The feel of inclusivity

Collaboration between diverse groups with different perspectives promotes scientific discovery. To enhance these partnerships and facilitate communication, innovative advances in universal design are essential

MYC sensitises cells to apoptosis by driving energetic demand

The MYC oncogene is a potent driver of growth and proliferation but also sensitises cells to apoptosis, which limits its oncogenic potential. MYC induces several biosynthetic programmes and primary cells overexpressing MYC are highly sensitive to glutamine withdrawal suggesting that MYC-induced sensitisation to apoptosis may be due to imbalance of metabolic/energetic supply and demand. Here we show that MYC elevates global transcription and translation, even in the absence of glutamine, revealing metabolic demand without corresponding supply. Glutamine withdrawal from MRC-5 fibroblasts depletes key tricarboxylic acid (TCA) cycle metabolites and, in combination with MYC activation, leads to AMP accumulation and nucleotide catabolism indicative of energetic stress. Further analyses reveal that glutamine supports viability through TCA cycle energetics rather than asparagine biosynthesis and that TCA cycle inhibition confers tumour suppression on MYC-driven lymphoma in vivo. In summary, glutamine supports the viability of MYC-overexpressing cells through an energetic rather than a biosynthetic mechanism

Bar coding MS2 spectra for metabolite identification

[Image: see text] Metabolite identifications are most frequently achieved in untargeted metabolomics by matching precursor mass and full, high-resolution MS(2) spectra to metabolite databases and standards. Here we considered an alternative approach for establishing metabolite identifications that does not rely on full, high-resolution MS(2) spectra. First, we select mass-to-charge regions containing the most informative metabolite fragments and designate them as bins. We then translate each metabolite fragmentation pattern into a binary code by assigning 1’s to bins containing fragments and 0’s to bins without fragments. With 20 bins, this binary-code system is capable of distinguishing 96% of the compounds in the METLIN MS(2) library. A major advantage of the approach is that it extends untargeted metabolomics to low-resolution triple quadrupole (QqQ) instruments, which are typically less expensive and more robust than other types of mass spectrometers. We demonstrate a method of acquiring MS(2) data in which the third quadrupole of a QqQ instrument cycles over 20 wide isolation windows (coinciding with the location and width of our bins) for each precursor mass selected by the first quadrupole. Operating the QqQ instrument in this mode yields diagnostic bar codes for each precursor mass that can be matched to the bar codes of metabolite standards. Furthermore, our data suggest that using low-resolution bar codes enables QqQ instruments to make MS(2)-based identifications in untargeted metabolomics with a specificity and sensitivity that is competitive to high-resolution time-of-flight technologies

Arteriovenous Blood Metabolomics: A Readout of Intra-Tissue Metabostasis.

The human circulatory system consists of arterial blood that delivers nutrients to tissues, and venous blood that removes the metabolic by-products. Although it is well established that arterial blood generally has higher concentrations of glucose and oxygen relative to venous blood, a comprehensive biochemical characterization of arteriovenous differences has not yet been reported. Here we apply cutting-edge, mass spectrometry-based metabolomic technologies to provide a global characterization of metabolites that vary in concentration between the arterial and venous blood of human patients. Global profiling of paired arterial and venous plasma from 20 healthy individuals, followed up by targeted analysis made it possible to measure subtle (<2 fold), yet highly statistically significant and physiologically important differences in water soluble human plasma metabolome. While we detected changes in lactic acid, alanine, glutamine, and glutamate as expected from skeletal muscle activity, a number of unanticipated metabolites were also determined to be significantly altered including Krebs cycle intermediates, amino acids that have not been previously implicated in transport, and a few oxidized fatty acids. This study provides the most comprehensive assessment of metabolic changes in the blood during circulation to date and suggests that such profiling approach may offer new insights into organ homeostasis and organ specific pathology

Mitochondrial pyruvate metabolism and glutaminolysis toggle steady-state and emergency myelopoiesis

To define the metabolic requirements of hematopoiesis, we examined blood lineages in mice conditionally deficient in genes required for long-chain fatty acid oxidation (Cpt2), glutaminolysis (Gls), or mitochondrial pyruvate import (Mpc2). Genetic ablation of Cpt2 or Gls minimally impacted most blood lineages. In contrast, deletion of Mpc2 led to a sharp decline in mature myeloid cells and a slower reduction in T cells, whereas other hematopoietic lineages were unaffected. Yet MPC2-deficient monocytes and neutrophils rapidly recovered due to a transient and specific increase in myeloid progenitor proliferation. Competitive bone marrow chimera and stable isotope tracing experiments demonstrated that this proliferative burst was progenitor intrinsic and accompanied by a metabolic switch to glutaminolysis. Myeloid recovery after loss of MPC2 or cyclophosphamide treatment was delayed in the absence of GLS. Reciprocally, MPC2 was not required for myeloid recovery after cyclophosphamide treatment. Thus, mitochondrial pyruvate metabolism maintains myelopoiesis under steady-state conditions, while glutaminolysis in progenitors promotes emergency myelopoiesis

Evidence that 2-hydroxyglutarate is not readily metabolized in colorectal carcinoma cells

BACKGROUND: Two-hydroxyglutarate (2HG) is present at low concentrations in healthy mammalian cells as both an L and D enantiomer. Both the L and D enantiomers have been implicated in regulating cellular physiology by mechanisms that are only partially characterized. In multiple human cancers, the D enantiomer accumulates due to gain-of-function mutations in the enzyme isocitrate dehydrogenase (IDH) and has been hypothesized to drive malignancy through mechanisms that remain incompletely understood. While much attention has been dedicated to identifying the route of 2HG synthesis, the metabolic fate of 2HG has not been studied in detail. Yet the metabolism of 2HG may have important mechanistic consequences influencing cell function and cancer pathogenesis, such as modulating redox potential or producing unknown products with unique modes of action. RESULTS: By applying our isotope-based metabolomic platform, we unbiasedly and comprehensively screened for products of L- and D-2HG in HCT116 colorectal carcinoma cells harboring a mutation in IDH1. After incubating HCT116 cells in uniformly (13)C-labeled 2HG for 24 h, we used liquid chromatography/mass spectrometry to track the labeled carbons in small molecules. Strikingly, we did not identify any products of 2HG metabolism from the thousands of metabolomic features that we screened. Consistent with these results, we did not detect any significant changes in the labeling patterns of tricarboxylic acid cycle metabolites from wild type or IDH1 mutant cells cultured in (13)C-labeled glucose upon the addition of L, D, or racemic mixtures of 2HG. A more sensitive, targeted analysis revealed trace levels of isotopic enrichment (<1 %) in some central carbon metabolites from (13)C-labeled 2HG. However, we found that cells do not deplete 2HG from the media at levels above our detection limit over a 48 h time period. CONCLUSIONS: Taken together, we conclude that 2HG carbon is not readily transformed in the HCT116 cell line. These data indicate that the phenotypic alterations induced by 2HG are not a result of its metabolic products

Uridine-derived ribose fuels glucose-restricted pancreatic cancer

Pancreatic ductal adenocarcinoma (PDA) is a lethal disease notoriously resistant to therap

Deletion of Glut1 in early postnatal cartilage reprograms chondrocytes toward enhanced glutamine oxidation

Abstract Glucose metabolism is fundamental for the functions of all tissues, including cartilage. Despite the emerging evidence related to glucose metabolism in the regulation of prenatal cartilage development, little is known about the role of glucose metabolism and its biochemical basis in postnatal cartilage growth and homeostasis. We show here that genetic deletion of the glucose transporter Glut1 in postnatal cartilage impairs cell proliferation and matrix production in growth plate (GPs) but paradoxically increases cartilage remnants in the metaphysis, resulting in shortening of long bones. On the other hand, articular cartilage (AC) with Glut1 deficiency presents diminished cellularity and loss of proteoglycans, which ultimately progress to cartilage fibrosis. Moreover, predisposition to Glut1 deficiency severely exacerbates injury-induced osteoarthritis. Regardless of the disparities in glucose metabolism between GP and AC chondrocytes under normal conditions, both types of chondrocytes demonstrate metabolic plasticity to enhance glutamine utilization and oxidation in the absence of glucose availability. However, uncontrolled glutamine flux causes collagen overmodification, thus affecting extracellular matrix remodeling in both cartilage compartments. These results uncover the pivotal and distinct roles of Glut1-mediated glucose metabolism in two of the postnatal cartilage compartments and link some cartilage abnormalities to altered glucose/glutamine metabolism

A multidimensional metabolomics workflow to image biodistribution and evaluate pharmacodynamics in adult zebrafish

An integrated evaluation of the tissue distribution and pharmacodynamic properties of a therapeutic is essential for successful translation to the clinic. To date, however, cost-effective methods to measure these parameters at the systems level in model organisms are lacking. Here, we introduce a multidimensional workflow to evaluate drug activity that combines mass spectrometry-based imaging, absolute drug quantitation across different biological matrices, in vivo isotope tracing and global metabolome analysis in the adult zebrafish. As a proof of concept, we quantitatively determined the whole-body distribution of the anti-rheumatic agent hydroxychloroquine sulfate (HCQ) and measured the systemic metabolic impacts of drug treatment. We found that HCQ distributed to most organs in the adult zebrafish 24 h after addition of the drug to water, with the highest accumulation of both the drug and its metabolites being in the liver, intestine and kidney. Interestingly, HCQ treatment induced organ-specific alterations in metabolism. In the brain, for example, HCQ uniquely elevated pyruvate carboxylase activity to support increased synthesis of the neuronal metabolite, N-acetylaspartate. Taken together, this work validates a multidimensional metabolomics platform for evaluating the mode of action of a drug and its potential off-target effects in the adult zebrafish. This article has an associated First Person interview with the first author of the paper