bims-mepmim 2022-11-27 papers

bims-mepmim

Biomed News

on Metabolites in pathological microenvironments and immunometabolism

Issue of 2022‒11‒27
forty papers selected by
Erika Mariana Palmieri
NIH/NCI Laboratory of Cancer ImmunoMetabolism

DOT1L regulates lipid biosynthesis and inflammatory responses in macrophages and promotes atherosclerotic plaque stability.
Metabolic symbiosis in pancreatic cancer.
Molecular Mechanisms for Ketone Body Metabolism, Signaling Functions, and Therapeutic Potential in Cancer.
Stearate-derived very long-chain fatty acids are indispensable to tumor growth.
Differential integrated stress response and asparagine production drive symbiosis and therapy resistance of pancreatic adenocarcinoma cells.
GPSM1 impairs metabolic homeostasis by controlling a pro-inflammatory pathway in macrophages.
Temporal and spatial metabolite dynamics impart control in adipogenesis.
How neutrophil metabolism affects bacterial killing.
Glucagon Changes Substrate Preference in Gluconeogenesis.
Branched-chain amino acid catabolism breaks glutamine addiction to sustain hepatocellular carcinoma progression.
Macrophage acetyl-CoA carboxylase regulates acute inflammation through control of glucose and lipid metabolism.
Metabolic control of innate lymphoid cells in health and disease.
Metabolic Recycling Enhances Proliferation in MYC-Transformed Lymphoma B Cells.
Glutamine-dependent effects of nitric oxide on cancer cells subjected to hypoxia-reoxygenation.
Mitophagy bridges DNA sensing with metabolic adaption to expand lung cancer stem-like cells.
Alveolar macrophage metabolic programming via a C-type lectin receptor protects against lipo-toxicity and cell death.
The leucine-rich repeat (LRR) domain of NLRP3 is required for NLRP3 inflammasome activation in macrophages.
Role of miR-199a-5p in the post-transcriptional regulation of ABCA1 in response to hypoxia in peritoneal macrophages.
Macrophage-Derived 25-Hydroxycholesterol Promotes Vascular Inflammation, Atherogenesis, and Lesion Remodeling.
Gsk3β regulates the resolution of liver ischemia reperfusion injury via MerTK.
Manipulating PP2Acα-ASK-JNK signaling to favor apoptotic over necroptotic hepatocyte fate reduces the extent of necrosis and fibrosis upon acute liver injury.
G-Protein-Coupled Lactate Receptor GPR81 Control of VMNvl Glucoregulatory Neurotransmitter and 5'-AMP-Activated Protein Kinase Expression.
Directional regulation of cytosolic PEPCK catalysis is mediated by competitive binding of anions.
Fructose Impairs Fat Oxidation: Implications for the Mechanism of Western diet-induced NAFLD.
HGFAC is a ChREBP regulated hepatokine that enhances glucose and lipid homeostasis.
Sphingosine-1-phosphate controls endothelial sphingolipid homeostasis via ORMDL.
Copper homeostasis and cuproptosis in health and disease.
Integration of Liver Glycogen and Triglyceride NMR Isotopomer Analyses Provides a Comprehensive Coverage of Hepatic Glucose and Fructose Metabolism.
Control of Gαq signaling dynamics and GPCR cross-talk by GRKs.
COX7A1 enhances the sensitivity of human NSCLC cells to cystine deprivation-induced ferroptosis via regulating mitochondrial metabolism.
SRSF6 balances mitochondrial-driven innate immune outcomes through alternative splicing of BAX.
A deubiquitination module essential for Treg fitness in the tumor microenvironment.
Cyclic stretch regulates immune responses via tank-binding kinase 1 expression in macrophages.
The phosphorylation of AMPKβ1 is critical for increasing autophagy and maintaining mitochondrial homeostasis in response to fatty acids.
Structure of human phagocyte NADPH oxidase in the resting state.
Multi-omics profiling reveals Chitinase-3-like protein 1 as a key mediator in the crosstalk between sarcopenia and liver cancer.
Mitochondrial ferritin expression in human macrophages is facilitated by thrombin-mediated cleavage under hypoxia.
Hydroxylation site-specific and production-dependent effects of endogenous oxysterols on cholesterol homeostasis: Implications for SREBP-2 and LXR.
Cholesterol transfer via endoplasmic reticulum contacts mediates lysosome damage repair.
Modulation of oxidative neurometabolism in ischemia/reperfusion by nitrite.

Cell Rep. 2022 Nov 22. pii: S2211-1247(22)01577-7. [Epub ahead of print]41(8): 111703

DOT1L regulates lipid biosynthesis and inflammatory responses in macrophages and promotes atherosclerotic plaque stability.

Lisa Willemsen, Koen H M Prange, Annette E Neele, Cindy P A A van Roomen, Marion Gijbels, Guillermo R Griffith, Myrthe den Toom, Linda Beckers, Ricky Siebeler, Nathanael J Spann, Hung-Jen Chen, Laura A Bosmans, Andrej Gorbatenko, Suzanne van Wouw, Noam Zelcer, Heinz Jacobs, Fred van Leeuwen, Menno P J de Winther.

  Macrophages are critical immune cells in inflammatory diseases, and their differentiation and function are tightly regulated by histone modifications. H3K79 methylation is a histone modification associated with active gene expression, and DOT1L is the only histone methyltransferase for H3K79. Here we determine the role of DOT1L in macrophages by applying a selective DOT1L inhibitor in mouse and human macrophages and using myeloid-specific Dot1l-deficient mice. We found that DOT1L directly regulates macrophage function by controlling lipid biosynthesis gene programs including central lipid regulators like sterol regulatory element-binding proteins SREBP1 and SREBP2. DOT1L inhibition also leads to macrophage hyperactivation, which is associated with disrupted SREBP pathways. In vivo, myeloid Dot1l deficiency reduces atherosclerotic plaque stability and increases the activation of inflammatory plaque macrophages. Our data show that DOT1L is a crucial regulator of macrophage inflammatory responses and lipid regulatory pathways and suggest a high relevance of H3K79 methylation in inflammatory disease.

Keywords:  CP: Molecular biology; DOT1L; H3K79; atherosclerosis; epigenetics; immune system; inflammation; lipid metabolism; macrophage; methyltransferase

DOI:  https://doi.org/10.1016/j.celrep.2022.111703
Nat Cancer. 2022 Nov 21.

Metabolic symbiosis in pancreatic cancer.

Dylan Gerard Ryan, Christian Frezza.

DOI: https://doi.org/10.1038/s43018-022-00454-2
Nutrients. 2022 Nov 21. pii: 4932. [Epub ahead of print]14(22):

Molecular Mechanisms for Ketone Body Metabolism, Signaling Functions, and Therapeutic Potential in Cancer.

Chi Yeon Hwang, Wonchae Choe, Kyung-Sik Yoon, Joohun Ha, Sung Soo Kim, Eui-Ju Yeo, Insug Kang.

  The ketone bodies (KBs) β-hydroxybutyrate and acetoacetate are important alternative energy sources for glucose during nutrient deprivation. KBs synthesized by hepatic ketogenesis are catabolized to acetyl-CoA through ketolysis in extrahepatic tissues, followed by the tricarboxylic acid cycle and electron transport chain for ATP production. Ketogenesis and ketolysis are regulated by the key rate-limiting enzymes, 3-hydroxy-3-methylglutaryl-CoA synthase 2 and succinyl-CoA:3-oxoacid-CoA transferase, respectively. KBs participate in various cellular processes as signaling molecules. KBs bind to G protein-coupled receptors. The most abundant KB, β-hydroxybutyrate, regulates gene expression and other cellular functions by inducing post-translational modifications. KBs protect tissues by regulating inflammation and oxidative stress. Recently, interest in KBs has been increasing due to their potential for treatment of various diseases such as neurological and cardiovascular diseases and cancer. Cancer cells reprogram their metabolism to maintain rapid cell growth and proliferation. Dysregulation of KB metabolism also plays a role in tumorigenesis in various types of cancer. Targeting metabolic changes through dietary interventions, including fasting and ketogenic diets, has shown beneficial effects in cancer therapy. Here, we review current knowledge of the molecular mechanisms involved in the regulation of KB metabolism and cellular signaling functions, and the therapeutic potential of KBs and ketogenic diets in cancer.

Keywords:  cancer; inflammation; ketogenic diet; ketone bodies; oxidative stress; post-translational modifications; β-hydroxybutyrate

DOI:  https://doi.org/10.3390/nu14224932
EMBO J. 2022 Nov 21. e111268

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.

  Reprogramming of lipid metabolism is emerging as a hallmark of cancer, yet involvement of specific fatty acids (FA) species and related enzymes in tumorigenesis remains unclear. While previous studies have focused on involvement of long-chain fatty acids (LCFAs) including palmitate in cancer, little attention has been paid to the role of very long-chain fatty acids (VLCFAs). Here, we show that depletion of acetyl-CoA carboxylase (ACC1), a critical enzyme involved in the biosynthesis of fatty acids, inhibits both de novo synthesis and elongation of VLCFAs in human cancer cells. ACC1 depletion markedly reduces cellular VLCFA but only marginally influences LCFA levels, including palmitate that can be nutritionally available. Therefore, tumor growth is specifically susceptible to regulation of VLCFAs. We further demonstrate that VLCFA deficiency results in a significant decrease in ceramides as well as downstream glucosylceramides and sphingomyelins, which impairs mitochondrial morphology and renders cancer cells sensitive to oxidative stress and cell death. Taken together, our study highlights that VLCFAs are selectively required for cancer cell survival and reveals a potential strategy to suppress tumor growth.

Keywords:  acetyl-CoA carboxylase; fatty acid elongation; fatty acid synthase; mitochondria potential; very long-chain fatty acids

DOI:  https://doi.org/10.15252/embj.2022111268
Nat Cancer. 2022 Nov 21.

Differential integrated stress response and asparagine production drive symbiosis and therapy resistance of pancreatic adenocarcinoma cells.

Christopher J Halbrook, Galloway Thurston, Seth Boyer, Cecily Anaraki, Jennifer A Jiménez, Amy McCarthy, Nina G Steele, Samuel A Kerk, Hanna S Hong, Lin Lin, Fiona V Law, Catherine Felton, Lorenzo Scipioni, Peter Sajjakulnukit, Anthony Andren, Alica K Beutel, Rima Singh, Barbara S Nelson, Fran Van Den Bergh, Abigail S Krall, Peter J Mullen, Li Zhang, Sandeep Batra, Jennifer P Morton, Ben Z Stanger, Heather R Christofk, Michelle A Digman, Daniel A Beard, Andrea Viale, Ji Zhang, Howard C Crawford, Marina Pasca di Magliano, Claus Jorgensen, Costas A Lyssiotis.

The pancreatic tumor microenvironment drives deregulated nutrient availability. Accordingly, pancreatic cancer cells require metabolic adaptations to survive and proliferate. Pancreatic cancer subtypes have been characterized by transcriptional and functional differences, with subtypes reported to exist within the same tumor. However, it remains unclear if this diversity extends to metabolic programming. Here, using metabolomic profiling and functional interrogation of metabolic dependencies, we identify two distinct metabolic subclasses among neoplastic populations within individual human and mouse tumors. Furthermore, these populations are poised for metabolic cross-talk, and in examining this, we find an unexpected role for asparagine supporting proliferation during limited respiration. Constitutive GCN2 activation permits ATF4 signaling in one subtype, driving excess asparagine production. Asparagine release provides resistance during impaired respiration, enabling symbiosis. Functionally, availability of exogenous asparagine during limited respiration indirectly supports maintenance of aspartate pools, a rate-limiting biosynthetic precursor. Conversely, depletion of extracellular asparagine with PEG-asparaginase sensitizes tumors to mitochondrial targeting with phenformin.

DOI: https://doi.org/10.1038/s43018-022-00463-1
Nat Commun. 2022 Nov 25. 13(1): 7260

GPSM1 impairs metabolic homeostasis by controlling a pro-inflammatory pathway in macrophages.

Jing Yan, Yuemei Zhang, Hairong Yu, Yicen Zong, Daixi Wang, Jiangfei Zheng, Li Jin, Xiangtian Yu, Caizhi Liu, Yi Zhang, Feng Jiang, Rong Zhang, Xiangnan Fang, Ting Xu, Mingyu Li, Jianzhong Di, Yan Lu, Xinran Ma, Jian Zhang, Weiping Jia, Cheng Hu.

G-protein-signaling modulator 1 (GPSM1) exhibits strong genetic association with Type 2 diabetes (T2D) and Body Mass Index in population studies. However, how GPSM1 carries out such control and in which types of cells are poorly understood. Here, we demonstrate that myeloid GPSM1 promotes metabolic inflammation to accelerate T2D and obesity development. Mice with myeloid-specific GPSM1 ablation are protected against high fat diet-induced insulin resistance, glucose dysregulation, and liver steatosis via repression of adipose tissue pro-inflammatory states. Mechanistically, GPSM1 deficiency mainly promotes TNFAIP3 transcription via the Gαi3/cAMP/PKA/CREB axis, thus inhibiting TLR4-induced NF-κB signaling in macrophages. In addition, we identify a small-molecule compound, AN-465/42243987, which suppresses the pro-inflammatory phenotype by inhibiting GPSM1 function, which could make it a candidate for metabolic therapy. Furthermore, GPSM1 expression is upregulated in visceral fat of individuals with obesity and is correlated with clinical metabolic traits. Overall, our findings identify macrophage GPSM1 as a link between metabolic inflammation and systemic homeostasis.

DOI: https://doi.org/10.1038/s41467-022-34998-9
J Cell Biol. 2022 Dec 05. pii: e202210021. [Epub ahead of print]221(12):

Temporal and spatial metabolite dynamics impart control in adipogenesis.

Kristin Eckel-Mahan.

The process of adipogenesis is critical for forming new, healthy adipocytes that are capable of storing lipids. In this issue, Sánchez-Ramírez and Ung et al. (2022. J. Cell Biol.https://doi.org/10.1083/jcb.202111137) reveal a novel role for the metabolite nicotinamide adenine dinucleotide in controlling differentiation of mesenchymal stromal cells into adipocytes.

DOI: https://doi.org/10.1083/jcb.202210021
Open Biol. 2022 Nov;12(11): 220248

How neutrophil metabolism affects bacterial killing.

Juliana E Toller-Kawahisa, Luke A J O'Neill.

  Neutrophils are front line cells in immunity that quickly recognize and eliminate pathogens, relying mainly on glycolysis to exert their killing functions. Even though investigations into the influence of metabolic pathways in neutrophil function started in the 1930s, the knowledge of how neutrophils metabolically adapt during a bacterial infection remains poorly understood. In this review, we discuss the current knowledge about the metabolic regulation underlying neutrophils response to bacterial infection. Glycogen metabolism has been shown to be important for multiple neutrophil functions. The potential contribution of metabolic pathways other than glycolysis, such as mitochondrial metabolism, for neutrophil function has recently been explored, including fatty acid oxidation in neutrophil differentiation. Complex III in the mitochondria might also control glycolysis via glycerol-3-phosphate oxidation. Future studies should yield new insights into the role of metabolic change in the anti-bacterial response in neutrophils.

Keywords:  bacterial infection; glucose metabolism; glutamine metabolism; mitochondrial metabolism; neutrophil function

DOI:  https://doi.org/10.1098/rsob.220248
J Biol Chem. 2022 Nov 16. pii: S0021-9258(22)01151-6. [Epub ahead of print] 102708

Glucagon Changes Substrate Preference in Gluconeogenesis.

Huiting Xu, Yujue Wang, Hyokjoon Kwon, Ankit Shah, Katarzyna Kalemba, Xiaoyang Su, Ling He, Fredric E Wondisford.

Fasting hyperglycemia in diabetes mellitus is caused by unregulated glucagon secretion that activates gluconeogenesis (GNG) and increases the use of pyruvate, lactate, amino acids, and glycerol. Studies of GNG in hepatocytes, however, tend to test a limited number of substrates at non-physiologic concentrations. Therefore, we treated cultured primary hepatocytes with three identical substrate mixtures of pyruvate/lactate, glutamine, and glycerol at serum fasting concentrations, where a different U-13C or 2-13C labeled substrate was substituted in each mix. In the absence of glucagon stimulation, 80% of glucose produced in primary hepatocytes incorporated either one or two 13C-labeled glycerol molecules in a 1:1 ratio, reflecting the high overall activity of this pathway. In contrast, glucose produced from 13C-labeled pyruvate/lactate or glutamine rarely incorporated two labeled molecules. While glucagon increased glycerol and pyruvate/lactate contribution to glucose carbon by 1.6- and 1.8-fold, respectively, glutamine contribution to glucose carbon was increased 6.4-fold in primary hepatocytes. To account for substrate 13C carbon loss during metabolism, we also performed a metabolic flux analysis, which confirmed that the majority of glucose carbon produced by primary hepatocytes was from glycerol. In vivo studies using a PKA-activation mouse model that represents elevated glucagon activity confirmed that most circulating lactate carbons originated from glycerol, but very little glycerol was derived from lactate carbons, reflecting glycerol's importance as a carbon donor to GNG. Given diverse entry points for GNG substrates, hepatic glucagon action is unlikely to be due to a single mechanism.

DOI: https://doi.org/10.1016/j.jbc.2022.102708
Cell Rep. 2022 Nov 22. pii: S2211-1247(22)01565-0. [Epub ahead of print]41(8): 111691

Branched-chain amino acid catabolism breaks glutamine addiction to sustain hepatocellular carcinoma progression.

Dongdong Yang, Haiying Liu, Yongping Cai, Kangyang Lu, Xiuying Zhong, Songge Xing, Wei Song, Yaping Zhang, Ling Ye, Xia Zhu, Ting Wang, Pinggen Zhang, Shi-Ting Li, Jiaqian Feng, Weidong Jia, Huafeng Zhang, Ping Gao.

  Branched-chain amino acid (BCAA) catabolism is related to tumorigenesis. However, the underlying mechanism and specific contexts in which BCAAs affect tumor progression remain unclear. Here, we demonstrate that BCAA catabolism is activated in liver cancer cells without glutamine. Enhanced BCAA catabolism leads to BCAA-derived carbon and nitrogen flow toward nucleotide synthesis, stimulating cell-cycle progression and promoting cell survival. Mechanistically, O-GlcNAcylation increases under glutamine-deprivation conditions and stabilizes the PPM1K protein, leading to dephosphorylation of BCKDHA and enhanced decomposition of BCAAs. Dephosphorylation of BCKDHA and high expression of PPM1K promote tumorigenesis in vitro and in vivo and are closely related to the poor prognosis of clinical patients with hepatocellular carcinoma (HCC). Inhibition of BCAA and glutamine metabolism can further retard HCC growth in vivo. These results not only elucidate a mechanism by which BCAA catabolism affects tumorigenesis but also identify pBCKDHA and PPM1K as potential therapeutic targets and predictive biomarkers.

Keywords:  CP: Cancer; CP: Metabolism; HCC progression; O-GlcNAcylation; branch-chain amino acid; glutamine-depravation

DOI:  https://doi.org/10.1016/j.celrep.2022.111691
Sci Adv. 2022 Nov 25. 8(47): eabq1984

Macrophage acetyl-CoA carboxylase regulates acute inflammation through control of glucose and lipid metabolism.

Scott Yeudall, Clint M Upchurch, Philip V Seegren, Caitlin M Pavelec, Jan Greulich, Michael C Lemke, Thurl E Harris, Bimal N Desai, Kyle L Hoehn, Norbert Leitinger.

Acetyl-CoA carboxylase (ACC) regulates lipid synthesis; however, its role in inflammatory regulation in macrophages remains unclear. We generated mice that are deficient in both ACC isoforms in myeloid cells. ACC deficiency altered the lipidomic, transcriptomic, and bioenergetic profile of bone marrow-derived macrophages, resulting in a blunted response to proinflammatory stimulation. In response to lipopolysaccharide (LPS), ACC is required for the early metabolic switch to glycolysis and remodeling of the macrophage lipidome. ACC deficiency also resulted in impaired macrophage innate immune functions, including bacterial clearance. Myeloid-specific deletion or pharmacological inhibition of ACC in mice attenuated LPS-induced expression of proinflammatory cytokines interleukin-6 (IL-6) and IL-1β, while pharmacological inhibition of ACC increased susceptibility to bacterial peritonitis in wild-type mice. Together, we identify a critical role for ACC in metabolic regulation of the innate immune response in macrophages, and thus a clinically relevant, unexpected consequence of pharmacological ACC inhibition.

DOI: https://doi.org/10.1126/sciadv.abq1984
Nat Metab. 2022 Nov 24.

Metabolic control of innate lymphoid cells in health and disease.

Lei Zhou, Qingxia Lin, Gregory F Sonnenberg.

Innate lymphoid cells (ILCs) are a family of predominantly tissue-resident lymphocytes that critically orchestrate immunity, inflammation, tolerance and repair at barrier surfaces of the mammalian body. Heterogeneity among ILC subsets is comparable to that of adaptive CD4+ T helper cell counterparts, and emerging studies demonstrate that ILC biology is also dictated by cellular metabolism that adapts bioenergetic requirements during activation, proliferation or cytokine production. Accumulating evidence in mouse models and human samples indicates that ILCs exhibit profound roles in shaping states of metabolic health and disease. Here we summarize and discuss our current knowledge of the cell-intrinsic and cell-extrinsic metabolic factors controlling ILC responses, as well as highlight contributions of ILCs to organismal metabolism. It is expected that continued research in this area will advance our understanding of how to manipulate ILCs or their metabolism for therapeutic strategies that benefit human health.

DOI: https://doi.org/10.1038/s42255-022-00685-8
Adv Biol (Weinh). 2022 Nov 23. e2200233

Metabolic Recycling Enhances Proliferation in MYC-Transformed Lymphoma B Cells.

Cissy Zhang, Giang Hoang, Nabeel Attarwala, Arthur J L Cooper, Ryoichi Asaka, Anne Le.

  Relapses negatively impact cancer patient survival due to the tumorigenesis ability of surviving cancer cells post-therapy. Efforts are needed to better understand and combat this problem. This study hypothesized that dead cell debris post-radiation therapy creates an advantageous microenvironment rich in metabolic materials promoting the growth of remaining live cancer cells. In this study, live cancer cells are co-cultured with dead cancer cells eradicated by UV radiation to mimic a post-therapy environment. Isotopic labeling metabolomics is used to investigate the metabolic behavior of cancer cells grown in a post-radiation-therapy environment. It is found that post-UV-eradicated dead cancer cells serve as nutritional sources of "off-the-shelf" and precursor metabolites for surviving cancer cells. The surviving cancer cells then take up these metabolites, integrate and upregulate multiple vital metabolic processes, thereby significantly increasing growth in vitro and probably in vivo beyond their intrinsic fast-growing characteristics. Importantly, this active metabolite uptake behavior is only observed in oncogenic but not in non-oncogenic cells, presenting opportunities for therapeutic approaches to interrupt the active uptake process of oncogenic cells without affecting normal cells. The process by which living cancer cells re-use vital metabolites released by dead cancer cells post-therapy is coined in this study as "metabolic recycling" of oncogenic cells.

Keywords:  13C6-glucose labeling; MYC-transformed lymphoma B cells; cancer metabolism; glucose metabolism; mass spectrometry; metabolic recycling; metabolomics

DOI:  https://doi.org/10.1002/adbi.202200233
Nitric Oxide. 2022 Nov 19. pii: S1089-8603(22)00122-7. [Epub ahead of print]

Glutamine-dependent effects of nitric oxide on cancer cells subjected to hypoxia-reoxygenation.

Dianna Xing, Gloria A Benavides, Michelle S Johnson, Ran Tian, Stephen Barnes, Victor M Darley-Usmar.

  Limited O2 availability can decrease essential processes in energy metabolism. However, cancers have developed distinct metabolic adaptations to these conditions. For example, glutaminolysis can maintain energy metabolism and hypoxia signaling. Additionally, it has been observed that nitric oxide (NO) possesses concentration-dependent, biphasic effects in cancer. NO has potent anti-tumor effects through modulating events such as angiogenesis and metastasis at low physiological concentrations and inducing cell death at higher concentrations. In this study, Ewing Sarcoma cells (A-673), MIA PaCa, and SKBR3 cells were treated with DetaNONOate (DetaNO) in a model of hypoxia (1% O2) and reoxygenation (21% O2). All 3 cell types showed NO-dependent inhibition of cellular O2 consumption which was enhanced as O2-tension decreased. L-Gln depletion suppressed the mitochondrial response to decreasing O2 tension in all 3 cell types and resulted in inhibition of Complex I activity. In A-673 cells the O2 tension dependent change in mitochondrial O2 consumption and increase in glycolysis was dependent on the presence of L-Gln. The response to hypoxia and Complex I activity were restored by α-ketoglutarate. NO exposure resulted in the A-673 cells showing greater sensitivity to decreasing O2 tension. Under conditions of L-Gln depletion, NO restored HIF-1α levels and the mitochondrial response to O2 tension possibly through the increase of 2-hydroxyglutarate. NO also resulted in suppression of cellular bioenergetics and further inhibition of Complex I which was not rescued by α-ketoglutarate. Taken together these data suggest that NO modulates the mitochondrial response to O2 differentially in the absence and presence of L-Gln. These data suggest a combination of metabolic strategies targeting glutaminolysis and Complex I in cancer cells.

Keywords:  Cancer; Glutaminolysis; Mitochondria; Mitochondrial function; Nitric oxide

DOI:  https://doi.org/10.1016/j.niox.2022.11.003
EMBO Rep. 2022 Nov 23. e54006

Mitophagy bridges DNA sensing with metabolic adaption to expand lung cancer stem-like cells.

Zhen Liu, Shan Shan, Zixin Yuan, Fengying Wu, Ming Zheng, Ying Wang, Jun Gui, Wei Xu, Chunhong Wang, Tao Ren, Zhenke Wen.

  While previous studies have identified cancer stem-like cells (CSCs) as a crucial driver for chemoresistance and tumor recurrence, the underlying mechanisms for populating the CSC pool remain unclear. Here, we identify hypermitophagy as a feature of human lung CSCs, promoting metabolic adaption via the Notch1-AMPK axis to drive CSC expansion. Specifically, mitophagy is highly active in CSCs, resulting in increased mitochondrial DNA (mtDNA) content in the lysosome. Lysosomal mtDNA acts as an endogenous ligand for Toll-like receptor 9 (TLR9) that promotes Notch1 activity. Notch1 interacts with AMPK to drive lysosomal AMPK activation by inducing metabolic stress and LKB1 phosphorylation. This TLR9-Notch1-AMPK axis supports mitochondrial metabolism to fuel CSC expansion. In patient-derived xenograft chimeras, targeting mitophagy and TLR9-dependent Notch1-AMPK pathway restricts tumor growth and CSC expansion. Taken together, mitochondrial hemostasis is interlinked with innate immune sensing and Notch1-AMPK activity to increase the CSC pool of human lung cancer.

Keywords:  AMPK; Notch1; TLR9; cancer stem-like cell; mitophagy

DOI:  https://doi.org/10.15252/embr.202154006
Nat Commun. 2022 Nov 25. 13(1): 7272

Alveolar macrophage metabolic programming via a C-type lectin receptor protects against lipo-toxicity and cell death.

Michal Scur, Ahmad Bakur Mahmoud, Sayanti Dey, Farah Abdalbarri, Iona Stylianides, Daniel Medina-Luna, Gayani S Gamage, Aaron Woblistin, Alexa N M Wilson, Haggag S Zein, Ashley Stueck, Andrew Wight, Oscar A Aguilar, Francesca Di Cara, Brendon D Parsons, Mir Munir A Rahim, James R Carlyle, Andrew P Makrigiannis.

Alveolar macrophages (AM) hold lung homeostasis intact. In addition to the defense against inhaled pathogens and deleterious inflammation, AM also maintain pulmonary surfactant homeostasis, a vital lung function that prevents pulmonary alveolar proteinosis. Signals transmitted between AM and pneumocytes of the pulmonary niche coordinate these specialized functions. However, the mechanisms that guide the metabolic homeostasis of AM remain largely elusive. We show that the NK cell-associated receptor, NKR-P1B, is expressed by AM and is essential for metabolic programming. Nkrp1b-/- mice are vulnerable to pneumococcal infection due to an age-dependent collapse in the number of AM and the formation of lipid-laden AM. The AM of Nkrp1b-/- mice show increased uptake but defective metabolism of surfactant lipids. We identify a physical relay between AM and alveolar type-II pneumocytes that is dependent on pneumocyte Clr-g expression. These findings implicate the NKR-P1B:Clr-g signaling axis in AM-pneumocyte communication as being important for maintaining metabolism in AM.

DOI: https://doi.org/10.1038/s41467-022-34935-w
J Biol Chem. 2022 Nov 17. pii: S0021-9258(22)01160-7. [Epub ahead of print] 102717

The leucine-rich repeat (LRR) domain of NLRP3 is required for NLRP3 inflammasome activation in macrophages.

Yanhui Duan, Jihong Wang, Juan Cai, Nathan Kelley, Yuan He.

  The NLRP3 inflammasome is a critical component of innate immunity that defends the host from microbial infections. However, its aberrant activation contributes to the pathogenesis of several inflammatory diseases. Activation of the NLRP3 inflammasome induces the secretion of proinflammatory cytokines IL-1β and IL-18, and pyroptotic cell death. NLRP3 contains a leucine-rich repeat (LRR) domain at its C-terminus. Although posttranslational modifications in this LRR domain have been shown to regulate NLRP3 inflammasome activation, the role of the entire LRR domain in NLRP3 inflammasome activation remains controversial. Here, we generated mouse macrophages that express an endogenous NLRP3 mutant lacking the LRR domain. Deletion of the LRR domain diminished NLRP3 inflammasome activation in macrophages. Furthermore, using NLRP3-deficient macrophages that are reconstituted with NLRP3 mutants lacking the LRR domain, we found that deletion of the LRR domain inhibited NLRP3 inflammasome activation. Mechanistically, deletion of the LRR domain inhibited NLRP3 self-association, oligomerization, and interaction with the essential regulator NEK7. Our results demonstrate a critical role for the LRR domain in NLRP3 inflammasome activation.

Keywords:  Innate immunity; NLRP3; Nod-like receptor; inflammasome; inflammation

DOI:  https://doi.org/10.1016/j.jbc.2022.102717
Front Cardiovasc Med. 2022 ;9 994080

Role of miR-199a-5p in the post-transcriptional regulation of ABCA1 in response to hypoxia in peritoneal macrophages.

Juan Francisco Aranda, Ana Pérez-García, Marta Torrecilla-Parra, Mario Fernández-de Frutos, Yolanda Martín-Martín, Pedro A Mateos-Gómez, Virginia Pardo-Marqués, Rebeca Busto, Cristina M Ramírez.

  Hypoxia is a crucial factor contributing to maintenance of atherosclerotic lesions. The ability of ABCA1 to stimulate the efflux of cholesterol from cells in the periphery, particularly foam cells in atherosclerotic plaques, is an important anti-atherosclerotic mechanism. The posttranscriptional regulation by miRNAs represents a key regulatory mechanism of a number of signaling pathways involved in atherosclerosis. Previously, miR-199a-5p has been shown to be implicated in the endocytic and retrograde intracellular transport. Although the regulation of miR-199a-5p and ABCA1 by hypoxia has been already reported independently, the role of miR-199a-5p in macrophages and its possible role in atherogenic processes such us regulation of lipid homeostasis through ABCA1 has not been yet investigated. Here, we demonstrate that both ABCA1 and miR-199a-5p show an inverse regulation by hypoxia and Ac-LDL in primary macrophages. Moreover, we demonstrated that miR-199a-5p regulates ABCA1 mRNA and protein levels by directly binding to its 3'UTR. As a result, manipulation of cellular miR-199a-5p levels alters ABCA1 expression and cholesterol efflux in primary mouse macrophages. Taken together, these results indicate that the correlation between ABCA1-miR-199a-5p could be exploited to control macrophage cholesterol efflux during the onset of atherosclerosis, where cholesterol alterations and hypoxia play a pathogenic role.

Keywords:  ABCA1; atherosclerosis; cholesterol efflux; hypoxia; macrophage; miRNAs

DOI:  https://doi.org/10.3389/fcvm.2022.994080
Circulation. 2022 Nov 23.

Macrophage-Derived 25-Hydroxycholesterol Promotes Vascular Inflammation, Atherogenesis, and Lesion Remodeling.

Alberto Canfrán-Duque, Noemi Rotllan, Xinbo Zhang, Irene Andrés-Blasco, Bonne M Thompson, Jonathan Sun, Nathan L Price, Marta Fernández-Fuertes, Joseph W Fowler, Diego Gómez-Coronado, William C Sessa, Chiara Giannarelli, Robert J Schneider, George Tellides, Jeffrey G McDonald, Carlos Fernández-Hernando, Yajaira Suárez.

  BACKGROUND: Cross-talk between sterol metabolism and inflammatory pathways has been demonstrated to significantly affect the development of atherosclerosis. Cholesterol biosynthetic intermediates and derivatives are increasingly recognized as key immune regulators of macrophages in response to innate immune activation and lipid overloading. 25-Hydroxycholesterol (25-HC) is produced as an oxidation product of cholesterol by the enzyme cholesterol 25-hydroxylase (CH25H) and belongs to a family of bioactive cholesterol derivatives produced by cells in response to fluctuating cholesterol levels and immune activation. Despite the major role of 25-HC as a mediator of innate and adaptive immune responses, its contribution during the progression of atherosclerosis remains unclear.METHODS: The levels of 25-HC were analyzed by liquid chromatography-mass spectrometry, and the expression of CH25H in different macrophage populations of human or mouse atherosclerotic plaques, respectively. The effect of CH25H on atherosclerosis progression was analyzed by bone marrow adoptive transfer of cells from wild-type or Ch25h-/- mice to lethally irradiated Ldlr-/- mice, followed by a Western diet feeding for 12 weeks. Lipidomic, transcriptomic analysis and effects on macrophage function and signaling were analyzed in vitro from lipid-loaded macrophage isolated from Ldlr-/- or Ch25h-/-;Ldlr-/- mice. The contribution of secreted 25-HC to fibrous cap formation was analyzed using a smooth muscle cell lineage-tracing mouse model, Myh11ERT2CREmT/mG;Ldlr-/-, adoptively transferred with wild-type or Ch25h-/- mice bone marrow followed by 12 weeks of Western diet feeding.
RESULTS: We found that 25-HC accumulated in human coronary atherosclerotic lesions and that macrophage-derived 25-HC accelerated atherosclerosis progression, promoting plaque instability through autocrine and paracrine actions. 25-HC amplified the inflammatory response of lipid-loaded macrophages and inhibited the migration of smooth muscle cells within the plaque. 25-HC intensified inflammatory responses of lipid-laden macrophages by modifying the pool of accessible cholesterol in the plasma membrane, which altered Toll-like receptor 4 signaling, promoted nuclear factor-κB-mediated proinflammatory gene expression, and increased apoptosis susceptibility. These effects were independent of 25-HC-mediated modulation of liver X receptor or SREBP (sterol regulatory element-binding protein) transcriptional activity.
CONCLUSIONS: Production of 25-HC by activated macrophages amplifies their inflammatory phenotype, thus promoting atherogenesis.

Keywords:  25-hydroxycholesterol; atherosclerosis; inflammation; macrophages

DOI:  https://doi.org/10.1161/CIRCULATIONAHA.122.059062
JCI Insight. 2022 Nov 24. pii: e151819. [Epub ahead of print]

Gsk3β regulates the resolution of liver ischemia reperfusion injury via MerTK.

Hanwen Zhang, Ming Ni, Han Wang, Jing Zhang, Dan Jin, Ronald W Busuttil, Jerzy W Kupiec-Weglinski, Wei Li, Xuehao Wang, Yuan Zhai.

  Although glycogen synthase kinase β (Gsk3β) has been shown to regulate tissue inflammation, whether and how it regulates inflammation resolution vs. inflammation activation is unclear. In a murine liver partial warm ischemia/reperfusion injury (IRI) model, we found that Gsk3β inhibitory phosphorylation increased at both the early activation and late resolution stages of the disease. Myeloid Gsk3β deficiency not only alleviated liver injuries, but also facilitated the restoration of liver homeostasis. Depletion of Kupffer cells (KCs) prior to the onset of liver ischemia diminished the differences between the WT and Gsk3β KO mice in the activation of liver IRI. However, the resolution of liver IRI remained accelerated in the Gsk3β KO mice. In CD11b-DTR mice, Gsk3β deficient bone marrow-derived macrophages (BMMs) facilitated the resolution of liver IRI as compared with WT cells. Furthermore, Gsk3β deficiency promoted the reparative phenotype differentiation in vivo in liver infiltrating macrophages and in vitro in BMMs. Gsk3 pharmacological inhibition promoted the resolution of liver IRI in WT, but not myeloid MerTK deficient, mice. Thus, Gsk3β regulates liver IRI at both activation and resolution stages of the disease. Gsk3 inactivation enhances the pro-resolving function of liver infiltrating macrophages in MerTK-dependent manner.

Keywords:  Hepatology; Immunology; Innate immunity; Macrophages

DOI:  https://doi.org/10.1172/jci.insight.151819
Cell Death Dis. 2022 Nov 22. 13(11): 985

Manipulating PP2Acα-ASK-JNK signaling to favor apoptotic over necroptotic hepatocyte fate reduces the extent of necrosis and fibrosis upon acute liver injury.

Ke Lu, Si-Yu Shen, Ou-Yang Luo, Yue Lu, Tian-Shu Shi, Jing Wu, Qi Cheng, Hua-Jian Teng, Di Chen, Xiang Lu, Chao-Jun Li, Qing Jiang, Lei Fang, Bin Xue.

In the widely used Carbon tetrachloride (CCl4)-induced acute liver injury (ALI) mouse model, hepatocytes are known to die from programmed cell death (PCD) processes including apoptosis and necroptosis. Both in vivo and in vitro experiments showed that CCl4 treatment could induce both apoptosis and necroptosis. Treatment of mice with the apoptosis inducer SMAC mimetic reduced necroptosis, led to less pronounced liver damage, and improved overall liver function. By LC-MS/MS, we found that PP2Acα expression was increased in ALI mice liver, and we confirmed its high expression in subacute hepatitis patients. We observed that ALI severity (including aggravated fibrogenesis) was significantly alleviated in hepatocyte-specific PP2Acα conditional knockout (PP2Acα cKO) mice. Furthermore, the relative extent of apoptosis over necroptosis was increased in the PP2Acα cKO ALI mice. Pursuing the idea that biasing the type of PCD towards apoptosis may reduce liver damage, we found that treatment of PP2Acα cKO ALI mice with the apoptosis inhibitor z-Vad-fmk increased the extent of necroptosis and caused severer damage. Mechanistically, disruption of PP2Acα prevents the dephosphorylation of pASK1(Ser967), thereby preventing the sustained activation of JNK. Inhibition of PP2Acα prevents CCl4-induced liver injury and fibrogenesis by disrupting ASK/JNK pathway mediated PCD signaling, ultimately improving liver function by biasing hepatocytes towards an apoptotic rather than necroptotic cell fate. Thus, targeting PP2A and/or ASK1 to favor apoptotic over necroptotic hepatocyte fate may represent an attractive therapeutic strategy for treating ALI.

DOI: https://doi.org/10.1038/s41419-022-05353-z
Am J Physiol Regul Integr Comp Physiol. 2022 Nov 21.

G-Protein-Coupled Lactate Receptor GPR81 Control of VMNvl Glucoregulatory Neurotransmitter and 5'-AMP-Activated Protein Kinase Expression.

Sagor Chandra Roy, Prabhat R Napit, Madhu Babu Pasula, Khaggeswar Bheemanapally, Karen P Briski.

  Astrocytes store glycogen as energy and promote neuro-metabolic stability through supply of oxidizable L-lactate. Whether lactate regulates ventromedial hypothalamic nucleus (VMN) glucostatic function as a metabolic volume transmitter is unknown. Current research investigated whether G-protein-coupled lactate receptor GPR81 controls astrocyte glycogen metabolism and glucose-regulatory neurotransmission in the ventrolateral VMN (VMNvl), where glucose-regulatory neurons reside. Female rats were pretreated by intra-VMN GPR81 or scramble siRNA infusion before insulin or vehicle injection. VMNvl cell or tissue samples were acquired by laser-catapult- or micropunch microdissection for Western blot protein or uHPLC-electrospray ionization-mass spectrometric glycogen analyses. Data show that GPR81 regulates eu- and/or hypoglycemic patterns of VMNvl astrocyte glycogen metabolic enzyme and 5'-AMP-activated protein kinase (AMPK) protein expression according to VMNvl segment. GPR81 inhibits baseline rostral and caudal VMNvl glycogen accumulation, but mediates glycogen breakdown in the former site during hypoglycemia. During euglycemia, GPR81 suppresses the transmitter marker neuronal nitric oxide synthase (nNOS) in rostral and caudal VMNvl nitrergic neurons, but stimulates (rostral VMNvl) or inhibits (caudal VMNvl) GABAergic neuron glutamate decarboxylase65/67 (GAD)protein. During hypoglycemia, GPR81 regulates AMPK activation in nitrergic and GABAergic neurons located in the rostral, but not caudal. VMNvl. VMN GPR81 knockdown amplified hypoglycemic hypercorticosteronemia, but not hyperglucagonemia. Results provide novel proof that VMNvl astrocyte and glucose-regulatory neurons express GPR81 protein. Data identify neuroanatomical subpopulations of VMNvl astrocytes and glucose-regulatory neurons that exhibit differential reactivity to GPR81 input. Heterogeneous GPR81 effects during eu- versus hypoglycemia infer that energy state may affect cellular sensitivity to or post-receptor processing of lactate transmitter signaling.

Keywords:  AMPK; GPR81; glycogen; neuronal nitric oxide synthase; ventrolateral ventromedial hypothalamic nucleus

DOI:  https://doi.org/10.1152/ajpregu.00100.2022
Biochem Biophys Res Commun. 2022 Nov 11. pii: S0006-291X(22)01566-2. [Epub ahead of print]637 218-223

Directional regulation of cytosolic PEPCK catalysis is mediated by competitive binding of anions.

Sarah A E Barwell, Ramona Duman, Armin Wagner, Todd Holyoak.

Phosphoenolpyruvate carboxykinase (PEPCK) is a well-characterized enzyme involved in primary glucose metabolism, responsible for catalyzing one of the key steps of gluconeogenesis. It is well demonstrated that PEPCK can efficiently catalyze the reversible interconversion of oxaloacetic acid (OAA) to phosphoenolpyruvate (PEP) in vitro, but the enzyme is typically ascribed a metabolic role that requires preferential catalysis in the direction of PEP synthesis in vivo. Here we present structural and functional data that demonstrate the preferential synthesis of PEP from OAA catalyzed by PEPCK in vivo is facilitated by anion-mediated enzyme inhibition that reduces enzyme activity more significantly in the direction of OAA synthesis than in the direction of PEP synthesis. From our studies we conclude that the specific binding of small, ubiquitous anions like chloride, present in millimolar concentrations under normal cellular conditions allows for metabolic control by restricting PEPCK to function in the direction of PEP synthesis.

DOI: https://doi.org/10.1016/j.bbrc.2022.11.025
J Nutr Biochem. 2022 Nov 17. pii: S0955-2863(22)00292-3. [Epub ahead of print] 109224

Fructose Impairs Fat Oxidation: Implications for the Mechanism of Western diet-induced NAFLD.

Mustafa Kutlu Inci, Se-Hyung Park, Robert N Helsley, Suzanna L Attia, Samir Softic.

  Increased fructose intake from sugar-sweetened beverages and highly processed sweets is a well-recognized risk factor for the development of obesity and its complications. Fructose strongly supports lipogenesis on a normal chow diet by providing both, a substrate for lipid synthesis and activation of lipogenic transcription factors. However, the negative health consequences of dietary sugar are best observed with the concomitant intake of a HFD. Indeed, the most commonly used obesogenic research diets, such as "Western diet", contain both fructose and a high amount of fat. In spite of its common use, how the combined intake of fructose and fat synergistically supports development of metabolic complications is not fully elucidated. Here we present the preponderance of evidence that fructose consumption decreases oxidation of dietary fat in human and animal studies. We provide a detailed review of the mitochondrial β-oxidation pathway. Fructose affects hepatic activation of fatty acyl-CoAs, decreases acylcarnitine production and impairs the carnitine shuttle. Mechanistically, fructose suppresses transcriptional activity of PPARα and its target CPT1α, the rate limiting enzyme of acylcarnitine production. These effects of fructose may be, in part, mediated by protein acetylation. Acetylation of PGC1α, a co-activator of PPARα and acetylation of CPT1α, in part, account for fructose-impaired acylcarnitine production. Interestingly, metabolic effects of fructose in the liver can be largely overcome by carnitine supplementation. In summary, fructose decreases oxidation of dietary fat in the liver, in part, by impairing acylcarnitine production, offering one explanation for the synergistic effects of these nutrients on the development of metabolic complications, such as NAFLD.

Keywords:  Western diet; fatty acid oxidation (FAO); fructose; non-alcoholic fatty liver disease (NAFLD); sugar

DOI:  https://doi.org/10.1016/j.jnutbio.2022.109224
JCI Insight. 2022 Nov 22. pii: e153740. [Epub ahead of print]

HGFAC is a ChREBP regulated hepatokine that enhances glucose and lipid homeostasis.

Ashot Sargsyan, Ludivine Doridot, Sarah Anissa Hannou, Wenxin Tong, Harini Srinivasan, Rachael Ivison, Ruby Monn, Henry H Kou, Jonathan M Haldeman, Michelle Arlotto, Phillip J White, Paul A Grimsrud, Inna Astapova, Linus T-Y Tsai, Mark A Herman.

  Carbohydrate Responsive Element-Binding Protein (ChREBP) is a carbohydrate sensing transcription factor that regulates both adaptive and maladaptive genomic responses in coordination of systemic fuel homeostasis. Genetic variants in the ChREBP locus associate with diverse metabolic traits in humans, including circulating lipids. To identify novel ChREBP-regulated hepatokines that contribute to its systemic metabolic effects, we integrated ChREBP ChIP-seq analysis in mouse liver with human genetic and genomic data for lipid traits and identified Hepatocyte Growth Factor Activator (HGFAC) as a promising ChREBP-regulated candidate in mice and humans. HGFAC is a protease that activates the pleiotropic hormone Hepatocyte Growth Factor (HGF). We demonstrate that HGFAC KO mice have phenotypes concordant with putative loss-of-function variants in human HGFAC. Moreover, in gain- and loss-of-function genetic mouse models, we demonstrate that HGFAC enhances lipid and glucose homeostasis, which may be mediated in part through actions to activate hepatic PPARγ activity. Together, our studies show that ChREBP mediates an adaptive response to overnutrition via activation of HGFAC in the liver to preserve glucose and lipid homeostasis.

Keywords:  Carbohydrate metabolism; Glucose metabolism; Growth factors; Metabolism

DOI:  https://doi.org/10.1172/jci.insight.153740
EMBO Rep. 2022 Nov 21. e54689

Sphingosine-1-phosphate controls endothelial sphingolipid homeostasis via ORMDL.

Linda Sasset, Kamrul H Chowdhury, Onorina L Manzo, Luisa Rubinelli, Csaba Konrad, J Alan Maschek, Giovanni Manfredi, William L Holland, Annarita Di Lorenzo.

  Disruption of sphingolipid homeostasis and signaling has been implicated in diabetes, cancer, cardiometabolic, and neurodegenerative disorders. Yet, mechanisms governing cellular sensing and regulation of sphingolipid homeostasis remain largely unknown. In yeast, serine palmitoyltransferase, catalyzing the first and rate-limiting step of sphingolipid de novo biosynthesis, is negatively regulated by Orm1 and 2. Lowering sphingolipids triggers Orms phosphorylation, upregulation of serine palmitoyltransferase activity and sphingolipid de novo biosynthesis. However, mammalian orthologs ORMDLs lack the N-terminus hosting the phosphosites. Thus, which sphingolipid(s) are sensed by the cells, and mechanisms of homeostasis remain largely unknown. Here, we identify sphingosine-1-phosphate (S1P) as key sphingolipid sensed by cells via S1PRs to maintain homeostasis. The increase in S1P-S1PR signaling stabilizes ORMDLs, restraining SPT activity. Mechanistically, the hydroxylation of ORMDLs at Pro137 allows a constitutive degradation of ORMDLs via ubiquitin-proteasome pathway, preserving SPT activity. Disrupting S1PR/ORMDL axis results in ceramide accrual, mitochondrial dysfunction, impaired signal transduction, all underlying endothelial dysfunction, early event in the onset of cardio- and cerebrovascular diseases. Our discovery may provide the molecular basis for therapeutic intervention restoring sphingolipid homeostasis.

Keywords:  ORMDL; ceramide; endothelial dysfunction; serine palmitoyltransferase; sphingolipid

DOI:  https://doi.org/10.15252/embr.202254689
Signal Transduct Target Ther. 2022 Nov 23. 7(1): 378

Copper homeostasis and cuproptosis in health and disease.

Liyun Chen, Junxia Min, Fudi Wang.

As an essential micronutrient, copper is required for a wide range of physiological processes in virtually all cell types. Because the accumulation of intracellular copper can induce oxidative stress and perturbing cellular function, copper homeostasis is tightly regulated. Recent studies identified a novel copper-dependent form of cell death called cuproptosis, which is distinct from all other known pathways underlying cell death. Cuproptosis occurs via copper binding to lipoylated enzymes in the tricarboxylic acid (TCA) cycle, which leads to subsequent protein aggregation, proteotoxic stress, and ultimately cell death. Here, we summarize our current knowledge regarding copper metabolism, copper-related disease, the characteristics of cuproptosis, and the mechanisms that regulate cuproptosis. In addition, we discuss the implications of cuproptosis in the pathogenesis of various disease conditions, including Wilson's disease, neurodegenerative diseases, and cancer, and we discuss the therapeutic potential of targeting cuproptosis.

DOI: https://doi.org/10.1038/s41392-022-01229-y
Metabolites. 2022 Nov 19. pii: 1142. [Epub ahead of print]12(11):

Integration of Liver Glycogen and Triglyceride NMR Isotopomer Analyses Provides a Comprehensive Coverage of Hepatic Glucose and Fructose Metabolism.

Ivan Viegas, Giada Di Nunzio, Getachew D Belew, Alejandra N Torres, João G Silva, Luis Perpétuo, Cristina Barosa, Ludgero C Tavares, John G Jones.

  Dietary glucose and fructose are both efficiently assimilated by the liver but a comprehensive measurement of this process starting from their conversion to sugar phosphates, involvement of the pentose phosphate pathway (PPP), and conversion to glycogen and lipid storage products, remains incomplete. Mice were fed a chow diet supplemented with 35 g/100 mL drinking water of a 55/45 fructose/glucose mixture for 18 weeks. On the final night, the sugar mixture was enriched with either [U-13C]glucose or [U-13C]fructose, and deuterated water (2H2O) was also administered. 13C-isotopomers representing newly synthesized hepatic glucose-6-phosphate (glucose-6-P), glycerol-3-phosphate, and lipogenic acetyl-CoA were quantified by 2H and 13C NMR analysis of post-mortem liver glycogen and triglyceride. These data were applied to a metabolic model covering glucose-6-P, PPP, triose-P, and de novo lipogenesis (DNL) fluxes. The glucose supplement was converted to glucose-6-P via the direct pathway, while the fructose supplement was metabolized by the liver to gluconeogenic triose-P via fructokinase-aldolase-triokinase. Glucose-6-P from all carbohydrate sources accounted for 40-60% of lipogenic acetyl-CoA and 10-12% was oxidized by the pentose phosphate pathway (PPP). The yield of NADPH from PPP flux accounted for a minority (~30%) of the total DNL requirement. In conclusion, this approach integrates measurements of glucose-6-P, PPP, and DNL fluxes to provide a holistic and informative assessment of hepatic glucose and fructose metabolism.

Keywords:  13C NMR; acetyl-CoA; lipogenesis; pentose phosphate pathway; triose phosphates

DOI:  https://doi.org/10.3390/metabo12111142
Sci Adv. 2022 Nov 25. 8(47): eabq3363

Control of Gαq signaling dynamics and GPCR cross-talk by GRKs.

Guoqing Xiang, Amanda Acosta-Ruiz, Arthur Radoux-Mergault, Melanie Kristt, Jihye Kim, Jared D Moon, Johannes Broichhagen, Asuka Inoue, Francis S Lee, Miriam Stoeber, Jeremy S Dittman, Joshua Levitz.

Numerous processes contribute to the regulation of G protein-coupled receptors (GPCRs), but relatively little is known about rapid mechanisms that control signaling on the seconds time scale or regulate cross-talk between receptors. Here, we reveal that the ability of some GPCR kinases (GRKs) to bind Gαq both drives acute signaling desensitization and regulates functional interactions between GPCRs. GRK2/3-mediated acute desensitization occurs within seconds, is rapidly reversible, and can occur upon local, subcellular activation. This rapid desensitization is kinase independent, insensitive to pharmacological inhibition, and generalizable across receptor families and effectors. We also find that the ability of GRK2 to bind G proteins also enables it to regulate the extent and timing of Gαq-dependent signaling cross-talk between GPCRs. Last, we find that G protein/GRK2 interactions enable a novel form of GPCR trafficking cross-talk. Together, this work reveals potent forms of Gαq-dependent GPCR regulation with wide-ranging pharmacological and physiological implications.

DOI: https://doi.org/10.1126/sciadv.abq3363
Cell Death Dis. 2022 Nov 23. 13(11): 988

COX7A1 enhances the sensitivity of human NSCLC cells to cystine deprivation-induced ferroptosis via regulating mitochondrial metabolism.

Yetong Feng, Jiayi Xu, Mengjiao Shi, Rongrong Liu, Lei Zhao, Xin Chen, Miaomiao Li, Yaping Zhao, Jiahui Chen, Wenjing Du, Pengfei Liu.

COX7A1, a subunit of cytochrome c oxidase, holds an important position in the super-assembly which integrates into multi-unit heteromeric complexes peripherally in the mitochondrial electron transport chain (ETC). Recently, some studies indicated the significant potential of COX7A1 in cancer metabolism and therapy. However, the underlying metabolic process and therapy mechanism remain unclear. In this study, COX7A1-overexpressed cell line was established via lentivirus transduction. The relationship between COX7A1 and ferroptosis, a novel form of cell death driven by iron-dependent lipid peroxidation, was further analyzed in different human non-small-cell lung carcinoma (NSCLC) cells respectively. Our results showed that COX7A1 increased the sensitivity of NSCLC cells to the ferroptosis induced by cysteine deprivation via enhancing the tricarboxylic acid (TCA) cycle and the activity of complex IV in mitochondrial ETC. Meanwhile, COX7A1 suppressed mitochondrial dynamics as well as mitochondrial biogenesis and mitophagy through blocking autophagic flux. The autophagy activator, rapamycin, relieved the autophagic blockage and further strengthened the sensitivity to cysteine deprivation-induced ferroptosis of NSCLC cells in vitro and in vivo. Taken together, our data indicate the close association of COX7A1 with cysteine deprivation-induced ferroptosis, and provide a novel insight into the therapy mode against human NSCLC.

DOI: https://doi.org/10.1038/s41419-022-05430-3
Elife. 2022 Nov 21. pii: e82244. [Epub ahead of print]11

SRSF6 balances mitochondrial-driven innate immune outcomes through alternative splicing of BAX.

Allison R Wagner, Chi G Weindel, Kelsi O West, Haley M Scott, Robert O Watson, Kristin L Patrick.

  To mount a protective response to infection while preventing hyperinflammation, gene expression in innate immune cells must be tightly regulated. Despite the importance of pre-mRNA splicing in shaping the proteome, its role in balancing immune outcomes remains understudied. Transcriptomic analysis of murine macrophage cell lines identified Serine/Arginine Rich Splicing factor 6 (SRSF6) as a gatekeeper of mitochondrial homeostasis. SRSF6-dependent orchestration of mitochondrial health is directed in large part by alternative splicing of the pro-apoptosis pore-forming protein BAX. Loss of SRSF6 promotes accumulation of BAX-k, a variant that sensitizes macrophages to undergo cell death and triggers upregulation of interferon stimulated genes through cGAS sensing of cytosolic mitochondrial DNA. Upon pathogen sensing, macrophages regulate SRSF6 expression to control the liberation of immunogenic mtDNA and adjust the threshold for entry into programmed cell death. This work defines BAX alternative splicing by SRSF6 as a critical node not only in mitochondrial homeostasis, but also in the macrophage's response to pathogens.

Keywords:  chromosomes; gene expression; immunology; inflammation; mouse

DOI:  https://doi.org/10.7554/eLife.82244
Sci Adv. 2022 Nov 25. 8(47): eabo4116

A deubiquitination module essential for Treg fitness in the tumor microenvironment.

Elena Montauti, Samuel E Weinberg, Peng Chu, Shuvam Chaudhuri, Nikita L Mani, Radhika Iyer, Yuanzhang Zhou, Yusi Zhang, Changhong Liu, Chen Xin, Shana Gregory, Juncheng Wei, Yana Zhang, Wantao Chen, Zhaolin Sun, Ming Yan, Deyu Fang.

The tumor microenvironment (TME) enhances regulatory T (Treg) cell stability and immunosuppressive functions through up-regulation of lineage transcription factor Foxp3, a phenomenon known as Treg fitness or adaptation. Here, we characterize previously unknown TME-specific cellular and molecular mechanisms underlying Treg fitness. We demonstrate that TME-specific stressors including transforming growth factor-β (TGF-β), hypoxia, and nutrient deprivation selectively induce two Foxp3-specific deubiquitinases, ubiquitin-specific peptidase 22 (Usp22) and Usp21, by regulating TGF-β, HIF, and mTOR signaling, respectively, to maintain Treg fitness. Simultaneous deletion of both USPs in Treg cells largely diminishes TME-induced Foxp3 up-regulation, alters Treg metabolic signatures, impairs Treg-suppressive function, and alleviates Treg suppression on cytotoxic CD8+ T cells. Furthermore, we developed the first Usp22-specific small-molecule inhibitor, which dramatically reduced intratumoral Treg Foxp3 expression and consequently enhanced antitumor immunity. Our findings unveil previously unappreciated mechanisms underlying Treg fitness and identify Usp22 as an antitumor therapeutic target that inhibits Treg adaptability in the TME.

DOI: https://doi.org/10.1126/sciadv.abo4116
FEBS Open Bio. 2022 Nov 23.

Cyclic stretch regulates immune responses via tank-binding kinase 1 expression in macrophages.

Anna Nakagawa, Sumio Hayakawa, Yinglan Cheng, Azusa Honda, Ryo Yuzawa, Rei Ogawa, Yumiko Oishi.

  Macrophages distributed in tissues throughout the body contribute to homeostasis. In the inflammatory state, macrophages undergo mechanical stress that regulates the signal transduction of immune responses and various cellular functions. However, the effects of the inflammatory response on macrophages under physiological cyclic stretch are unclear. We found that physiological cyclic stretch suppresses inflammatory cytokine expression in macrophages by regulating NF-κB activity. NF-κB phosphorylation at Ser536 in macrophages was inhibited, suggesting that tank-binding kinase (TBK1) regulates NF-κB activity during physiological stress. Moreover, TBK1 expression was suppressed by physiological stretch, and TBK1 knockdown by siRNA induced the suppression of NF-κB phosphorylation at Ser536. In conclusion, physiological stretch triggers suppression of a TBK1-dependent excessive inflammatory response, which is necessary to maintain tissue homeostasis.

Keywords:  NF-κB phosphorylation; cyclic stretch; inflammatory cytokines; macrophage; tank-binding kinase

DOI:  https://doi.org/10.1002/2211-5463.13526
Proc Natl Acad Sci U S A. 2022 Nov 29. 119(48): e2119824119

The phosphorylation of AMPKβ1 is critical for increasing autophagy and maintaining mitochondrial homeostasis in response to fatty acids.

Eric M Desjardins, Brennan K Smith, Emily A Day, Serge Ducommun, Matthew J Sanders, Joshua P Nederveen, Rebecca J Ford, Stephen L Pinkosky, Logan K Townsend, Robert M Gutgesell, Rachel Lu, Kei Sakamoto, Gregory R Steinberg.

  Fatty acids are vital for the survival of eukaryotes, but when present in excess can have deleterious consequences. The AMP-activated protein kinase (AMPK) is an important regulator of multiple branches of metabolism. Studies in purified enzyme preparations and cultured cells have shown that AMPK is allosterically activated by small molecules as well as fatty acyl-CoAs through a mechanism involving Ser108 within the regulatory AMPK β1 isoform. However, the in vivo physiological significance of this residue has not been evaluated. In the current study, we generated mice with a targeted germline knock-in (KI) mutation of AMPKβ1 Ser108 to Ala (S108A-KI), which renders the site phospho-deficient. S108A-KI mice had reduced AMPK activity (50 to 75%) in the liver but not in the skeletal muscle. On a chow diet, S108A-KI mice had impairments in exogenous lipid-induced fatty acid oxidation. Studies in mice fed a high-fat diet found that S108A-KI mice had a tendency for greater glucose intolerance and elevated liver triglycerides. Consistent with increased liver triglycerides, livers of S108A-KI mice had reductions in mitochondrial content and respiration that were accompanied by enlarged mitochondria, suggestive of impairments in mitophagy. Subsequent studies in primary hepatocytes found that S108A-KI mice had reductions in palmitate- stimulated Cpt1a and Ppargc1a mRNA, ULK1 phosphorylation and autophagic/mitophagic flux. These data demonstrate an important physiological role of AMPKβ1 Ser108 phosphorylation in promoting fatty acid oxidation, mitochondrial biogenesis and autophagy under conditions of high lipid availability. As both ketogenic diets and intermittent fasting increase circulating free fatty acid levels, AMPK activity, mitochondrial biogenesis, and mitophagy, these data suggest a potential unifying mechanism which may be important in mediating these effects.

Keywords:  AMPK; NAFLD; autophagy; fat oxidation; mitochondria

DOI:  https://doi.org/10.1073/pnas.2119824119
Elife. 2022 Nov 22. pii: e83743. [Epub ahead of print]11

Structure of human phagocyte NADPH oxidase in the resting state.

Rui Liu, Kangcheng Song, Jing-Xiang Wu, Xiao-Peng Geng, Liming Zheng, Xiaoyin Gao, Hailin Peng, Lei Chen.

  Phagocyte oxidase plays an essential role in the first line of host defense against pathogens. It oxidizes intracellular NADPH to reduce extracellular oxygen to produce superoxide anions that participate in pathogen killing. The resting phagocyte oxidase is a heterodimeric complex formed by two transmembrane proteins NOX2 and p22. Despite the physiological importance of this complex, its structure remains elusive. Here we reported the cryo-EM structure of the functional human NOX2-p22 complex in nanodisc in the resting state. NOX2 shows a canonical 6-TM architecture of NOX and p22 has four transmembrane helices. M3, M4, and M5 of NOX2 and M1 and M4 helices of p22 are involved in the hetero-dimer formation. DH domain of NOX2 in the resting state is not optimally docked onto the transmembrane domain (TMD), leading to inefficient electron transfer and NADPH binding. Structural analysis suggests that the cytosolic factors might activate the NOX2-p22 complex by stabilizing the dehydrogenase domain (DH) in a productive docked conformation.

Keywords:  human; immunology; inflammation; molecular biophysics; structural biology

DOI:  https://doi.org/10.7554/eLife.83743
Redox Biol. 2022 Nov 12. pii: S2213-2317(22)00310-X. [Epub ahead of print]58 102538

Multi-omics profiling reveals Chitinase-3-like protein 1 as a key mediator in the crosstalk between sarcopenia and liver cancer.

Di Lu, Zuyuan Lin, Rui Wang, Zun Chen, Jianyong Zhuo, Li Xu, Linhui Pan, Huihui Li, Xinyu Yang, Chiyu He, Wei Shen, Modan Yang, Huigang Li, Hao Chen, Winyen Tan, Xuyong Wei, Shusen Zheng, Xiao Xu.

  Sarcopenia is prevalent in patients with hepatocellular carcinoma (HCC), and can adversely affect their outcomes. This study aims to explore the key mechanisms in the crosstalk between sarcopenia and HCC based on multi-omics profiling. A total of 136 male patients with HCC were enrolled. Sarcopenia was an independent risk factor for poor outcomes after liver transplantation (p < 0.05). Inflammatory cytokine and metabolomic profiling on these patients identified elevated plasma sTNF-R1/CHI3L1 and dysregulated lipid metabolism as related to sarcopenia and tumor recurrence risk concurrently (p < 0.05). Integrated analysis revealed close relationship between CHI3L1 and fatty acid metabolism. In mouse cachectic models by intraperitoneal injection of H22 cells, CHI3L1 was significantly elevated in the atrophic muscle tissue, as well as in circulation. In-vitro, CHI3L1 was up-regulated in muscle cells to protect itself from inflammatory damage through TNF-α/TNF-R1 signaling. CHI3L1 secreted by the muscle cells promoted the invasion of co-cultured HCC cells. Tumor tissue transcriptome data for 73 out of the 136 patients revealed that CHI3L1 may regulate fatty acid metabolism and oxidative stress. In vitro, CHI3L1 caused ROS and lipid accumulation. Targeted lipid profiling further proved that CHI3L1 was able to activate arachidonic acid metabolism, leading to lipid peroxide (LPO) accumulation. Meanwhile, LPO inhibition could compromise the remarkable pro-cancerous effects of CHI3L1. In conclusion, sarcopenia adversely affects the outcomes of liver transplantation for HCC. In sarcopenic patients, CHI3L1 was up-regulated and secreted by the skeletal muscle to protect itself through TNF-α/TNF-R1 signaling, which, in turn, can promote HCC tumor progression by inducing LPO accumulation.

Keywords:  CHI3L1; Hepatocellular carcinoma; Lipid peroxide; Metabolomic; Sarcopenia

DOI:  https://doi.org/10.1016/j.redox.2022.102538
FEBS Lett. 2022 Nov 23.

Mitochondrial ferritin expression in human macrophages is facilitated by thrombin-mediated cleavage under hypoxia.

Dominik C Fuhrmann, Sabrina Becker, Bernhard Brüne.

  Ferritins are iron storage proteins, which maintain cellular iron homeostasis. Among these proteins, the ferritin heavy chain is well characterized, but the regulatory principles of mitochondrial ferritin (FTMT) remain elusive. FTMT appears to be cleaved from a 27 kDa to a 22 kDa form. In human macrophages, FTMT increased under hypoxia in a hypoxia-inducible factor 2-dependent manner. Occurrence of FTMT resulted from cleavage by thrombin, which was supplied by serum. Inhibition of thrombin as well as serum removal decreased FTMT, while supplementation of thrombin under serum-deprived conditions restored its expression. Besides hypoxia, thrombin facilitated FTMT expression after treatment with the ferroptosis inducer RSL3 and the pro-inflammatory stimulus lipopolysaccharide. This study provides insights into the regulation of FTMT under hypoxia and identifies thrombin as a FTMT maturation-associated peptidase.

Keywords:  RSL3; Thrombin; ferroptosis; hypoxia inducible factor; oxidative stress

DOI:  https://doi.org/10.1002/1873-3468.14545
J Biol Chem. 2022 Nov 21. pii: S0021-9258(22)01176-0. [Epub ahead of print] 102733

Hydroxylation site-specific and production-dependent effects of endogenous oxysterols on cholesterol homeostasis: Implications for SREBP-2 and LXR.

Hodaka Saito, Wakana Tachiura, Mizuki Nishimura, Makoto Shimizu, Ryuichiro Sato, Yoshio Yamauchi.

  The cholesterol metabolites oxysterols play central roles in cholesterol feedback control. They modulate the activity of two master transcription factors that control cholesterol homeostatic responses, sterol regulatory element-binding protein-2 (SREBP-2) and liver X receptor (LXR). Although the role of exogenous oxysterols in regulating these transcription factors has been well established, whether endogenously synthesized oxysterols similarly control both SREBP-2 and LXR remains poorly explored. Here, we carefully validate the role of oxysterols enzymatically synthesized within cells in cholesterol homeostatic responses. We first show that SREBP-2 responds more sensitively to exogenous oxysterols than LXR in CHO cells and rat primary hepatocytes. We then show that 25-hydroxycholesterol (25-HC), 27-HC, and 24S-HC endogenously synthesized by CH25H, CYP27A1, and CYP46A1, respectively, suppress SREBP-2 activity at different degrees by stabilizing Insig proteins, whereas 7α-HC has little impact on SREBP-2. These results demonstrate the role of site-specific hydroxylation of endogenous oxysterols. In contrast, the expression of CH25H, CYP46A1, CYP27A1, or CYP7A1 fails to induce LXR target gene expression. We also show the 25-HC production-dependent suppression of SREBP-2 using a tetracycline-inducible CH25H expression system. To induce 25-HC production physiologically, murine macrophages are stimulated with a toll-like receptor 4 ligand, and its effect on SREBP-2 and LXR is examined. The results also suggest that de novo synthesis of 25-HC preferentially regulates SREBP-2 activity. Finally, we quantitatively determine the specificity of the four cholesterol hydroxylases in living cells. Based on our current findings, we conclude that endogenous side-chain oxysterols primarily regulate the activity of SREBP-2, not LXR.

Keywords:  CH25H; CYP27A1; CYP46A1; Cholesterol; LXR; SREBP-2; cholesterol metabolism; cholesterol regulation; lipid metabolism; lipid signaling; oxysterols

DOI:  https://doi.org/10.1016/j.jbc.2022.102733
EMBO J. 2022 Nov 21. e112677

Cholesterol transfer via endoplasmic reticulum contacts mediates lysosome damage repair.

Maja Radulovic, Eva Maria Wenzel, Sania Gilani, Lya Kk Holland, Alf Håkon Lystad, Santosh Phuyal, Vesa M Olkkonen, Andreas Brech, Marja Jäättelä, Kenji Maeda, Camilla Raiborg, Harald Stenmark.

  Lysosome integrity is essential for cell viability, and lesions in lysosome membranes are repaired by the ESCRT machinery. Here, we describe an additional mechanism for lysosome repair that is activated independently of ESCRT recruitment. Lipidomic analyses showed increases in lysosomal phosphatidylserine and cholesterol after damage. Electron microscopy demonstrated that lysosomal membrane damage is rapidly followed by the formation of contacts with the endoplasmic reticulum (ER), which depends on the ER proteins VAPA/B. The cholesterol-binding protein ORP1L was recruited to damaged lysosomes, accompanied by cholesterol accumulation by a mechanism that required VAP-ORP1L interactions. The PtdIns 4-kinase PI4K2A rapidly produced PtdIns4P on lysosomes upon damage, and knockout of PI4K2A inhibited damage-induced accumulation of ORP1L and cholesterol and led to the failure of lysosomal membrane repair. The cholesterol-PtdIns4P transporter OSBP was also recruited upon damage, and its depletion caused lysosomal accumulation of PtdIns4P and resulted in cell death. We conclude that ER contacts are activated on damaged lysosomes in parallel to ESCRTs to provide lipids for membrane repair, and that PtdIns4P generation and removal are central in this response.

Keywords:  cholesterol; lysosome; membrane contact site; membrane repair; phosphoinositide

DOI:  https://doi.org/10.15252/embj.2022112677
Free Radic Biol Med. 2022 Nov 17. pii: S0891-5849(22)00984-4. [Epub ahead of print]

Modulation of oxidative neurometabolism in ischemia/reperfusion by nitrite.

C Dias, C F Lourenço, J Laranjinha, A Ledo.

  Nitrite has been viewed in the past essentially as an inert metabolic endpoint of nitric oxide (•NO). However, it has become evident that, under certain conditions, nitrite can be a source of •NO. In the brain, this alternative source of •NO production independent of nitric oxide synthase activity may be particularly relevant in ischemia/reperfusion (I/R), where low oxygen availability limits enzymatic production of •NO. Notably, in vivo concentration of nitrite can be easily increased with diet, through the ingestion of nitrate-rich foods, opening the window for a therapeutic intervention based on diet. Considering the modulation of mitochondrial respiration by •NO, we have hypothesized that the protective action of nitrite in I/R may also result from modulation of mitochondrial function. We used high-resolution respirometry to evaluate the effects of nitrite in two in vitro models of I/R. In both cases, an increase in oxygen flux was observed following reoxygenation, a phenomenon that has been coined "oxidative burst". The amplitude of this "oxidative burst" was decreased by nitrite in a concentration-dependent manner. Additionally, a pilot in vivo study in which animals received a nitrate-rich diet as a strategy to increase circulating and tissue levels of nitrite also revealed that the "oxidative burst" was decreased in the nitrate-treated animals. These results may provide mechanistic support to the observation of a protective effect of nitrite in situations of brain ischemia.

Keywords:  Hippocampus; Ischemia-reperfusion; Nitric oxide; Nitrite; Oxidative burst; Respirometry

DOI:  https://doi.org/10.1016/j.freeradbiomed.2022.11.021