveals that these de-differentiated adipocytes lose their original identities and transform into multiple cell types, including myofibroblast- and macrophage-like cells, with their characteristic features involved in immune response, inflammation, and extracellular matrix remodeling. The de-differentiated cells are metabolically distinct from tumor-associated fibroblasts but exhibit comparable effects on tumor cell proliferation. Inducing de-differentiation by Xbp1s overexpression promotes tumor progression despite lower adiposity. In contrast, promoting lipid-storage capacity in adipocytes through MitoNEET overexpression curbs tumor growth despite greater adiposity. Collectively, the metabolic interplay between tumor cells and adipocytes induces adipocyte mesenchymal transition and contributes to reconfigure the stroma into a more tumor-friendly microenvironment.
    Keywords:  CP: Cancer; CP: Metabolism; adipocyte; breast cancer; de-differentiation; obesity
    DOI:  https://doi.org/10.1016/j.celrep.2022.111362
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