bims-mepmim 2023-07-30 papers

bims-mepmim

Biomed News

on Metabolites in pathological microenvironments and immunometabolism

Issue of 2023‒07‒30
34 papers selected by
Erika Mariana Palmieri, NIH/NCI Laboratory of Cancer ImmunoMetabolism

Macrophage-to-endothelial cell crosstalk by the cholesterol metabolite 27HC promotes atherosclerosis in male mice.
The Virulence Factor Macrophage Infectivity Potentiator (Mip) Influences Branched-Chain Amino Acid Metabolism and Pathogenicity of Legionella pneumophila.
Lipoylation is dependent on the ferredoxin FDX1 and dispensable under hypoxia in human cells.
The lncRNA HOXA11os regulates mitochondrial function in myeloid cells to maintain intestinal homeostasis.
Branched-chain amino acid synthesis is coupled to TOR activation early in the cell cycle in yeast.
Moderate hypoxia induces metabolic divergence in circulating monocytes and tissue resident macrophages from Berkeley sickle cell anemia mice.
Cutaneous squamous cell carcinoma-derived extracellular vesicles exert an oncogenic role by activating cancer-associated fibroblasts.
Cell-autonomous regulation of complement C3 by factor H limits macrophage efferocytosis and exacerbates atherosclerosis.
Metabolite-sensing GPCRs controlling interactions between adipose tissue and inflammation.
Melanocortin 1 receptor regulates cholesterol and bile acid metabolism in the liver.
The vitamin B5/coenzyme A axis: A target for immunomodulation?
Resolvin D1 prevents injurious neutrophil swarming in transplanted lungs.
Identification of structural determinants of nicotinamide phosphoribosyl transferase (NAMPT) activity and substrate selectivity.
Lipid Signatures and Inter-Cellular Heterogeneity of Naı̈ve and Lipopolysaccharide-Stimulated Human Microglia-like Cells.
Loss of metabolic fitness drives tumor resistance after CAR-NK cell therapy and can be overcome by cytokine engineering.
Arginine dependency is a therapeutically exploitable vulnerability in chronic myeloid leukaemic stem cells.
Interleukin-17 as a key player in neuroimmunometabolism.
A GSTP1-mediated lactic acid signaling promotes tumorigenesis through the PPP oxidative branch.
Stearate-derived very long-chain fatty acids are indispensable to tumor growth.
Nitric oxide-induced ribosome collision activates ribosomal surveillance mechanisms.
Tumor aerobic glycolysis confers immune evasion through modulating sensitivity to T cell-mediated bystander killing via TNF-α.
Invariant natural killer T cells drive hepatic homeostasis in Non-alcoholic fatty liver disease via sustained IL-10 expression in CD170+ Kupffer cells.
Tumor interstitial fluid analysis enables the study of microenvironment-cell interactions in cancers.
Alcohol-sourced acetate impairs T cell function by promoting cortactin acetylation.
Acidosis attenuates CPT-I supported bioenergetics as a potential mechanism limiting lipid oxidation.
A transient increase of HIF-1α during the G1 phase (G1-HIF) ensures cell survival under nutritional stress.
Nascent mitochondrial proteins initiate the localized condensation of cytosolic protein aggregates on the mitochondrial surface.
Supersulphides provide airway protection in viral and chronic lung diseases.
Combined GLUT1 and OXPHOS inhibition eliminates acute myeloid leukemia cells by restraining their metabolic plasticity.
Phosphatidylinositol 3-phosphate regulates iron transport via PI3P-binding CgPil1 protein.
Platelet-derived lipids promote insulin secretion of pancreatic β cells.
LRRK2 suppresses lysosome degradative activity in macrophages and microglia through MiT-TFE transcription factor inhibition.
PGAM5 is an MFN2 phosphatase that plays an essential role in the regulation of mitochondrial dynamics.
Competition for cysteine acylation by C16:0 and C18:0 derived lipids is a global phenomenon in the proteome.

Nat Commun. 2023 07 25. 14(1): 4101

Macrophage-to-endothelial cell crosstalk by the cholesterol metabolite 27HC promotes atherosclerosis in male mice.

Liming Yu, Lin Xu, Haiyan Chu, Jun Peng, Anastasia Sacharidou, Hsi-Hsien Hsieh, Ada Weinstock, Sohaib Khan, Liqian Ma, José Gabriel Barcia Durán, Jeffrey McDonald, Erik R Nelson, Sunghee Park, Donald P McDonnell, Kathryn J Moore, Lily Jun-Shen Huang, Edward A Fisher, Chieko Mineo, Linzhang Huang, Philip W Shaul.

Hypercholesterolemia and vascular inflammation are key interconnected contributors to the pathogenesis of atherosclerosis. How hypercholesterolemia initiates vascular inflammation is poorly understood. Here we show in male mice that hypercholesterolemia-driven endothelial activation, monocyte recruitment and atherosclerotic lesion formation are promoted by a crosstalk between macrophages and endothelial cells mediated by the cholesterol metabolite 27-hydroxycholesterol (27HC). The pro-atherogenic actions of macrophage-derived 27HC require endothelial estrogen receptor alpha (ERα) and disassociation of the cytoplasmic scaffolding protein septin 11 from ERα, leading to extranuclear ERα- and septin 11-dependent activation of NF-κB. Furthermore, pharmacologic inhibition of cyp27a1, which generates 27HC, affords atheroprotection by reducing endothelial activation and monocyte recruitment. These findings demonstrate cell-to-cell communication by 27HC, and identify a major causal linkage between the hypercholesterolemia and vascular inflammation that partner to promote atherosclerosis. Interventions interrupting this linkage may provide the means to blunt vascular inflammation without impairing host defense to combat the risk of atherosclerotic cardiovascular disease that remains despite lipid-lowering therapies.

DOI: https://doi.org/10.1038/s41467-023-39586-z
Metabolites. 2023 Jul 11. pii: 834. [Epub ahead of print]13(7):

The Virulence Factor Macrophage Infectivity Potentiator (Mip) Influences Branched-Chain Amino Acid Metabolism and Pathogenicity of Legionella pneumophila.

Fabian Nikolka, Mustafa Safa Karagöz, Mohamed Zakaria Nassef, Karsten Hiller, Michael Steinert, Thekla Cordes.

  Legionella pneumophila (Lp) is a common etiological agent of bacterial pneumonia that causes Legionnaires' disease (LD). The bacterial membrane-associated virulence factor macrophage infectivity potentiator (Mip) exhibits peptidyl-prolyl-cis/trans-isomerase (PPIase) activity and contributes to the intra- and extracellular pathogenicity of Lp. Though Mip influences disease outcome, little is known about the metabolic consequences of altered Mip activity during infections. Here, we established a metabolic workflow and applied mass spectrometry approaches to decipher how Mip activity influences metabolism and pathogenicity. Impaired Mip activity in genetically engineered Lp strains decreases intracellular replication in cellular infection assays, confirming the contribution of Mip for Lp pathogenicity. We observed that genetic and chemical alteration of Mip using the PPIase inhibitors rapamycin and FK506 induces metabolic reprogramming in Lp, specifically branched-chain amino acid (BCAA) metabolism. Rapamycin also inhibits PPIase activity of mammalian FK506 binding proteins, and we observed that rapamycin induces a distinct metabolic signature in human macrophages compared to bacteria, suggesting potential involvement of Mip in normal bacteria and in infection. Our metabolic studies link Mip to alterations in BCAA metabolism and may help to decipher novel disease mechanisms associated with LD.

Keywords:  Legionella pneumophila; branched-chain amino acid; infection model; macrophage; macrophage infectivity potentiator (Mip); mass spectrometry; metabolism; tracing; virulence factor

DOI:  https://doi.org/10.3390/metabo13070834
J Biol Chem. 2023 Jul 20. pii: S0021-9258(23)02103-8. [Epub ahead of print] 105075

Lipoylation is dependent on the ferredoxin FDX1 and dispensable under hypoxia in human cells.

Pallavi R Joshi, Shayan Sadre, Xiaoyan A Guo, Jason G McCoy, Vamsi K Mootha.

  Iron sulfur clusters (ISC) are essential cofactors that participate in electron transfer, environmental sensing, and catalysis. Amongst the most ancient ISC containing proteins are the ferredoxin family of electron carriers. Humans have two ferredoxins, FDX1 and FDX2, both of which are localized to mitochondria, and the latter of which is itself important for ISC synthesis. We have previously shown that hypoxia can eliminate the requirement for some components of the ISC biosynthetic pathway, but ferredoxins were not included in that study. Here we report that FDX1, but not FDX2, is dispensable under 1% O2 in cultured human cells. We find that FDX1 is essential for production of the lipoic acid cofactor, which is synthesized by the ISC containing enzyme lipoyl synthase (LIAS). While hypoxia can rescue the growth phenotype of either FDX1 or LIAS knockout cells, lipoylation in these same cells is not rescued, arguing against an alternative biosynthetic route or salvage pathway for lipoate in hypoxia. Our work reveals the divergent roles of FDX1 and FDX2 in mitochondria, identifies a role for FDX1 in lipoate synthesis, and suggests that loss of lipoic acid can be tolerated under low oxygen tensions in cell culture.

Keywords:  Energy Metabolism; Hypoxia; Iron-Sulfur Protein; Lipoate; Mitochondria; S-adenosyl methionine

DOI:  https://doi.org/10.1016/j.jbc.2023.105075
Cell Metab. 2023 Jul 20. pii: S1550-4131(23)00250-4. [Epub ahead of print]

The lncRNA HOXA11os regulates mitochondrial function in myeloid cells to maintain intestinal homeostasis.

Liraz Shmuel-Galia, Fiachra Humphries, Tim Vierbuchen, Zhaozhao Jiang, Nolan Santos, John Johnson, Boris Shklyar, Leonel Joannas, Nicholas Mustone, Shany Sherman, Doyle Ward, JeanMarie Houghton, Christina E Baer, Aisling O'Hara, Jorge Henao-Mejia, Kasper Hoebe, Katherine A Fitzgerald.

  This study reveals a previously uncharacterized mechanism to restrict intestinal inflammation via a regulatory RNA transcribed from a noncoding genomic locus. We identified a novel transcript of the lncRNA HOXA11os specifically expressed in the distal colon that is reduced to undetectable levels in colitis. HOXA11os is localized to mitochondria under basal conditions and interacts with a core subunit of complex 1 of the electron transport chain (ETC) to maintain its activity. Deficiency of HOXA11os in colonic myeloid cells results in complex I deficiency, dysfunctional oxidative phosphorylation (OXPHOS), and the production of mitochondrial reactive oxygen species (mtROS). As a result, HOXA11os-deficient mice develop spontaneous intestinal inflammation and are hypersusceptible to colitis. Collectively, these studies identify a new regulatory axis whereby a lncRNA maintains intestinal homeostasis and restricts inflammation in the colon through the regulation of complex I activity.

Keywords:  IBD; Krebs cycle; OXPHOS; colitis; complex I; intestinal inflammation; lncRNA; mitochondria; mtROS; mucosal inflammation; ncRNA

DOI:  https://doi.org/10.1016/j.cmet.2023.06.019
EMBO Rep. 2023 Jul 27. e57372

Branched-chain amino acid synthesis is coupled to TOR activation early in the cell cycle in yeast.

Heidi M Blank, Carsten Reuse, Kerstin Schmidt-Hohagen, Staci E Hammer, Karsten Hiller, Michael Polymenis.

  How cells coordinate their metabolism with division determines the rate of cell proliferation. Dynamic patterns of metabolite synthesis during the cell cycle are unexplored. We report the first isotope tracing analysis in synchronous, growing budding yeast cells. Synthesis of leucine, a branched-chain amino acid (BCAA), increases through the G1 phase of the cell cycle, peaking later during DNA replication. Cells lacking Bat1, a mitochondrial aminotransferase that synthesizes BCAAs, grow slower, are smaller, and are delayed in the G1 phase, phenocopying cells in which the growth-promoting kinase complex TORC1 is moderately inhibited. Loss of Bat1 lowers the levels of BCAAs and reduces TORC1 activity. Exogenous provision of valine and, to a lesser extent, leucine to cells lacking Bat1 promotes cell division. Valine addition also increases TORC1 activity. In wild-type cells, TORC1 activity is dynamic in the cell cycle, starting low in early G1 but increasing later in the cell cycle. These results suggest a link between BCAA synthesis from glucose to TORC1 activation in the G1 phase of the cell cycle.

Keywords:  BCAA; BCAT; TORC1; cell size; isotope tracing

DOI:  https://doi.org/10.15252/embr.202357372
Front Med (Lausanne). 2023 ;10 1149005

Moderate hypoxia induces metabolic divergence in circulating monocytes and tissue resident macrophages from Berkeley sickle cell anemia mice.

Christina Lisk, Francesca Cendali, David I Pak, Delaney Swindle, Kathryn Hassell, Rachelle Nuss, Gemlyn George, Pavel Davizon-Castillo, Paul W Buehler, Angelo D'Alessandro, David C Irwin.

  Introduction: Human and murine sickle cell disease (SCD) associated pulmonary hypertension (PH) is defined by hemolysis, nitric oxide depletion, inflammation, and thrombosis. Further, hemoglobin (Hb), heme, and iron accumulation are consistently observed in pulmonary adventitial macrophages at autopsy and in hypoxia driven rodent models of SCD, which show distribution of ferric and ferrous Hb as well as HO-1 and ferritin heavy chain. The anatomic localization of these macrophages is consistent with areas of significant vascular remodeling. However, their contributions toward progressive disease may include unique, but also common mechanisms, that overlap with idiopathic and other forms of pulmonary hypertension. These processes likely extend to the vasculature of other organs that are consistently impaired in advanced SCD.Methods: To date, limited information is available on the metabolism of macrophages or monocytes isolated from lung, spleen, and peripheral blood in humans or murine models of SCD.
Results: Here we hypothesize that metabolism of macrophages and monocytes isolated from this triad of tissue differs between Berkley SCD mice exposed for ten weeks to moderate hypobaric hypoxia (simulated 8,000 ft, 15.4% O2) or normoxia (Denver altitude, 5000 ft) with normoxia exposed wild type mice evaluated as controls.
Discussion: This study represents an initial set of data that describes the metabolism in monocytes and macrophages isolated from moderately hypoxic SCD mice peripheral lung, spleen, and blood mononuclear cells.

Keywords:  hypoxia; metabolic disease; pulmonary hypertension; sickle cell disease; spleen

DOI:  https://doi.org/10.3389/fmed.2023.1149005
Cell Death Discov. 2023 Jul 26. 9(1): 260

Cutaneous squamous cell carcinoma-derived extracellular vesicles exert an oncogenic role by activating cancer-associated fibroblasts.

Chen Li, Chengxi Sun, Warangkana Lohcharoenkal, Mohamad Moustafa Ali, Pengwei Xing, Wenyi Zheng, André Görgens, Manuela O Gustafsson, Samir El Andaloussi, Enikö Sonkoly, Andor Pivarcsi.

Cutaneous squamous cell carcinoma (cSCC) is a fast-increasing cancer with metastatic potential. Extracellular vesicles (EVs) are small membrane-bound vesicles that play important roles in intercellular communication, particularly in the tumor microenvironment (TME). Here we report that cSCC cells secrete an increased number of EVs relative to normal human epidermal keratinocytes (NHEKs) and that interfering with the capacity of cSCC to secrete EVs inhibits tumor growth in vivo in a xenograft model of human cSCC. Transcriptome analysis of tumor xenografts by RNA-sequencing enabling the simultaneous quantification of both the human and the mouse transcripts revealed that impaired EV-production of cSCC cells prominently altered the phenotype of stromal cells, in particular genes related to extracellular matrix (ECM)-formation and epithelial-mesenchymal transition (EMT). In line with these results, co-culturing of human dermal fibroblasts (HDFs) with cSCC cells, but not with normal keratinocytes in vitro resulted in acquisition of cancer-associated fibroblast (CAF) phenotype. Interestingly, EVs derived from metastatic cSCC cells, but not primary cSCCs or NHEKs, were efficient in converting HDFs to CAFs. Multiplex bead-based flow cytometry assay and mass-spectrometry (MS)-based proteomic analyses revealed the heterogenous cargo of cSCC-derived EVs and that especially EVs derived from metastatic cSCCs carry proteins associated with EV-biogenesis, EMT, and cell migration. Mechanistically, EVs from metastatic cSCC cells result in the activation of TGFβ signaling in HDFs. Altogether, our study suggests that cSCC-derived EVs mediate cancer-stroma communication, in particular the conversion of fibroblasts to CAFs, which eventually contribute to cSCC progression.

DOI: https://doi.org/10.1038/s41420-023-01555-2
Immunity. 2023 Jul 18. pii: S1074-7613(23)00283-2. [Epub ahead of print]

Cell-autonomous regulation of complement C3 by factor H limits macrophage efferocytosis and exacerbates atherosclerosis.

Máté G Kiss, Nikolina Papac-Miličević, Florentina Porsch, Dimitrios Tsiantoulas, Tim Hendrikx, Minoru Takaoka, Huy Q Dinh, Marie-Sophie Narzt, Laura Göderle, Mária Ozsvár-Kozma, Michael Schuster, Nikolaus Fortelny, Anastasiya Hladik, Sylvia Knapp, Florian Gruber, Matthew C Pickering, Christoph Bock, Filip K Swirski, Klaus Ley, Alma Zernecke, Clément Cochain, Claudia Kemper, Ziad Mallat, Christoph J Binder.

  Complement factor H (CFH) negatively regulates consumption of complement component 3 (C3), thereby restricting complement activation. Genetic variants in CFH predispose to chronic inflammatory disease. Here, we examined the impact of CFH on atherosclerosis development. In a mouse model of atherosclerosis, CFH deficiency limited plaque necrosis in a C3-dependent manner. Deletion of CFH in monocyte-derived inflammatory macrophages propagated uncontrolled cell-autonomous C3 consumption without downstream C5 activation and heightened efferocytotic capacity. Among leukocytes, Cfh expression was restricted to monocytes and macrophages, increased during inflammation, and coincided with the accumulation of intracellular C3. Macrophage-derived CFH was sufficient to dampen resolution of inflammation, and hematopoietic deletion of CFH in atherosclerosis-prone mice promoted lesional efferocytosis and reduced plaque size. Furthermore, we identified monocyte-derived inflammatory macrophages expressing C3 and CFH in human atherosclerotic plaques. Our findings reveal a regulatory axis wherein CFH controls intracellular C3 levels of macrophages in a cell-autonomous manner, evidencing the importance of on-site complement regulation in the pathogenesis of inflammatory diseases.

Keywords:  atherosclerosis; cell-autonomous complement; complement factor H; complement protein C3; efferocytosis; inflammation; local complement production; macrophages; plaque necrosis

DOI:  https://doi.org/10.1016/j.immuni.2023.06.026
Front Endocrinol (Lausanne). 2023 ;14 1197102

Metabolite-sensing GPCRs controlling interactions between adipose tissue and inflammation.

Elaine M Duncan, Luca Vita, Bethany Dibnah, Brian D Hudson.

  Metabolic disorders including obesity, diabetes and non-alcoholic steatohepatitis are a group of conditions characterised by chronic low-grade inflammation of metabolic tissues. There is now a growing appreciation that various metabolites released from adipose tissue serve as key signalling mediators, influencing this interaction with inflammation. G protein-coupled receptors (GPCRs) are the largest family of signal transduction proteins and most historically successful drug targets. The signalling pathways for several key adipose metabolites are mediated through GPCRs expressed both on the adipocytes themselves and on infiltrating macrophages. These include three main groups of GPCRs: the FFA4 receptor, which is activated by long chain free fatty acids; the HCA2 and HCA3 receptors, activated by hydroxy carboxylic acids; and the succinate receptor. Understanding the roles these metabolites and their receptors play in metabolic-immune interactions is critical to establishing how these GPCRs may be exploited for the treatment of metabolic disorders.

Keywords:  G protein-coupled receptor; adipose; free fatty acid; hydroxy carboxylic acids; inflammation; metabolite signalling; succinate

DOI:  https://doi.org/10.3389/fendo.2023.1197102
Elife. 2023 Jul 25. pii: e84782. [Epub ahead of print]12

Melanocortin 1 receptor regulates cholesterol and bile acid metabolism in the liver.

Keshav Thapa, James J Kadiri, Karla Saukkonen, Iida Pennanen, Bishwa Ghimire, Minying Cai, Eriika Savontaus, Petteri Rinne.

  Melanocortin 1 receptor (MC1-R) is widely expressed in melanocytes and leukocytes and is thus strongly implicated in the regulation of skin pigmentation and inflammation. MC1-R has also been found in the rat and human liver, but its functional role has remained elusive. We hypothesized that MC1-R is functionally active in the liver and involved in the regulation of cholesterol and bile acid metabolism. We generated hepatocyte-specific MC1-R knock-out (Mc1r LKO) mice and phenotyped the mouse model for lipid profiles, liver histology, and bile acid levels. Mc1r LKO mice had significantly increased liver weight, which was accompanied by elevated levels of total cholesterol and triglycerides in the liver as well as in the plasma. These mice demonstrated also enhanced liver fibrosis and a disturbance in bile acid metabolism as evidenced by markedly reduced bile acid levels in the plasma and feces. Mechanistically, using HepG2 cells as an in vitro model, we found that selective activation of MC1-R in HepG2 cells reduced cellular cholesterol content and enhanced uptake of low- and high-density lipoprotein particles via a cAMP-independent mechanism. In conclusion, the present results demonstrate that MC1-R signaling in hepatocytes regulates cholesterol and bile acid metabolism and its deficiency leads to hypercholesterolemia and enhanced lipid accumulation and fibrosis in the liver.

Keywords:  bile acid; cell biology; cholesterol; fibrosis; medicine; melanocortin 1 receptor; mouse; steatosis; triglyceride

DOI:  https://doi.org/10.7554/eLife.84782
Eur J Immunol. 2023 Jul 22. e2350435

The vitamin B5/coenzyme A axis: A target for immunomodulation?

Miallot Richard, Millet Virginie, Galland Franck, Naquet Philippe.

  Coenzyme A (CoA) serves as a vital cofactor in numerous enzymatic reactions involved in energy production, lipid metabolism, and synthesis of essential molecules. Dysregulation of CoA-dependent metabolic pathways can contribute to chronic diseases, such as inflammatory diseases, obesity, diabetes, cancer, and cardiovascular disorders. Additionally, CoA influences immune cell activation by modulating the metabolism of these cells, thereby affecting their proliferation, differentiation, and effector functions. Targeting CoA metabolism presents a promising avenue for therapeutic intervention, as it can potentially restore metabolic balance, mitigate chronic inflammation, and enhance immune cell function. This might ultimately improve the management and outcomes for these diseases. This review will more specifically focus on the contribution of pathways regulating the availability of the CoA precursor Vitamin B5/pantothenate in vivo and modulating the development of Th17-mediated inflammation, CD8-dependent anti-tumor immunity but also tissue repair processes in chronic inflammatory or degenerative diseases.

Keywords:  Coenzyme A; Inflammation; Tissue repair; Tumor immunity; Vitamin B5

DOI:  https://doi.org/10.1002/eji.202350435
Proc Natl Acad Sci U S A. 2023 08;120(31): e2302938120

Resolvin D1 prevents injurious neutrophil swarming in transplanted lungs.

Wenjun Li, Hailey M Shepherd, Yuriko Terada, Ashley E Shay, Amit I Bery, Andrew E Gelman, Kory J Lavine, Charles N Serhan, Daniel Kreisel.

  Neutrophils are the primary cell type involved in lung ischemia-reperfusion injury (IRI), which remains a frequent and morbid complication after organ transplantation. Endogenous lipid mediators that become activated during acute inflammation-resolution have gained increasing recognition for their protective role(s) in promoting the restoration of homeostasis, but their influence on early immune responses following transplantation remains to be uncovered. Resolvin D1, 7S,8R,17S-trihydroxy-4Z,9E,11E,13Z,15E,19Z-docosahexaenoic acid (RvD1), is a potent stereoselective mediator that exhibits proresolving and anti-inflammatory actions in the setting of tissue injury. Here, using metabololipidomics, we demonstrate that endogenous proresolving mediators including RvD1 are increased in human and murine lung grafts immediately following transplantation. In mouse grafts, we observe lipid mediator class switching early after reperfusion. We use intravital two-photon microscopy to reveal that RvD1 treatment significantly limits early neutrophil infiltration and swarming, thereby ameliorating early graft dysfunction in transplanted syngeneic lungs subjected to severe IRI. Through integrated analysis of single-cell RNA sequencing data of donor and recipient immune cells from lung grafts, we identify transcriptomic changes induced by RvD1. These results support a role for RvD1 as a potent modality for preventing early neutrophil-mediated tissue damage after lung IRI that may be therapeutic in the clinics.

Keywords:  inflammation; leukocytes; lipid mediators; specialized proresolving mediators; transplantation

DOI:  https://doi.org/10.1073/pnas.2302938120
J Struct Biol. 2023 Jul 24. pii: S1047-8477(23)00067-9. [Epub ahead of print] 108004

Identification of structural determinants of nicotinamide phosphoribosyl transferase (NAMPT) activity and substrate selectivity.

Dorothée Houry, Arne Raasakka, Eugenio Ferrario, Marc Niere, Ersilia Bifulco, Petri Kursula, Mathias Ziegler.

  NAD homeostasis in mammals requires the salvage of nicotinamide (Nam), which is cleaved from NAD± by sirtuins, PARPs, and other NAD±-dependent signaling enzymes. Nam phosphoribosyltransferase (NAMPT) catalyzes the rate-limiting step in vitamin B3 salvage, whereby Nam reacts with phosphoribosyl pyrophosphate (PRPP) to form nicotinamide mononucleotide. NAMPT has a high affinity towards Nam, which is further enhanced by autophosphorylation of His247. The mechanism of this enhancement has remained unknown. Here, we present high-resolution crystal structures and biochemical data that provide reasoning for the increased affinity of the phosphorylated NAMPT for its substrate. Structural and kinetic analyses suggest a mechanism that includes Mg2+ coordination by phospho-His247, such that PRPP is stabilized in a position highly favorable for catalysis. Under these conditions, nicotinic acid (NA) can serve as a substrate. Moreover, we demonstrate that a stretch of 10 amino acids, present only in NAMPTs from deuterostomes, facilitates conformational plasticity and stabilizes the chemically unstable phosphorylation of His247. Thereby the apparent substrate affinity is considerably enhanced compared to prokaryotic NAMPTs. Collectively, our study provides a structural basis for the important function of NAMPT to recycle Nam into NAD biosynthesis with high affinity.

Keywords:  NAD biosynthesis; NAMPT; PARP; SAXS; X-ray crystallography; nicotinamide; nicotinic acid; phosphoribosyltransferase; sirtuins

DOI:  https://doi.org/10.1016/j.jsb.2023.108004
Anal Chem. 2023 Jul 28.

Lipid Signatures and Inter-Cellular Heterogeneity of Naı̈ve and Lipopolysaccharide-Stimulated Human Microglia-like Cells.

Max A Müller, Norman Zweig, Bernhard Spengler, Maria Weinert, Sven Heiles.

Microglia are non-neuronal cells, which reside in the central nervous system and are known to play an important role in health and disease. We investigated the lipidomic phenotypes of human naı̈ve and stimulated microglia-like cells by atmospheric-pressure scanning microprobe matrix-assisted laser desorption/ionization mass spectrometry imaging (AP-SMALDI MSI). With lateral resolutions between 5 and 1.5 μm pixel size, we were able to chart lipid compositions of individual cells, enabling differentiation of cell lines and stimulation conditions. This allowed us to reveal local lipid heterogeneities in naı̈ve and lipopolysaccharide (LPS)-stimulated cells. We were able to identify individual cells with elevated triglyceride (TG) levels and could show that the number of these TG-enriched cells increased with LPS stimulation as a hallmark for a proinflammatory phenotype. Additionally, the observed local abundance alterations of specific phosphatidylinositols (PIs) indicate a cell specific regulation of the PI metabolism.

DOI: https://doi.org/10.1021/acs.analchem.3c01533
Sci Adv. 2023 Jul 28. 9(30): eadd6997

Loss of metabolic fitness drives tumor resistance after CAR-NK cell therapy and can be overcome by cytokine engineering.

Li Li, Vakul Mohanty, Jinzhuang Dou, Yuefan Huang, Pinaki P Banerjee, Qi Miao, Jens G Lohr, Tushara Vijaykumar, Julia Frede, Birgit Knoechel, Luis Muniz-Feliciano, Tamara J Laskowski, Shaoheng Liang, Judy S Moyes, Vandana Nandivada, Rafet Basar, Mecit Kaplan, May Daher, Enli Liu, Ye Li, Sunil Acharya, Paul Lin, Mayra Shanley, Hind Rafei, David Marin, Stephan Mielke, Richard E Champlin, Elizabeth J Shpall, Ken Chen, Katayoun Rezvani.

Chimeric antigen receptor (CAR) engineering of natural killer (NK) cells is promising, with early-phase clinical studies showing encouraging responses. However, the transcriptional signatures that control the fate of CAR-NK cells after infusion and factors that influence tumor control remain poorly understood. We performed single-cell RNA sequencing and mass cytometry to study the heterogeneity of CAR-NK cells and their in vivo evolution after adoptive transfer, from the phase of tumor control to relapse. Using a preclinical model of noncurative lymphoma and samples from a responder and a nonresponder patient treated with CAR19/IL-15 NK cells, we observed the emergence of NK cell clusters with distinct patterns of activation, function, and metabolic signature associated with different phases of in vivo evolution and tumor control. Interaction with the highly metabolically active tumor resulted in loss of metabolic fitness in NK cells that could be partly overcome by incorporation of IL-15 in the CAR construct.

DOI: https://doi.org/10.1126/sciadv.add6997
EMBO Rep. 2023 Jul 25. e56279

Arginine dependency is a therapeutically exploitable vulnerability in chronic myeloid leukaemic stem cells.

Kevin M Rattigan, Martha M Zarou, Zuzana Brabcova, Bodhayan Prasad, Désirée Zerbst, Daniele Sarnello, Eric R Kalkman, Angela Ianniciello, Mary T Scott, Karen Dunn, Engy Shokry, David Sumpton, Mhairi Copland, Saverio Tardito, Johan Vande Voorde, Francis Mussai, Paul Cheng, G Vignir Helgason.

  To fuel accelerated proliferation, leukaemic cells undergo metabolic deregulation, which can result in specific nutrient dependencies. Here, we perform an amino acid drop-out screen and apply pre-clinical models of chronic phase chronic myeloid leukaemia (CML) to identify arginine as a nutrient essential for primary human CML cells. Analysis of the Microarray Innovations in Leukaemia (MILE) dataset uncovers reduced ASS1 levels in CML compared to most other leukaemia types. Stable isotope tracing reveals repressed activity of all urea cycle enzymes in patient-derived CML CD34+ cells, rendering them arginine auxotrophic. Thus, arginine deprivation completely blocks proliferation of CML CD34+ cells and induces significantly higher levels of apoptosis when compared to arginine-deprived cell lines. Similarly, primary CML cells, but not normal CD34+ samples, are particularly sensitive to treatment with the arginine-depleting enzyme, BCT-100, which induces apoptosis and reduces clonogenicity. Moreover, BCT-100 is highly efficacious in a patient-derived xenograft model, causing > 90% reduction in the number of human leukaemic stem cells (LSCs). These findings indicate arginine depletion to be a promising and novel strategy to eradicate therapy resistant LSCs.

Keywords:  amino acids; leukaemic stem cells; metabolism; therapy resistance

DOI:  https://doi.org/10.15252/embr.202256279
Nat Metab. 2023 Jul;5(7): 1088-1100

Interleukin-17 as a key player in neuroimmunometabolism.

Aaron Douglas, Brenneth Stevens, Lydia Lynch.

In mammals, interleukin (IL)-17 cytokines are produced by innate and adaptive lymphocytes. However, the IL-17 family has widespread expression throughout evolution, dating as far back as cnidaria, molluscs and worms, which predate lymphocytes. The evolutionary conservation of IL-17 suggests that it is involved in innate defence strategies, but also that this cytokine family has a fundamental role beyond typical host defence. Throughout evolution, IL-17 seems to have a major function in homeostatic maintenance at barrier sites. Most recently, a pivotal role has been identified for IL-17 in regulating cellular metabolism, neuroimmunology and tissue physiology, particularly in adipose tissue. Here we review the emerging role of IL-17 signalling in regulating metabolic processes, which may shine a light on the evolutionary role of IL-17 beyond typical immune responses. We propose that IL-17 helps to coordinate the cross-talk among the nervous, endocrine and immune systems for whole-body energy homeostasis as a key player in neuroimmunometabolism.

DOI: https://doi.org/10.1038/s42255-023-00846-3
Cell Death Dis. 2023 07 25. 14(7): 463

A GSTP1-mediated lactic acid signaling promotes tumorigenesis through the PPP oxidative branch.

Yandi Sun, Qian He, Jingjia Li, Ze Yang, Mashaal Ahmad, Yindan Lin, Di Wu, Lei Zheng, Jiangtao Li, Ben Wang, Chitty Chen, Yue Hu, Heng Luo, Yan Luo.

Lactic acidosis is a feature of solid tumors and plays fundamental role(s) rendering cancer cells to adapt to diverse metabolic stresses, but the mechanism underlying its roles in redox homeostasis remains elusive. Here we show that G6PD is phosphorylated at tyrosine 249/322 by the SRC through the formation of a GSTP1-G6PD-SRC complex. Lactic acid attenuates this formation and the phosphorylation of G6PD by non-covalently binding with GSTP1. Furthermore, lactic acid increases the activity of G6PD and facilitates the PPP (NADPH production) through its sensor GSTP1, thereby exhibiting resistance to reactive oxygen species when glucose is scarce. Abrogating a GSTP1-mediated lactic acid signaling showed attenuated tumor growth and reduced resistance to ROS in breast cancer cells. Importantly, positive correlations between immuno-enriched SRC protein and G6PD Y249/322 phosphorylation specifically manifest in ER/PR positive or HER negative types of breast cancer. Taken together, these results suggest that GSTP1 plays a key role in tumor development by functioning as a novel lactate sensor.

DOI: https://doi.org/10.1038/s41419-023-05998-4
EMBO J. 2023 Jul 27. e114802

Stearate-derived very long-chain fatty acids are indispensable to tumor growth.

Qiaoyun Chu, Ping Liu, Yihan Song, Ronghui Yang, Jing An, Xuewei Zhai, Jing Niu, Chuanzhen Yang, Binghui Li.

DOI: https://doi.org/10.15252/embj.2023114802
Cell Death Dis. 2023 07 26. 14(7): 467

Nitric oxide-induced ribosome collision activates ribosomal surveillance mechanisms.

Laura Ryder, Frederic Schrøder Arendrup, José Francisco Martínez, Goda Snieckute, Chiara Pecorari, Riyaz Ahmad Shah, Anders H Lund, Melanie Blasius, Simon Bekker-Jensen.

Impairment of protein translation can cause stalling and collision of ribosomes and is a signal for the activation of ribosomal surveillance and rescue pathways. Despite clear evidence that ribosome collision occurs stochastically at a cellular and organismal level, physiologically relevant sources of such aberrations are poorly understood. Here we show that a burst of the cellular signaling molecule nitric oxide (NO) reduces translational activity and causes ribosome collision in human cell lines. This is accompanied by activation of the ribotoxic stress response, resulting in ZAKα-mediated activation of p38 and JNK kinases. In addition, NO production is associated with ZNF598-mediated ubiquitination of the ribosomal protein RPS10 and GCN2-mediated activation of the integrated stress response, which are well-described responses to the collision of ribosomes. In sum, our work implicates a novel role of NO as an inducer of ribosome collision and activation of ribosomal surveillance mechanisms in human cells.

DOI: https://doi.org/10.1038/s41419-023-05997-5
Cell Metab. 2023 Jul 21. pii: S1550-4131(23)00251-6. [Epub ahead of print]

Tumor aerobic glycolysis confers immune evasion through modulating sensitivity to T cell-mediated bystander killing via TNF-α.

Lijian Wu, Yiteng Jin, Xi Zhao, Kaiyang Tang, Yaoning Zhao, Linjie Tong, Xuerong Yu, Ke Xiong, Ce Luo, Jiajun Zhu, Fubing Wang, Zexian Zeng, Deng Pan.

  Metabolic reprogramming toward glycolysis is a hallmark of cancer malignancy. The molecular mechanisms by which the tumor glycolysis pathway promotes immune evasion remain to be elucidated. Here, by performing genome-wide CRISPR screens in murine tumor cells co-cultured with cytotoxic T cells (CTLs), we identified that deficiency of two important glycolysis enzymes, Glut1 (glucose transporter 1) and Gpi1 (glucose-6-phosphate isomerase 1), resulted in enhanced killing of tumor cells by CTLs. Mechanistically, Glut1 inactivation causes metabolic rewiring toward oxidative phosphorylation, which generates an excessive amount of reactive oxygen species (ROS). Accumulated ROS potentiate tumor cell death mediated by tumor necrosis factor alpha (TNF-α) in a caspase-8- and Fadd-dependent manner. Genetic and pharmacological inactivation of Glut1 sensitizes tumors to anti-tumor immunity and synergizes with anti-PD-1 therapy through the TNF-α pathway. The mechanistic interplay between tumor-intrinsic glycolysis and TNF-α-induced killing provides new therapeutic strategies to enhance anti-tumor immunity.

Keywords:  T cell-mediated killing; TNF-α; glycolysis; immune evasion

DOI:  https://doi.org/10.1016/j.cmet.2023.07.001
Eur J Immunol. 2023 Jul 25. e2350474

Invariant natural killer T cells drive hepatic homeostasis in Non-alcoholic fatty liver disease via sustained IL-10 expression in CD170+ Kupffer cells.

Mutian Han, Jinke Geng, Shuangshuang Zhang, Jia Rao, Yansong Zhu, Shaodong Xu, Fei Wang, Fang Ma, Meng Zhou, Hong Zhou.

  Kupffer cells (KCs) are liver-resident macrophages involved in hepatic inflammatory responses, including non-alcoholic fatty liver disease (NAFLD) development. However, the contribution of KC subsets to liver inflammation remains unclear. Here, using high-dimensional single-cell RNA sequencing, we characterised murine embryo-derived KCs (Em-KCs) and identified two KC populations with different gene expression profiles: KC-1 and KC-2. KC-1 expressed CD170, exhibiting immunoreactivity and immune-regulatory abilities, while KC-2 highly expressed lipid metabolism-associated genes. In a high-fat diet (HFD)-induced NAFLD model, KC-1 cells differentiated into pro-inflammatory phenotypes and initiated more frequent communications with invariant natural killer T (iNKT) cells. In KC-1, interleukin (IL)-10 expression was unaffected by the HFD but impaired by iNKT cell ablation and upregulated by iNKT cell adoptive transfer in vivo. Moreover, in a cellular co-culture system, primary hepatic iNKT cells promoted IL-10 expression in RAW264.7 and primary KC-1 cells. CD206 signal blocking in KC-1 or CD206 knockdown in RAW264.7 cells significantly reduced IL-10 expression. In conclusion, we identified two Em-KC subpopulations with distinct transcriptional profiles. The CD206-mediated crosstalk between iNKT and KC-1 cells maintains IL-10 expression in KC-1 cells, affecting hepatic immune balance. Therefore, KC-based therapeutic strategies must consider cellular heterogeneity and the local immune microenvironment for enhanced specificity and efficiency. This article is protected by copyright. All rights reserved.

Keywords:  Kupffer cells; immune homeostasis; invariant natural killer T cells; non-alcoholic fatty liver disease

DOI:  https://doi.org/10.1002/eji.202350474
Curr Opin Biotechnol. 2023 Jul 24. pii: S0958-1669(23)00080-0. [Epub ahead of print]83 102970

Tumor interstitial fluid analysis enables the study of microenvironment-cell interactions in cancers.

Juan J Apiz Saab, Alexander Muir.

The tumor microenvironment (TME) plays a crucial role in regulating the state and function of all cell types residing in the tumor and thus impacts many aspects of tumor biology. The importance of the TME has led to an interest in characterizing the composition of the TME and how TME components regulate cancer and stromal cell biology. Tumor interstitial fluid (TIF) is the local perfusate of the TME that carries metabolites, electrolytes, and soluble macromolecules to tumor-resident cells. Recently, techniques to isolate TIF have been coupled with analytical techniques to interrogate the composition of TIF, providing new insight into TME composition. In this review, we will discuss what TIF studies indicate about TME composition and new avenues TIF analysis provides to delineate how the TME regulates tumor biology.

DOI: https://doi.org/10.1016/j.copbio.2023.102970
iScience. 2023 Jul 21. 26(7): 107230

Alcohol-sourced acetate impairs T cell function by promoting cortactin acetylation.

Vugar Azizov, Michel Hübner, Michael Frech, Jörg Hofmann, Marketa Kubankova, Dennis Lapuente, Matthias Tenbusch, Jochen Guck, Georg Schett, Mario M Zaiss.

  Alcohol is among the most widely consumed dietary substances. Excessive alcohol consumption damages the liver, heart, and brain. Alcohol also has strong immunoregulatory properties. Here, we report how alcohol impairs T cell function via acetylation of cortactin, a protein that binds filamentous actin and facilitates branching. Upon alcohol consumption, acetate, the metabolite of alcohol, accumulates in lymphoid organs. T cells exposed to acetate, exhibit increased acetylation of cortactin. Acetylation of cortactin inhibits filamentous actin binding and hence reduces T cell migration, immune synapse formation and activation. While mutated, acetylation-resistant cortactin rescues the acetate-induced inhibition of T cell migration, primary mouse cortactin knockout T cells exhibited impaired migration. Acetate-induced cytoskeletal changes effectively inhibited activation, proliferation, and immune synapse formation in T cells in vitro and in vivo in an influenza infection model in mice. Together these findings reveal cortactin as a possible target for mitigation of T cell driven autoimmune diseases.

Keywords:  Immunology; Molecular biology

DOI:  https://doi.org/10.1016/j.isci.2023.107230
J Biol Chem. 2023 Jul 21. pii: S0021-9258(23)02107-5. [Epub ahead of print] 105079

Acidosis attenuates CPT-I supported bioenergetics as a potential mechanism limiting lipid oxidation.

Sara M Frangos, Geneviève J DesOrmeaux, Graham P Holloway.

  Fuel interactions in contracting muscle represent a complex interplay between enzymes regulating carbohydrate and fatty acid catabolism, converging in the mitochondrial matrix. While increasing exercise intensity promotes carbohydrate use at the expense of fatty acid oxidation, the mechanisms underlying this effect remain poorly elucidated. As a potential explanation, we investigated whether exercise-induced reductions in intramuscular pH (acidosis) attenuate carnitine palmitoyltransferase-I (CPT-I) supported bioenergetics, the rate-limiting step for fatty acid oxidation within mitochondria. Specifically, we assessed the effect of a physiologically relevant reduction in pH (pH 7.2 vs 6.8) on single and mixed substrate respiratory responses in murine skeletal muscle isolated mitochondria and permeabilized fibers. While pH did not influence OXPHOS stoichiometry (ADP/O ratios), coupling efficiency, oxygen affinity or ADP respiratory responses, acidosis impaired lipid bioenergetics by attenuating respiration with L-carnitine and palmitoyl-CoA, while enhancing the inhibitory effect of malonyl-CoA on CPT-I. These acidotic effects were largely retained following a single bout of intense exercise. At rest, pyruvate and succinate supported respiration were also impaired by acidosis. However, providing more pyruvate and ADP at pH 6.8 to model increases in glycolytic flux and ATP turnover with intense exercise, overcame the acidotic attenuation of carbohydrate-linked OXPHOS. Importantly, this situation is fundamentally different from lipids where CPT-I substrate sensitivity and availability is impaired at higher power outputs suggesting lipid metabolism may be more susceptible to the effects of acidosis, possibly contributing to fuel shifts with increasing exercise intensity.

Keywords:  CPT-I; Mitochondrial bioenergetics; exercise; fuel metabolism; lipid metabolism; pH; skeletal muscle

DOI:  https://doi.org/10.1016/j.jbc.2023.105079
Cell Death Dis. 2023 Jul 27. 14(7): 477

A transient increase of HIF-1α during the G1 phase (G1-HIF) ensures cell survival under nutritional stress.

Ratnal Belapurkar, Maximilian Pfisterer, Jan Dreute, Sebastian Werner, Sven Zukunft, Ingrid Fleming, Michael Kracht, M Lienhard Schmitz.

The family of hypoxia-inducible transcription factors (HIF) is activated to adapt cells to low oxygen conditions, but is also known to regulate some biological processes under normoxic conditions. Here we show that HIF-1α protein levels transiently increase during the G1 phase of the cell cycle (designated as G1-HIF) in an AMP-activated protein kinase (AMPK)-dependent manner. The transient elimination of G1-HIF by a degron system revealed its contribution to cell survival under unfavorable metabolic conditions. Indeed, G1-HIF plays a key role in the cell cycle-dependent expression of genes encoding metabolic regulators and the maintenance of mTOR activity under conditions of nutrient deprivation. Accordingly, transient elimination of G1-HIF led to a significant reduction in the concentration of key proteinogenic amino acids and carbohydrates. These data indicate that G1-HIF acts as a cell cycle-dependent surveillance factor that prevents the onset of starvation-induced apoptosis.

DOI: https://doi.org/10.1038/s41419-023-06012-7
Proc Natl Acad Sci U S A. 2023 08;120(31): e2300475120

Nascent mitochondrial proteins initiate the localized condensation of cytosolic protein aggregates on the mitochondrial surface.

Qingqing Liu, Benjamin Fong, Seungmin Yoo, Jay R Unruh, Fengli Guo, Zulin Yu, Jingjing Chen, Kausik Si, Rong Li, Chuankai Zhou.

  Eukaryotes organize cellular contents into membrane-bound organelles and membrane-less condensates, for example, protein aggregates. An unsolved question is why the ubiquitously distributed proteins throughout the cytosol give rise to spatially localized protein aggregates on the organellar surface, like mitochondria. We report that the mitochondrial import receptor Tom70 is involved in the localized condensation of protein aggregates in budding yeast and human cells. This is because misfolded cytosolic proteins do not autonomously aggregate in vivo; instead, they are recruited to the condensation sites initiated by Tom70's substrates (nascent mitochondrial proteins) on the organellar membrane using multivalent hydrophobic interactions. Knocking out Tom70 partially impairs, while overexpressing Tom70 increases the formation and association between cytosolic protein aggregates and mitochondria. In addition, ectopic targeting Tom70 and its substrates to the vacuole surface is able to redirect the localized aggregation from mitochondria to the vacuolar surface. Although other redundant mechanisms may exist, this nascent mitochondrial proteins-based initiation of protein aggregation likely explains the localized condensation of otherwise ubiquitously distributed molecules on the mitochondria. Disrupting the mitochondrial association of aggregates impairs their asymmetric retention during mitosis and reduces the mitochondrial import of misfolded proteins, suggesting a proteostasis role of the organelle-condensate interactions.

Keywords:  condensate; mitochondria; protein aggregation

DOI:  https://doi.org/10.1073/pnas.2300475120
Nat Commun. 2023 07 25. 14(1): 4476

Supersulphides provide airway protection in viral and chronic lung diseases.

Tetsuro Matsunaga, Hirohito Sano, Katsuya Takita, Masanobu Morita, Shun Yamanaka, Tomohiro Ichikawa, Tadahisa Numakura, Tomoaki Ida, Minkyung Jung, Seiryo Ogata, Sunghyeon Yoon, Naoya Fujino, Yorihiko Kyogoku, Yusaku Sasaki, Akira Koarai, Tsutomu Tamada, Atsuhiko Toyama, Takakazu Nakabayashi, Lisa Kageyama, Shigeru Kyuwa, Kenji Inaba, Satoshi Watanabe, Péter Nagy, Tomohiro Sawa, Hiroyuki Oshiumi, Masakazu Ichinose, Mitsuhiro Yamada, Hisatoshi Sugiura, Fan-Yan Wei, Hozumi Motohashi, Takaaki Akaike.

Supersulphides are inorganic and organic sulphides with sulphur catenation with diverse physiological functions. Their synthesis is mainly mediated by mitochondrial cysteinyl-tRNA synthetase (CARS2) that functions as a principal cysteine persulphide synthase (CPERS). Here, we identify protective functions of supersulphides in viral airway infections (influenza and COVID-19), in aged lungs and in chronic lung diseases, including chronic obstructive pulmonary disease (COPD), idiopathic pulmonary fibrosis (IPF). We develop a method for breath supersulphur-omics and demonstrate that levels of exhaled supersulphides increase in people with COVID-19 infection and in a hamster model of SARS-CoV-2 infection. Lung damage and subsequent lethality that result from oxidative stress and inflammation in mouse models of COPD, IPF, and ageing were mitigated by endogenous supersulphides production by CARS2/CPERS or exogenous administration of the supersulphide donor glutathione trisulphide. We revealed a protective role of supersulphides in airways with various viral or chronic insults and demonstrated the potential of targeting supersulphides in lung disease.

DOI: https://doi.org/10.1038/s41467-023-40182-4
Blood Adv. 2023 Jul 28. pii: bloodadvances.2023009967. [Epub ahead of print]

Combined GLUT1 and OXPHOS inhibition eliminates acute myeloid leukemia cells by restraining their metabolic plasticity.

Maria Rodriguez-Zabala, Ramprasad Ramakrishnan, Katrin Reinbach, Somadri Ghosh, Leal Oburoglu, Antoni Falqués-Costa, Kishan Bellamkonda, Mats Ehinger, Pablo Peña-Martínez, Noelia Puente-Moncada, Henrik Lilljebjörn, Jörg Cammenga, Cornelis Jan Pronk, Vladimir Lazarevic, Thoas Fioretos, Anna Hagström-Andersson, Niels-Bjarne Woods, Marcus Järås.

Acute myeloid leukemia (AML) is initiated and propagated by leukemia stem cells (LSCs), a self-renewing population of leukemia cells responsible for therapy resistance. Hence, there is an urgent need to identify new therapeutic opportunities targeting LSCs. Here we performed an in vivo CRISPR knockout screen to identify potential therapeutic targets by interrogating cell surface dependencies of LSCs. The facilitated glucose transporter type 1 (GLUT1) emerged as a critical in vivo metabolic dependency for LSCs in a murine MLL::AF9-driven model of AML. GLUT1 disruption by genetic ablation or pharmacological inhibition led to suppression of leukemia progression and improved survival of mice transplanted with LSCs. Metabolic profiling revealed that Glut1 inhibition suppressed glycolysis, decreased levels of tricarboxylic acid (TCA) cycle intermediates, and increased the levels of amino acids. This metabolic reprogramming was accompanied by an increase in autophagic activity and apoptosis. Moreover, Glut1 disruption caused transcriptional, morphological and immunophenotypic changes consistent with differentiation of AML cells. Notably, dual inhibition of GLUT1 and oxidative phosphorylation (OXPHOS) exhibited synergistic anti-leukemic effects in the majority of primary AML patient samples tested by restraining their metabolic plasticity. In particular, RUNX1-mutated primary leukemia cells displayed striking sensitivity to the combination treatment compared to normal CD34+ bone marrow and cord blood cells. Collectively, our study reveals a GLUT1 dependency of murine LSCs in the bone marrow microenvironment, and demonstrates that dual inhibition of GLUT1 and OXPHOS is a promising therapeutic approach for AML.

DOI: https://doi.org/10.1182/bloodadvances.2023009967
Cell Rep. 2023 Jul 24. pii: S2211-1247(23)00866-5. [Epub ahead of print]42(8): 112855

Phosphatidylinositol 3-phosphate regulates iron transport via PI3P-binding CgPil1 protein.

Fizza Askari, Bhogadi Vasavi, Rupinder Kaur.

  Iron homeostasis, which is pivotal to virulence, is regulated by the phosphatidylinositol 3-kinase CgVps34 in the human fungal pathogen Candida glabrata. Here, we identify CgPil1 as a phosphatidylinositol 3-phosphate (PI3P)-binding protein and unveil its role in retaining the high-affinity iron transporter CgFtr1 at the plasma membrane (PM), with PI3P negatively regulating CgFtr1-CgPil1 interaction. PI3P production and its PM localization are elevated in the high-iron environment. Surplus iron also leads to intracellular distribution and vacuolar delivery of CgPil1 and CgFtr1, respectively, from the PM. Loss of CgPil1 or CgFtr1 ubiquitination at lysines 391 and 401 results in CgFtr1 trafficking to the endoplasmic reticulum and a decrease in vacuole-localized CgFtr1. The E3-ubiquitin ligase CgRsp5 interacts with CgFtr1 and forms distinct CgRsp5-CgFtr1 puncta at the PM, with high iron resulting in their internalization. Finally, PI3P controls retrograde transport of many PM proteins. Altogether, we establish PI3P as a key regulator of membrane transport in C. glabrata.

Keywords:  CP: Cell biology; E3-ubiquitin ligase; PI3P; eisosome; human pathogenic fungi; iron transporter; phosphatidylinositol 3-phosphate; plasma membrane; ubiquitination

DOI:  https://doi.org/10.1016/j.celrep.2023.112855
EMBO Mol Med. 2023 Jul 25. e16858

Platelet-derived lipids promote insulin secretion of pancreatic β cells.

Till Karwen, Katarzyna Kolczynska-Matysiak, Carina Gross, Mona C Löffler, Mike Friedrich, Angel Loza-Valdes, Werner Schmitz, Magdalena Wit, Filip Dziaczkowski, Andrei Belykh, Jonathan Trujillo-Viera, Rabih El-Merahbi, Carsten Deppermann, Sameena Nawaz, Benoit Hastoy, Agnieszka Demczuk, Manuela Erk, Mariusz R Wieckowski, Patrik Rorsman, Katrin G Heinze, David Stegner, Bernhard Nieswandt, Grzegorz Sumara.

  Hyperreactive platelets are commonly observed in diabetic patients indicating a potential link between glucose homeostasis and platelet reactivity. This raises the possibility that platelets may play a role in the regulation of metabolism. Pancreatic β cells are the central regulators of systemic glucose homeostasis. Here, we show that factor(s) derived from β cells stimulate platelet activity and platelets selectively localize to the vascular endothelium of pancreatic islets. Both depletion of platelets and ablation of major platelet adhesion or activation pathways consistently resulted in impaired glucose tolerance and decreased circulating insulin levels. Furthermore, we found platelet-derived lipid classes to promote insulin secretion and identified 20-Hydroxyeicosatetraenoic acid (20-HETE) as the main factor promoting β cells function. Finally, we demonstrate that the levels of platelet-derived 20-HETE decline with age and that this parallels with reduced impact of platelets on β cell function. Our findings identify an unexpected function of platelets in the regulation of insulin secretion and glucose metabolism, which promotes metabolic fitness in young individuals.

Keywords:  20-HETE; diabetes; insulin secretion; platelet; β cell

DOI:  https://doi.org/10.15252/emmm.202216858
Proc Natl Acad Sci U S A. 2023 Aug;120(31): e2303789120

LRRK2 suppresses lysosome degradative activity in macrophages and microglia through MiT-TFE transcription factor inhibition.

Narayana Yadavalli, Shawn M Ferguson.

  Cells maintain optimal levels of lysosome degradative activity to protect against pathogens, clear waste, and generate nutrients. Here, we show that LRRK2, a protein that is tightly linked to Parkinson's disease, negatively regulates lysosome degradative activity in macrophages and microglia via a transcriptional mechanism. Depletion of LRRK2 and inhibition of LRRK2 kinase activity enhanced lysosomal proteolytic activity and increased the expression of multiple lysosomal hydrolases. Conversely, the kinase hyperactive LRRK2 G2019S Parkinson's disease mutant suppressed lysosomal degradative activity and gene expression. We identified MiT-TFE transcription factors (TFE3, TFEB, and MITF) as mediators of LRRK2-dependent control of lysosomal gene expression. LRRK2 negatively regulated the abundance and nuclear localization of these transcription factors and their depletion prevented LRRK2-dependent changes in lysosome protein levels. These observations define a role for LRRK2 in controlling lysosome degradative activity and support a model wherein LRRK2 hyperactivity may increase Parkinson's disease risk by suppressing lysosome degradative activity.

Keywords:  Parkinson’s disease; lysosome; macrophage; microglia

DOI:  https://doi.org/10.1073/pnas.2303789120
Cell Rep. 2023 Jul 26. pii: S2211-1247(23)00906-3. [Epub ahead of print]42(8): 112895

PGAM5 is an MFN2 phosphatase that plays an essential role in the regulation of mitochondrial dynamics.

Sudeshna Nag, Kaitlin Szederkenyi, Olena Gorbenko, Hannah Tyrrell, Christopher M Yip, G Angus McQuibban.

  Mitochondrial morphology is regulated by the post-translational modifications of the dynamin family GTPase proteins including mitofusin 1 (MFN1), MFN2, and dynamin-related protein 1 (DRP1). Mitochondrial phosphatase phosphoglycerate mutase 5 (PGAM5) is emerging as a regulator of these post-translational modifications; however, its precise role in the regulation of mitochondrial morphology is unknown. We show that PGAM5 interacts with MFN2 and DRP1 in a stress-sensitive manner. PGAM5 regulates MFN2 phosphorylation and consequently protects it from ubiquitination and degradation. Further, phosphorylation and dephosphorylation modification of MFN2 regulates its fusion ability. Phosphorylation enhances fission and degradation, whereas dephosphorylation enhances fusion. PGAM5 dephosphorylates MFN2 to promote mitochondrial network formation. Further, using a Drosophila genetic model, we demonstrate that the MFN2 homolog Marf and dPGAM5 are in the same biological pathway. Our results identify MFN2 dephosphorylation as a regulator of mitochondrial fusion and PGAM5 as an MFN2 phosphatase.

Keywords:  CP: Molecular biology; DRP1; MFN2; PGAM5; mitochondrial morphology

DOI:  https://doi.org/10.1016/j.celrep.2023.112895
J Biol Chem. 2023 Jul 24. pii: S0021-9258(23)02116-6. [Epub ahead of print] 105088

Competition for cysteine acylation by C16:0 and C18:0 derived lipids is a global phenomenon in the proteome.

Hana Nůsková, Fabiola Garcia Cortizo, Lena Sophie Schwenker, Timo Sachsenheimer, Egor Diakonov, Marcel Tiebe, Martin Schneider, Jasmin Lohbeck, Carissa Reid, Annette Kopp-Schneider, Dominic Helm, Britta Brügger, Aubry K Miller, Aurelio A Teleman.

  S-acylation is a reversible posttranslational protein modification consisting of attachment of a fatty acid to a cysteine via a thioester bond. Research over the last few years has shown that a variety of different fatty acids, such as palmitic acid (C16:0), stearate (C18:0) or oleate (C18:1), are used in cells to S-acylate proteins. We recently showed that GNAI proteins can be acylated on a single residue, Cys3, with either C16:0 or C18:1 and that the relative proportion of acylation with these fatty acids depends on the level of the respective fatty acid in the cell's environment. This has functional consequences for GNAI proteins, with the identity of the acylating fatty acid affecting the subcellular localization of GNAIs. Unclear is whether this competitive acylation is specific to GNAI proteins or a more general phenomenon in the proteome. We perform here a proteome screen to identify proteins acylated with different fatty acids. We identify 218 proteins acylated with C16:0 and 308 proteins acylated with C18-lipids, thereby uncovering novel targets of acylation. We find that most proteins that can be acylated by C16:0 can also be acylated with C18-fatty acids. For proteins with more than one acylation site, we find that this competitive acylation occurs on each individual cysteine residue. This raises the possibility that the function of many different proteins can be regulated by the lipid environment via differential S-acylation.

Keywords:  HRAS; LAMTOR1; S-oleoylation; S-palmitoylation; S-stearoylation; click chemistry; lipid; posttranslational modification; protein S-acylation; transferrin receptor 1

DOI:  https://doi.org/10.1016/j.jbc.2023.105088