bims-mascan Biomed News
on Mass spectrometry in cancer research
Issue of 2020‒03‒29
twenty papers selected by
Giovanny Rodriguez Blanco
The Beatson Institute for Cancer Research


  1. Anal Chem. 2020 Mar 23.
    Ding J, Kind T, Zhu QF, Wang Y, Yan JW, Fiehn O, Feng YQ.
      Fatty acid esters of hydroxy fatty acids (FAHFAs) are a family of recently discovered lipids with important physiological functions in mammals and plants. However, low detection sensitivity in negative ionization mode mass spectrometry makes low abundant FAHFA challenging to analyze. A 2-dimethylaminoethylamine (DMED) based chemical derivatization strategy was recently reported to improve the MS sensitivity of FAHFAs by labeling FAHFAs with a positively ionizable tertiary amine group. To facilitate a reliable, high throughput and automatic annotation of these compounds, a DMED-FAHFA in-silico library containing 4,290 high-resolution tandem mass spectra covering 264 different FAHFA classes was developed. The construction of the library was based on the heuristic information from MS/MS fragmentation patterns of DMED-FAHFA authentic standards, and then applied the patterns to computer-generated DMED-FAHFAs. The developed DMED-FAHFA in-silico library was demonstrated to be compatible with library search software NIST MS Search and the LC-MS/MS data processing tool MS-DIAL to guarantee high throughput and automatic annotations. Applying the in-silico library in Arabidopsis thaliana samples for profiling FAHFAs by high resolution LC-MS/MS enabled the annotation of 19 DMED-FAHFAs from 16 families, including 4 novel compounds. Using the in-silico library largely decreased the false positive annotation rate in comparison to low resolution LC-MS/MS. The developed library, MS/MS spectra and development templates are freely available for commercial and non-commercial use at https://zenodo.org/record/3606905.
    DOI:  https://doi.org/10.1021/acs.analchem.0c00172
  2. J Vis Exp. 2020 Mar 08.
    Mohammad K, Jiang H, Hossain MI, Titorenko VI.
      Lipids are structurally diverse amphipathic molecules that are insoluble in water. Lipids are essential contributors to the organization and function of biological membranes, energy storage and production, cellular signaling, vesicular transport of proteins, organelle biogenesis, and regulated cell death. Because the budding yeast Saccharomyces cerevisiae is a unicellular eukaryote amenable to thorough molecular analyses, its use as a model organism helped uncover mechanisms linking lipid metabolism and intracellular transport to complex biological processes within eukaryotic cells. The availability of a versatile analytical method for the robust, sensitive, and accurate quantitative assessment of major classes of lipids within a yeast cell is crucial for getting deep insights into these mechanisms. Here we present a protocol to use liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) for the quantitative analysis of major cellular lipids of S. cerevisiae. The LC-MS/MS method described is versatile and robust. It enables the identification and quantification of numerous species (including different isobaric or isomeric forms) within each of the 10 lipid classes. This method is sensitive and allows identification and quantitation of some lipid species at concentrations as low as 0.2 pmol/µL. The method has been successfully applied to assessing lipidomes of whole yeast cells and their purified organelles. The use of alternative mobile phase additives for electrospray ionization mass spectrometry in this method can increase the efficiency of ionization for some lipid species and can be therefore used to improve their identification and quantitation.
    DOI:  https://doi.org/10.3791/60616
  3. Methods Mol Biol. 2020 ;2138 207-216
    Jayasena T, Bustamante S, Clement J, Welschinger R, Caplan GA, Sachdev PS, Braidy N.
      Nicotinamide adenine dinucleotide (NAD+) and its related metabolites (NADome) are important endogenous analytes that are thought to play important roles in cellular metabolism, inflammation, oxidative stress, cancer, neurodegeneration, and aging in mammals. However, these analytes are unstable during the collection of biological fluids, which is a major limiting factor for their quantitation. Herein, we describe a highly robust and quantitative method using liquid chromatography coupled to tandem mass spectrometry to quantify the NADome in whole blood, plasma, mononuclear cells, platelets, cerebrospinal fluid (CSF), and urine. This methodology represents a "gold standard" of measure for understanding biological pathways and developing targeted pharmacological interventions to modulate NAD+ biosynthesis and NAD-dependent mediators in health and disease.
    Keywords:  Aging; Cancer; Inflammation; Metabolism; NAD+; NADome; Neurodegeneration; Oxidative stress
    DOI:  https://doi.org/10.1007/978-1-0716-0471-7_13
  4. Anal Chem. 2020 Mar 26.
    Heuillet M, Millard P, Y Cissé M, K Linares L, Letisse F, Manie SN, Le Cam L, Portais JC, Bellvert F.
      Studies of the topology, functioning and regulation of metabolic systems are based on two main types of information which can be measured by mass spectrometry: the (absolute or relative) concentration of metabolites and their isotope incorporation in 13C-labeling experiments. These data are currently obtained from two independent experiments because the 13C-labeled internal standard (IS) used to determine the concentration of a given metabolite overlaps the 13C-mass fractions from which its 13C-isotopologue distribution (CID) is quantified. Here, we developed a generic method with a dedicated processing workflow to obtain these two information simultaneously in a unique sample collected from a single cultivation, thereby reducing by a factor of two both the number of cultivations to perform and the number of samples to collect, prepare and analyze. The proposed approach is based on an IS labeled with other isotope(s) which can be resolved from the 13C-mass fractions of interest. As proof-of-principle, we analyzed amino acids using a doubly labeled 15N13C-cell extract as IS. Extensive evaluation of the proposed approach shows a similar accuracy and precision compared to state-of-the-art approaches. We demonstrate the value of this approach by investigating the dynamic response of amino acids metabolism in mammalian cells upon activation of the PERK kinase, a key component of the unfolded protein response. Integration of metabolite concentrations and isotopic profiles reveals a reduced de novo biosynthesis of amino-acids upon PERK activation. The proposed approach is generic and can be applied to other (micro)organisms, analytical platforms, isotopic tracers, or classes of metabolites.
    DOI:  https://doi.org/10.1021/acs.analchem.9b05709
  5. Anal Chem. 2020 Mar 27.
    Xiang P, Zhu Y, Yang Y, Zhao Z, Williams SM, Moore RJ, Kelly RT, Smith RD, Liu S.
      In many areas of application, key objectives of chemical separation and analysis are to minimize the sample quantity while maximizing the chemical information obtained. Increasing measurement sensitivity is especially critical for proteomics research, especially when processing trace samples and where multiple measurements are desired. A rich collection of technologies has been developed, but the resulting sensitivity remains insufficient for achieving in-depth coverage of proteomic samples as small as single cells. Here, we combine picoliter-scale liquid chromatography (picoLC) with mass spectrometry (MS) to address this issue. The picoLC employs a 2-μm-i.d. open tubular column to reduce the sample input needed to greatly increase the sensitivity achieved using electrospray ionization (ESI) with MS. With this picoLC-MS system, we show that we can identify ∼1000 proteins reliably using only 75 pg of tryptic peptides, representing a 10-100-fold sensitivity improvement compared with the state-of-the-art liquid chromatography (LC) or capillary electrophoresis (CE)-MS methods. PicoLC-MS extends the limit of separation science and is expected to be a powerful tool for single cell proteomics.
    DOI:  https://doi.org/10.1021/acs.analchem.9b05639
  6. Mol Metab. 2020 Feb 14. pii: S2212-8778(20)30007-7. [Epub ahead of print] 100941
    Trefely S, Lovell CD, Snyder NW, Wellen KE.
      BACKGROUND: Many metabolites serve as important signalling molecules to adjust cellular activities and functions based on nutrient availability. Links between acetyl-CoA metabolism, histone lysine acetylation, and gene expression have been documented and studied over the past decade. In recent years, several additional acyl modifications to histone lysine residues have been identified, which depend on acyl-coenzyme A thioesters (acyl-CoAs) as acyl donors. Acyl-CoAs are intermediates of multiple distinct metabolic pathways, and substantial evidence has emerged that histone acylation is metabolically sensitive. Nevertheless, the metabolic sources of acyl-CoAs used for chromatin modification in most cases remain poorly understood. Elucidating how these diverse chemical modifications are coupled to and regulated by cellular metabolism is important in deciphering their functional significance.SCOPE OF REVIEW: In this article, we review the metabolic pathways that produce acyl-CoAs, as well as emerging evidence for functional roles of diverse acyl-CoAs in chromatin regulation. Because acetyl-CoA has been extensively reviewed elsewhere, we will focus on four other acyl-CoA metabolites integral to major metabolic pathways that are also known to modify histones: succinyl-CoA, propionyl-CoA, crotonoyl-CoA, and butyryl-CoA. We also briefly mention several other acyl-CoA species, which present opportunities for further research; malonyl-CoA, glutaryl-CoA, 3-hydroxybutyryl-CoA, 2-hydroxyisobutyryl-CoA, and lactyl-CoA. Each acyl-CoA species has distinct roles in metabolism, indicating the potential to report shifts in the metabolic status of the cell. For each metabolite, we consider the metabolic pathways in which it participates and the nutrient sources from which it is derived, the compartmentalisation of its metabolism, and the factors reported to influence its abundance and potential nuclear availability. We also highlight reported biological functions of these metabolically-linked acylation marks. Finally, we aim to illuminate key questions in acyl-CoA metabolism as they relate to the control of chromatin modification.
    MAJOR CONCLUSIONS: A majority of acyl-CoA species are annotated to mitochondrial metabolic processes. Since acyl-CoAs are not known to be directly transported across mitochondrial membranes, they must be synthesized outside of mitochondria and potentially within the nucleus to participate in chromatin regulation. Thus, subcellular metabolic compartmentalisation likely plays a key role in the regulation of histone acylation. Metabolite tracing in combination with targeting of relevant enzymes and transporters will help to map the metabolic pathways that connect acyl-CoA metabolism to chromatin modification. The specific function of each acyl-CoA may be determined in part by biochemical properties that affect its propensity for enzymatic versus non-enzymatic protein modification, as well as the various enzymes that can add, remove and bind each modification. Further, competitive and inhibitory effects of different acyl-CoA species on these enzymes make determining the relative abundance of acyl-CoA species in specific contexts important to understand the regulation of chromatin acylation. An improved and more nuanced understanding of metabolic regulation of chromatin and its roles in physiological and disease-related processes will emerge as these questions are answered.
    Keywords:  Acyl-CoA; Acylation; Compartmentalisation; Histone; Metabolism
    DOI:  https://doi.org/10.1016/j.molmet.2020.01.005
  7. J Clin Invest. 2020 Mar 23. pii: 131800. [Epub ahead of print]
    Zhou X, He C, Ren J, Dai C, Stevens SR, Wang Q, Zamler D, Shingu T, Yuan L, Chandregowda CR, Wang Y, Ravikumar V, Rao AU, Zhou F, Zheng H, Rasband MN, Chen Y, Lan F, Heimberger AB, Segal BM, Hu J.
      Lipid-rich myelin forms electrically insulating, axon-wrapping multilayers that are essential for neural function, and mature myelin is traditionally considered metabolically inert. Surprisingly, we discovered that mature myelin lipids undergo rapid turnover, and quaking (Qki) is a major regulator of myelin lipid homeostasis. Oligodendrocyte-specific Qki depletion, without affecting oligodendrocyte survival, resulted in rapid demyelination, within 1 week, and gradually neurological deficits in adult mice. Myelin lipids, especially the monounsaturated fatty acids and very-long-chain fatty acids, were dramatically reduced by Qki depletion, whereas the major myelin proteins remained intact, and the demyelinating phenotypes of Qki-depleted mice were alleviated by a high-fat diet. Mechanistically, Qki serves as a coactivator of the PPARβ-RXRα complex, which controls the transcription of lipid-metabolism genes, particularly those involved in fatty acid desaturation and elongation. Treatment of Qki-depleted mice with PPARβ/RXR agonists significantly alleviated neurological disability and extended survival durations. Furthermore, a subset of lesions from patients with primary progressive multiple sclerosis were characterized by preferential reductions in myelin lipid contents, activities of various lipid metabolism pathways, and expression level of QKI-5 in human oligodendrocytes. Together, our results demonstrate that continuous lipid synthesis is indispensable for mature myelin maintenance and highlight an underappreciated role of lipid metabolism in demyelinating diseases.
    Keywords:  Demyelinating disorders; Metabolism; Multiple sclerosis; Neuroscience
    DOI:  https://doi.org/10.1172/JCI131800
  8. J Proteome Res. 2020 Mar 27.
    Pinto FG, Mahmud I, Harmon TA, Rubio VY, Garrett TJ.
      Spectrometric methods with rapid biomarker detection capacity through untargeted metabolomics are becoming essential in the clinical cancer research. Liquid chromatography-mass spectrometry (LC-MS) is a rapidly developing metabolomic-based biomarker technique due its high sensitivity, reproducibility, and separation efficiency. However, its transla-tion to clinical diagnostics is often limited due to long data acquisition times (~20 min/sample) and laborious sample extraction procedures when employed for large-scale metabolomics studies. Here, we developed a segmented flow approach coupled with high-resolution mass spectrometry (SF-HRMS) for untargeted metabolomics which has the capability to acquire data in less than 1.5 min/sample with robustness and reproducibility relative to LC-HRMS. The SF-HRMS results demonstrate the capability for screening metabolite-based urinary biomarkers associated with prostate cancer (PCa). The study shows that SF-HRMS-based global metabolomics has the potential to evolve into a rapid biomarker screening tool for clinical research.
    DOI:  https://doi.org/10.1021/acs.jproteome.0c00006
  9. Anal Chim Acta. 2020 Apr 22. pii: S0003-2670(20)30191-4. [Epub ahead of print]1107 92-100
    Wang T, Chen X, Luan C, Wu J.
      Cell heterogeneity of tumor tissues is one of the causes of cancer recurrence after chemotherapy. Cell subtype identification in tumor tissues of specific cancer is critical for precision medicine and prognosis. As the structural and functional components of cells, lipids are closely related to the apparent morphology of cells. They are potential biomarkers of species of cancers and can be used to classify different cancer cell types, but it remains a challenge to establish a stable cell differentiation model and extend it to tumor tissue cell subtype differentiation. Here we describe a lipid profiling method based on nanostructure assisted laser desorption/ionization mass spectrometry (NALDI-MS), which could classify five hepatocellular carcinoma (HCC) cell lines and discriminate subtype of mixed cells and tumor tissues. The NALDI target was patterned with array of sample spots containing vertical silicon nanowires (Si NWs). Owing to its high ability to absorb laser energy, the vertical Si NWs can help to generate abundant lipid ions of cell extracts without need of organic matrix. Combined with statistical analysis methods, twenty-two ion peaks distributed in four MS peak clusters were selected as potential biomarkers to distinguish the subtype of the five HCC cell lines. Peak normalization was performed within each MS peak cluster to reduce the variation of peak intensity in batch to batch analysis. Compared to full-spectrum normalization method, the inner-cluster normalization method could help to distinguish cell subtype more stably and accurately. The molecular structure of these biomarkers was identified and sorted into two classes including phosphatidylcholine (PE, PI, PG, PA, PS) and glycosphingolipid (LacCer, ST). Furthermore, the established method was successfully applied to identify the major HCC cell subtype in mixed cell samples and xenograft tumor tissues as well as drug response test, showing great potential in precision medicine and prognosis.
    Keywords:  Cell and tissue subtype; Drug susceptibility; High throughput silicon nanowires array chip; Lipidomics; Matrix-free laser desorption/ionization
    DOI:  https://doi.org/10.1016/j.aca.2020.02.019
  10. Genes Dis. 2020 Jun;7(2): 185-198
    Lane AN, Higashi RM, Fan TW.
      The genetic alterations associated with cell transformation are in large measure expressed in the metabolic phenotype as cancer cells proliferate and change their local environment, and prepare for metastasis. Qualitatively, the fundamental biochemistry of cancer cells is generally the same as in the untransformed cells, but the cancer cells produce a local environment, the TME, that is hostile to the stromal cells, and compete for nutrients. In order to proliferate, cells need sufficient nutrients, either those that cannot be made by the cells themselves, or must be made from simpler precursors. However, in solid tumors, the nutrient supply is often limiting given the potential for rapid proliferation, and the poor quality of the vasculature. Thus, cancer cells may employ a variety of strategies to obtain nutrients for survival, growth and metastasis. Although much has been learned using established cell lines in standard culture conditions, it is becoming clear from in vivo metabolic studies that this can also be misleading, and which nutrients are used for energy production versus building blocks for synthesis of macromolecules can vary greatly from tumor to tumor, and even within the same tumor. Here we review the operation of metabolic networks, and how recent understanding of nutrient supply in the TME and utilization are being revealed using stable isotope tracers in vivo as well as in vitro.
    Keywords:  2OG, 2-oxoglutarate; ACO1,2, aconitase 1,2; CP-MAS, Cross polarization Magic Angle Spinning; Cancer metabolism; DMEM, Dulbeccos Modified Eagles Medium; ECAR, extracellular acidification rate; ECM, extracellular matrix; EMP, Embden-Meyerhof Pathway; IDH1,2, isocitrate dehydrogenase 1,2 (NADP+dependent); IF, interstitial fluid; ME, malic enzyme; Metabolic flux; Nutrient supply; RPMI, Roswell Park Memorial Institute; SIRM, Stable Isotope Resolved Metabolomics; Stable isotope resolved metabolomics; TIL, tumor infiltrating lymphocyte; TIM/TPI, triose phosphate isomerase; TME, Tumor Micro Environment; Tumor microenvironment
    DOI:  https://doi.org/10.1016/j.gendis.2019.10.007
  11. J Biol Chem. 2020 Mar 23. pii: jbc.RA119.012420. [Epub ahead of print]
    Bradley MC, Yang K, Fernández-Del-Río L, Ngo J, Ayer A, Tsui HS, Novales NA, Stocker R, Shirihai OS, Barros MH, Clarke CF.
      Coenzyme Q (Qn) is a vital lipid component of the electron transport chain that functions in cellular energy metabolism and as a membrane antioxidant. In the yeast Saccharomyces cerevisiae, coq1-coq9 deletion mutants are respiratory incompetent, sensitive to lipid peroxidation stress, and unable to synthesize Q6 The yeast coq10 deletion mutant is also respiratory deficient and sensitive to lipid peroxidation, yet continues to produce Q6 at an impaired rate. Thus, Coq10 is required for the function of Q6 in respiration and as an antioxidant and is believed to chaperone Q6 from its site of synthesis to the respiratory complexes. In several fungi, Coq10 is encoded as a fusion polypeptide with Coq11, a recently identified protein of unknown function required for efficient Q6 biosynthesis. Because "fused" proteins are often involved in similar biochemical pathways, here we examined the putative functional relationship between Coq10 and Coq11 in yeast. We used plate growth and Seahorse assays and LC-MS/MS analysis to show that COQ11 deletion rescues respiratory deficiency, sensitivity to lipid peroxidation, and decreased Q6 biosynthesis of the coq10Δ mutant. Additionally, immunoblotting indicated that yeast coq11Δ mutants accumulate increased amounts of certain Coq polypeptides and display a stabilized CoQ synthome. These effects suggest that Coq11 modulates Q6 biosynthesis, and that its absence increases mitochondrial Q6 content in the coq10Δcoq11Δ double mutant. This augmented mitochondrial Q6 content counteracts the respiratory deficiency and lipid peroxidation sensitivity phenotypes of the coq10Δ mutant. This study further clarifies the intricate connection between Q6 biosynthesis, trafficking, and function in mitochondrial metabolism.
    Keywords:  CoQ synthome; Coenzyme Q; Coq10; Coq11; Saccharomyces cerevisiae; lipid; mitochondrial metabolism; ubiquinone; yeast
    DOI:  https://doi.org/10.1074/jbc.RA119.012420
  12. Biomolecules. 2020 Mar 24. pii: E493. [Epub ahead of print]10(3):
    Martínez-Sena T, Soluyanova P, Guzmán C, Valdivielso JM, Castell JV, Jover R.
      The vitamin D receptor (VDR) must be relevant to liver lipid metabolism because VDR deficient mice are protected from hepatosteatosis. Therefore, our objective was to define the role of VDR on the overall lipid metabolism in human hepatocytes. We developed an adenoviral vector for human VDR and performed transcriptomic and metabolomic analyses of cultured human hepatocytes upon VDR activation by vitamin D (VitD). Twenty percent of the VDR responsive genes were related to lipid metabolism, including MOGAT1, LPGAT1, AGPAT2, and DGAT1 (glycerolipid metabolism); CDS1, PCTP, and MAT1A (phospholipid metabolism); and FATP2, SLC6A12, and AQP3 (uptake of fatty acids, betaine, and glycerol, respectively). They were rapidly induced (4-6 h) upon VDR activation by 10 nM VitD or 100 µM lithocholic acid (LCA). Most of these genes were also upregulated by VDR/VitD in mouse livers in vivo. Ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS) metabolomics demonstrated intracellular accumulation of triglycerides, with concomitant decreases in diglycerides and phosphatidates, at 8 and 24 h upon VDR activation. Significant alterations in phosphatidylcholines, increases in lyso-phosphatidylcholines and decreases in phosphatidylethanolamines and phosphatidylethanolamine plasmalogens were also observed. In conclusion, active VitD/VDR signaling in hepatocytes triggers an unanticipated coordinated gene response leading to triglyceride synthesis and to important perturbations in glycerolipids and phospholipids.
    Keywords:  human hepatocytes; lipid metabolism; vitamin D; vitamin D receptor
    DOI:  https://doi.org/10.3390/biom10030493
  13. Methods Mol Biol. 2020 ;2138 49-81
    Poljak A, Duncan MW, Jayasena T, Sachdev PS.
      The apolipoproteins are well known for their roles in both health and disease, as components of plasma lipoprotein particles, such as high-density lipoprotein (HDL), low-density lipoprotein (LDL), very-low-density lipoprotein (VLDL), chylomicrons, and metabolic, vascular- and inflammation-related disorders, such as cardiovascular disease, atherosclerosis, metabolic syndrome, and diabetes. Increasingly, their roles in neurovascular and neurodegenerative disorders are also being elucidated. They play major roles in lipid and cholesterol transport between blood and organs and are, therefore, critical to maintenance and homeostasis of the lipidome, with apolipoprotein-lipid interactions, including cholesterol, fatty acids, triglycerides, phospholipids, and isoprostanes. Further, they have important pleiotropic roles related to aging and longevity, which are largely managed through their many structural variants, including multiple isoforms, and a diversity of post-translational modifications. Consequently, tools for the characterization and accurate quantification of apolipoproteins, including their diverse array of variant forms, are required to understand their salutary and disease related roles. In this chapter we outline three distinct quantitative approaches suitable for targeting apolipoproteins: (1) multiplex immunoassays, (2) mass spectrometric immunoassay, and (3) multiple reaction monitoring, mass spectrometric quantification. We also discuss management of pre-analytical and experimental design variables.
    Keywords:  Apolipoproteins; Atherosclerosis; Cardiovascular disease; Diabetes; Mass spectrometric immunoassay; Metabolic syndrome; Multiple reaction monitoring; Multiplex immunoassay
    DOI:  https://doi.org/10.1007/978-1-0716-0471-7_3
  14. Annu Rev Anal Chem (Palo Alto Calif). 2020 Mar 25.
    Sämfors S, Fletcher JS.
      Lipids are an important class of biomolecules with many roles within cells and tissue. As targets for study, they present several challenges. They are difficult to label, as many labels lack the specificity to the many different lipid species or the labels maybe larger than the lipids themselves, thus severely perturbing the natural chemical environment. Mass spectrometry provides exceptional specificity and is often used to examine lipid extracts from different samples. However, spatial information is lost during extraction. Of the different imaging mass spectrometry methods available, secondary ion mass spectrometry (SIMS) is unique in its ability to analyze very small features, with probe sizes <50 nm available. It also offers high surface sensitivity and 3D imaging capability on a subcellular scale. This article reviews the current capabilities and some remaining challenges associated with imaging the diverse lipids present in cell and tissue samples. We show how the technique has moved beyond show-and-tell, proof-of-principle analysis and is now being used to address real biological challenges. These include imaging the microenvironment of cancer tumors, probing the pathophysiology of traumatic brain injury, or tracking the lipid composition through bacterial membranes. Expected final online publication date for the Annual Review of Analytical Chemistry, Volume 13 is June 12, 2020. Please see http://www.annualreviews.org/page/journal/pubdates for revised estimates.
    DOI:  https://doi.org/10.1146/annurev-anchem-091619-103512
  15. Nature. 2020 Mar;579(7800): 586-591
    Zhao S, Jang C, Liu J, Uehara K, Gilbert M, Izzo L, Zeng X, Trefely S, Fernandez S, Carrer A, Miller KD, Schug ZT, Snyder NW, Gade TP, Titchenell PM, Rabinowitz JD, Wellen KE.
      Consumption of fructose has risen markedly in recent decades owing to the use of sucrose and high-fructose corn syrup in beverages and processed foods1, and this has contributed to increasing rates of obesity and non-alcoholic fatty liver disease2-4. Fructose intake triggers de novo lipogenesis in the liver4-6, in which carbon precursors of acetyl-CoA are converted into fatty acids. The ATP citrate lyase (ACLY) enzyme cleaves cytosolic citrate to generate acetyl-CoA, and is upregulated after consumption of carbohydrates7. Clinical trials are currently pursuing the inhibition of ACLY as a treatment for metabolic diseases8. However, the route from dietary fructose to hepatic acetyl-CoA and lipids remains unknown. Here, using in vivo isotope tracing, we show that liver-specific deletion of Acly in mice is unable to suppress fructose-induced lipogenesis. Dietary fructose is converted to acetate by the gut microbiota9, and this supplies lipogenic acetyl-CoA independently of ACLY10. Depletion of the microbiota or silencing of hepatic ACSS2, which generates acetyl-CoA from acetate, potently suppresses the conversion of bolus fructose into hepatic acetyl-CoA and fatty acids. When fructose is consumed more gradually to facilitate its absorption in the small intestine, both citrate cleavage in hepatocytes and microorganism-derived acetate contribute to lipogenesis. By contrast, the lipogenic transcriptional program is activated in response to fructose in a manner that is independent of acetyl-CoA metabolism. These data reveal a two-pronged mechanism that regulates hepatic lipogenesis, in which fructolysis within hepatocytes provides a signal to promote the expression of lipogenic genes, and the generation of microbial acetate feeds lipogenic pools of acetyl-CoA.
    DOI:  https://doi.org/10.1038/s41586-020-2101-7
  16. Metabolism. 2020 Mar 21. pii: S0026-0495(20)30079-2. [Epub ahead of print] 154215
    Liu J, Muturi HT, Khuder SS, Helal RA, Ghadieh HE, Ramakrishnan SK, Kaw MK, Lester SG, Al-Khudhair A, Conran PB, Chin KV, Gatto-Weis C, Najjar SM.
      OBJECTIVE: PTEN haploinsufficiency plays an important role in prostate cancer development in men. However, monoallelic deletion of Pten gene failed to induce high prostate intraepithelial neoplasia (PIN) until Pten+/- mice aged or fed a high-calorie diet. Because CEACAM1, a cell adhesion molecule with a potential tumor suppression activity, is induced in Pten+/- prostates, the study aimed at examining whether the rise of Ceacam1 limited neoplastic progression in Pten+/- prostates.METHODS: Pten+/- were crossbred with Cc1-/- mice harboring a null deletion of Ceacam1 gene to produce Pten+/-/Cc1-/- double mutants. Prostates from 7-month old male mice were analyzed histologically and biochemically for PIN progression.
    RESULTS: Deleting Ceacam1 in Pten+/- mice caused an early development of high-grade PIN in parallel to hyperactivation of PI3 kinase/Akt and Ras/MAP kinase pathways, with an increase in cell proliferation, epithelial-to-mesenchymal transition, angiogenesis and inflammation relative to Pten+/- and Cc1-/- individual mutants. It also caused a remarkable increase in lipogenesis in prostate despite maintaining insulin sensitivity. Concomitant Ceacam1 deletion with Pten+/- activated the IL-6/STAT3 signaling pathways to suppress Irf-8 transcription that in turn, led to a decrease in the expression level of promyelocytic leukemia gene, a well characterized tumor suppressor in prostate.
    CONCLUSIONS: Ceacam1 deletion accelerated high-grade prostate intraepithelial neoplasia in Pten haploinsufficient mice while preserving insulin sensitivity. This demonstrated that the combined loss of Ceacam1 and Pten advanced prostate cancer by increasing lipogenesis and modifying the STAT3-dependent inflammatory microenvironment of prostate.
    Keywords:  CEACAM1; Fatty acid synthase; Neoplasia; Normoinsulinemia; PML; PTEN tumor suppression; Prostate intraepithelial neoplasia
    DOI:  https://doi.org/10.1016/j.metabol.2020.154215
  17. Anal Chem. 2020 Mar 26.
    McLean C, Kujawinski EB.
      Untargeted metabolomics experiments provide a snapshot of cellular metabolism, but remain challenging to interpret due to the computational complexity involved in data processing and analysis. Prior to any interpretation, raw data must be processed to remove noise and to align mass-spectral peaks across samples. This step requires selection of dataset-specific parameters, as erroneous parameters can result in noise inflation. While several algorithms exist to automate parameter selection, each depends on gradient descent optimization functions. In contrast, our new parameter optimization algorithm, AutoTuner, obtains parameter estimates from raw data in a single step as opposed to many iterations. Here, we tested the accuracy and the run time of AutoTuner in comparison to isotopologue parameter optimization (IPO), the most commonly-used parameter selection tool, and compared the resulting parameters' influence on the quality of feature tables after processing. We performed a Monte Carlo experiment to test the robustness of AutoTuner parameter selection, and found that AutoTuner generated similar parameter estimates from random subsets of samples. We conclude that AutoTuner is a desirable alternative to existing tools, because it is scalable, highly robust, and very fast (~100-1000X speed improvement from other algorithms going from days to minutes). AutoTuner is freely available as an R package through BioConductor.
    DOI:  https://doi.org/10.1021/acs.analchem.9b04804
  18. Biochem J. 2020 Mar 27. pii: BCJ20200099. [Epub ahead of print]
    Sikder MOF, Sivaprakasam S, Brown TP, Thangaraju M, Bhutia YD, Ganapathy V.
      SLC6A14 is a Na+/Cl--coupled transporter for neutral and cationic amino acids. It is expressed at basal levels in normal colon but is upregulated in colon cancer. However, the relevance of this upregulation to cancer progression and the mechanisms involved in the upregulation remain unknown. Here we show that SLC6A14 is essential for colon cancer and that its upregulation involves, at least partly, Wnt signaling. Upregulation of the transporter is evident in most human colon cancer cell lines and also in a majority of patient-derived xenografts. These findings are supported by publicly available TCGA (The Cancer Genome Atlas) database. Treatment of colon cancer cells with α-methyltryptophan (α-MT), a blocker of SLC6A14, induces amino acid deprivation, decreases mTOR activity, increases autophagy, promotes apoptosis, and suppresses cell proliferation and invasion. In xenograft and syngeneic mouse tumor models, silencing of SLC6A14 by shRNA or blocking its function by α-MT reduces tumor growth. Similarly, deletion of Slc6a14in mice protects against colon cancer in two different experimental models (inflammation-associated colon cancer and genetically driven colon cancer). In colon cancer cells, expression of the transporter is reduced by Wnt antagonist or by silencing of b-catenin whereas Wnt agonist or overexpression of b-catenin shows the opposite effect. Finally, SLC6A14 as a target for b-catenin is confirmed by chromatin immunoprecipitation. These studies demonstrate that SLC6A14 plays a critical role in promotion of colon cancer and that its upregulation in cancer involves Wnt signaling. These findings identify SLC6A14 as a promising drug target for treatment of colon cancer.
    Keywords:  Slc6a14-null mouse; Wnt signaling; colon cancer; mTOR; α-methyltryptophan; β-catenin
    DOI:  https://doi.org/10.1042/BCJ20200099
  19. Rapid Commun Mass Spectrom. 2020 Mar 24. e8792
    Radchenko T, Kochansky CJ, Cancilla M, Wrona MD, Mortishire-Smith RJ, Kirk J, Murray G, Fontaine F, Zamora I.
      RATIONALE: Liquid chromatography/mass spectrometry is an essential tool for efficient and reliable quantitative and qualitative analysis and underpins much of contemporary drug metabolism and pharmacokinetics. Data-independent acquisition methods such as MSE have reduced potential to miss metabolites, but do not formally generate quadrupole-resolved product ion spectra. The addition of ion mobility separation to these approaches, for example, in High Definition MSE (HDMSE ) has the potential to reduce the time needed to set up an experiment and maximize the chance all metabolites present can be resolved and characterized. We compared High Definition Data-Dependent Acquisition (HD-DDA), MSE and HDMSE approaches using automated software processing with MassMetaSite and WebMetabase.METHODS: Metabolite identification was performed on incubations of glucagon-like peptide-1 (7-37) (GLP-1) and verapamil hydrochloride. The HD-DDA, MSE and HDMSE experiments were conducted on a Waters ACQUITY UPLC I-Class LC system with VION IMS QTof mass spectrometer operating under UNIFI control. All acquired data were processed using MassMetaSite able to read data from UNIFI 1.9.4. WebMetabase was used to review the detected chromatographic peaks and the spectral data interpretations.
    RESULTS: A comparison of outcomes obtained for MSE and HDMSE data demonstrated that the same structures were proposed for metabolites of both verapamil and GLP-1. The ratio of structurally matched to mismatched product ions found by MassMetaSite was slightly greater for HDMSE than for MSE , and HD-DDA, thus improving confidence in the structures proposed through the addition of ion mobility-based data acquisitions. CONCLUSIONS: HDMSE data acquisition is an effective approach for the elucidation of metabolite structures for both small molecules and peptides, with excellent accuracy and quality, requiring minimal tailoring for the compound under investigation.
    DOI:  https://doi.org/10.1002/rcm.8792
  20. Cell Metab. 2020 Mar 16. pii: S1550-4131(20)30119-4. [Epub ahead of print]
    Kurniawan H, Franchina DG, Guerra L, Bonetti L, -Baguet LS, Grusdat M, Schlicker L, Hunewald O, Dostert C, Merz MP, Binsfeld C, Duncan GS, Farinelle S, Nonnenmacher Y, Haight J, Das Gupta D, Ewen A, Taskesen R, Halder R, Chen Y, Jäger C, Ollert M, Wilmes P, Vasiliou V, Harris IS, Knobbe-Thomsen CB, Turner JD, Mak TW, Lohoff M, Meiser J, Hiller K, Brenner D.
      Regulatory T cells (Tregs) maintain immune homeostasis and prevent autoimmunity. Serine stimulates glutathione (GSH) synthesis and feeds into the one-carbon metabolic network (1CMet) essential for effector T cell (Teff) responses. However, serine's functions, linkage to GSH, and role in stress responses in Tregs are unknown. Here, we show, using mice with Treg-specific ablation of the catalytic subunit of glutamate cysteine ligase (Gclc), that GSH loss in Tregs alters serine import and synthesis and that the integrity of this feedback loop is critical for Treg suppressive capacity. Although Gclc ablation does not impair Treg differentiation, mutant mice exhibit severe autoimmunity and enhanced anti-tumor responses. Gclc-deficient Tregs show increased serine metabolism, mTOR activation, and proliferation but downregulated FoxP3. Limitation of cellular serine in vitro and in vivo restores FoxP3 expression and suppressive capacity of Gclc-deficient Tregs. Our work reveals an unexpected role for GSH in restricting serine availability to preserve Treg functionality.
    Keywords:  FoxP3; ROS; Treg; autoimmunity; cancer; diet; glutamate cysteine ligase; glutathione; one carbon metabolism; serine metabolism
    DOI:  https://doi.org/10.1016/j.cmet.2020.03.004