Metabolism
Abstract
Clear cell renal cell carcinomas (ccRCC) are largely driven by HIF2α and are avid consumers of glutamine. However, inhibitors of glutaminase1 (GLS1), the first step in glutaminolysis, have not shown benefit in phase III trials, and HIF2α inhibition, recently FDA-approved for treatment of ccRCC, shows significant but incomplete benefits. This highlights the need to better understand the interplay between glutamine metabolism and HIF2α in ccRCC. Here, we report that glutamine deprivation rapidly redistributes GLS1 into isolated clusters within mitochondria in diverse cell types, but not in ccRCC. GLS1 clustering occurs rapidly within 1 to 3 hours, is reversible, is specifically triggered by reduced intracellular glutamate, and is dependent on mitochondrial fission. Clustered GLS1 markedly enhances glutaminase activity and promotes cell death under glutamine-deprived conditions. HIF2α prevents GLS1 clustering, independently of its transcriptional activity, thereby maintaining low GLS activity and protecting ccRCC cells from glutamine deprivation-induced cell death. Forced clustering of GLS1, using constitutively clustering mutants, restores high GLS activity, promotes apoptosis, and suppresses ccRCC tumor growth in vivo. These findings reveal multiple insights into cellular glutamine handling, including a previously unrecognized process by which HIF2α promotes ccRCC: by suppressing GLS1 clustering and maintaining low GLS activity. This mechanism provides a potential explanation for the lack of clinical efficacy of GLS inhibitors in ccRCC and suggests a therapeutic avenue to combine HIF2α inhibition with strategies that restore GLS1 clustering.
Authors
Wencao Zhao, Sara M. Demczyszyn, Nathan J. Coffey, Yanqing Jiang, Boyoung Kim, Schuyler Bowers, Caitlyn E. Bowman, Michael C. Noji, Cholsoon Jang, M. Celeste Simon, Zoltan Arany, Boa Kim
Abstract
Angiopoietin-like 3 (ANGPTL3) is a major regulator of lipoprotein metabolism. ANGPTL3 deficiency results in lower levels of triglycerides, LDL-cholesterol (LDL-C), and HDL-cholesterol (HDL-C), and may protect from cardiovascular disease. ANGPTL3 oligomerizes with ANGPTL8 to inhibit lipoprotein lipase (LPL), the enzyme responsible for plasma triglyceride hydrolysis. Independent of ANGPTL8, oligomers of ANGPTL3 can inhibit endothelial lipase (EL), which regulates circulating HDL-C and LDL-C levels through the hydrolysis of lipoprotein phospholipids. The N-terminal region of ANGPTL3 is necessary for both oligomerization and lipase inhibition. However, our understanding of the specific residues that contribute to these functions is incomplete. In this study, we performed mutagenesis of the N-terminal region to identify residues important for EL inhibition and oligomerization. We also assessed the presence of different ANGPTL3 species in human plasma. We identified a motif important for lipase inhibition, and protein structure prediction suggested that this region interacted directly with EL. We also found that recombinant ANGPTL3 formed a homotrimer and was unable to inhibit EL activity when trimerization was disrupted. Surprisingly, we observed that human plasma contained more monomeric ANGPTL3 than trimeric ANGPTL3. An important implication of these findings is that previous correlations between circulating ANGPTL3 and circulating triglyceride-rich lipoproteins need to be revisited.
Authors
Sydney G. Walker, Yan Q. Chen, Kelli L. Sylvers-Davie, Alex Dou, Eugene Y. Zhen, Yuewei Qian, Yi Wen, Mariam E. Ehsani, Sydney A. Smith, Rakshya Thapa, Maxwell J. Mercer, Lucy Langmack, Bharat Raj Bhattarai, Michael Ploug, Robert J. Konrad, Brandon S.J. Davies
Abstract
Recent evidence suggests that cellular metabolism, including glycolysis and fatty acid synthesis in lymphatic endothelial cells (LECs), plays essential roles in developing functional lymphatic systems. Site-1 protease (S1P) proteolytically activates membrane-bound latent transcription factor sterol regulatory element-binding proteins (SREBPs), which are required to induce lipid biosynthesis. In this study, we generated mice with pan-endothelial or LEC-specific deficiency of either S1P or SREBP2. Mouse embryos with pan-endothelial deletion of S1P showed defective lymphatic vessel migration in skin and lymphedema, while their blood vasculature formation was relatively normal. Mice lacking S1P in LECs or SREBP2 in LECs exhibited chylous ascites, reduced lipogenic gene expression, and reduced VEGFR3 expression and progressively developed wasting, resulting in postnatal death by approximately 8 weeks of age. Additionally, mice with SREBP2 deletion in LECs exhibited dilated lacteal and mesenteric lymphatics and accumulation of lipids in the lacteal before weaning age, indicating apparent lymphatic malfunctioning. These data indicate that S1P-SREBP2–mediated cholesterol biosynthesis is pivotal in lymphatic vascular development. We also found that treating human dermal LECs with VEGF-C induced proteolytic activation of SREBP2 with concomitant phosphorylation of Akt and the expression of genes involved in cholesterol biosynthesis. Those effects were canceled out by treating the cells with an S1P inhibitor or SREBP inhibitor. These data demonstrate that the S1P/SREBP2 axis is critical in VEGF-C/VEGFR3 mitogenic signaling in LECs.
Authors
Yuji Kondo, Yizhi Jiang, Xin Geng, Jianhua Song, Summer Simeroth, J. Michael McDaniel, Pengchun Yu, R. Sathish Srinivasan, Lijun Xia
Abstract
Poor skeletal muscle fitness contributes to many chronic disease states including obesity, heart failure, primary muscle disorders, and age-related sarcopenia. Receptor Interacting Protein 140 (RIP140) is a striated muscle-enriched nuclear receptor coregulator known to suppress mitochondrial oxidative capacity. To investigate the role of RIP140 in skeletal muscle, striated muscle-specific RIP140-deficient (strNrip1-/-) mice were generated and characterized. strNrip1-/- mice displayed an enhanced endurance performance phenotype. RNA-sequence (RNA-seq) analysis of glycolytic fast-twitch muscle from strNrip1-/- mice identified a broad array of differentially upregulated metabolic and structural muscle genes known to be induced by endurance training, including pathways involved in mitochondrial biogenesis and respiration, fatty acid oxidation, slow muscle fiber type, and angiogenesis. In addition, muscle RIP140-deficiency induced expansive neuromuscular junction (NMJ) remodeling. Integration of RNA sequence results with CUT&RUN analysis of strNrip1-/- myotubes identified Wnt16 as a candidate effector for the NMJ biogenesis in RIP140-deficient skeletal myotubes. We conclude that RIP140 serves as a physiological “rheostat” for a broad coordinated network of metabolic and structural genes involved in skeletal muscle fitness.
Authors
Elizabeth Pruzinsky, Kirill Batmanov, Denis M. Medeiros, Sarah M. Sulon, Brian P. Sullivan, Tomoya Sakamoto, Teresa C. Leone, Tejvir S. Khurana, Daniel P. Kelly
Abstract
Impaired wound healing poses a major and increasingly frequent health problem. Among the key players in the healing process are fibroblasts, but their metabolic profile in healing wounds is largely unknown. Using a combination of transcriptomics, targeted proteomics and metabolomics, we identified retinol metabolism as a top regulated pathway in wound fibroblasts. This is functionally relevant, since even a mild retinol deficiency caused a delay in wound closure and re-epithelialization, which mainly resulted from misdirected keratinocyte migration on the new granulation tissue. Quantitative proteomics identified integrin alpha 11 (Itga11) as a less abundant protein in wounds of mice subjected to a retinol-deficient diet. Reduced levels of this fibroblast-specific protein likely altered the granulation tissue matrix, which in turn affected re-epithelialization. These results provide a comprehensive overview on the transcriptome, proteome and metabolome of wound fibroblasts and identify retinol metabolism in fibroblasts as a key regulator of tissue repair.
Authors
Till Wüstemann, Elizabeta Madzharova, Mateusz S. Wietecha, Norbert B. Ghyselinck, Marcus Höring, Gerhard Liebisch, Nicola Zamboni, Ulrich auf dem Keller, Sabine Werner
Abstract
Long chain fatty acid oxidation disorders (LC FAODs) cause energy deficits in heart and skeletal muscle that are only partially corrected by current medium chain lipid therapies such as triheptanoin. We find that heart and muscle lack medium chain acyl CoA synthetases, limiting the capacity for β-oxidation of medium-chain fatty acids. Instead, heart and muscle mitochondria robustly respire on medium-chain acylcarnitines. The mitochondrial matrix enzyme carnitine acetyltransferase (CrAT) efficiently converts orally delivered octanoylcarnitine (C8 carnitine) to octanoyl CoA for energy generation. C8-carnitine exhibits twice the oral bioavailability of triheptanoin and distributes to muscle and heart. A single oral dose significantly enhances grip strength and treadmill endurance while attenuating lactic acidosis in two mouse models of LC-FAODs. Thus, medium chain acylcarnitines overcome a previously unrecognized metabolic bottleneck in LC FAOD muscle and may represent an alternative to triglyceride based therapies for bioenergetic disorders.
Authors
Keaton J. Solo, Yuxun Zhang, Sivakama S. Bharathi, Bob B. Zhang, Adam C. Richert, Alexandra V. Schmidt, Clinton Van't Land, Olivia D'Annibale, Timothy C. Wood, Eric S. Goetzman
Abstract
BACKGROUND Icosapent ethyl (IPE), an ethyl ester of eicosapentaenoic acid (EPA), reduces cardiovascular disease (CVD), but the mechanism remains elusive. We examined the effect of IPE supplementation on lipoprotein subclasses, lipidomes, and pro-atherogenic properties.METHODS Using 3 independent metabolomic platforms, we examined the effect of high-dose IPE supplementation for 28 days on fatty acid profiles, lipoprotein subclasses, lipidomes, and pro-atherogenic properties in normolipidemic volunteers (n = 38).RESULTS IPE supplementation increased lipoprotein EPA on average 4-fold within 7 days, returning to baseline after a 7-day washout. Notably, the incorporation displayed marked interindividual variance, negatively correlating with baseline levels. We identified persistent participant-specific lipoprotein fingerprints despite uniform IPE-induced lipidome remodeling across all lipoprotein classes. This remodeling resulted in reductions in saturated, monounsaturated, and n-6 polyunsaturated fatty acids, resulting in reduced clinical risk markers, including triglyceride, remnant cholesterol, and apolipoprotein B (apoB) levels and 10-year CVD risk score. Of the pro-atherogenic properties tested, IPE significantly reduced apoB lipoprotein binding to proteoglycans, which correlated with lower apoB particle concentration, cholesterol content, and specific lipid species in LDL, including phosphatidylcholine 38:3 previously associated with CVD.CONCLUSION These findings highlight IPE’s rapid, uniform remodeling of lipoproteins and reduced proteoglycan binding, likely contributing to previously observed CVD risk reduction. Persistent interindividual lipidome signatures underscore the potential for personalized therapeutic approaches in atherosclerotic CVD treatment.TRIAL REGISTRATION NCT04152291.FUNDING Jenny and Antti Wihuri Foundation, Research Council of Finland, Sigrid Jusélius Foundation, Finnish Foundation for Cardiovascular Research, Emil Aaltonen Foundation, Ida Montin Foundation, Novo Nordisk Foundation, Finnish Cultural Foundation, and Jane and Aatos Erkko Foundation.
Authors
Lauri Äikäs, Petri T. Kovanen, Martina B. Lorey, Reijo Laaksonen, Minna Holopainen, Hanna Ruhanen, Reijo Käkelä, Matti Jauhiainen, Martin Hermansson, Katariina Öörni
Abstract
Atherosclerotic cardiovascular disease is a major contributor to the global disease burden. Atherosclerosis initiation depends on cholesterol accumulation in subendothelial macrophages (Mφs). To clarify the role of Bmal1 in Mφ function and atherosclerosis, we used several global and myeloid-specific Bmal1 deficient mouse models. Myeloid-specific Bmal1 deficient mice had higher Mφ cholesterol and displayed greater atherosclerosis compared to controls. Bmal1-deficient Mφs exhibited: (1) elevated expression of Cd36 and uptake of oxLDL; (2) diminished expression of Abca1 and Abcg1, and decreased cholesterol efflux and reverse cholesterol transport; and (3) reduced Npc1 and Npc2 expression, and diminished cholesterol egress from lysosomes. Molecular studies revealed that Bmal1 directly regulates basal and cyclic expression of Npc1 and Npc2 by binding the E-boxes in their promoters and indirectly regulates the basal and temporal regulation of Cd36 and Abca1/Abcg1 involving Rev-erbα and Znf202 repressors, respectively. In conclusion, Mφ Bmal1 is a key regulator of the uptake of modified lipoproteins, cholesterol efflux, lysosomal cholesterol egress and atherosclerosis, and therefore may be a master regulator of cholesterol metabolism in Mφs. Restoration of Mφ Bmal1 expression or blocking of factors that decrease its activity may be effective in preventing atherosclerosis.
Authors
Xiaoyue Pan, John O'Hare, Cyrus Mowdawalla, Samantha Mota, Nan Wang, M. Mahmood Hussain
Abstract
Glioblastoma IDH-wildtype is the most common and aggressive primary brain tumor in adults, with poor prognosis despite current therapies. To identify new therapeutic vulnerabilities, we investigated the role of CDK12, a transcription-associated cyclin-dependent kinase, in glioblastoma. Genetic or pharmacologic inactivation of CDK12 impaired tumor growth in patientderived xenograft (PDX) models and enhanced the efficacy of temozolomide. Metabolic profiling using extracellular flux analysis and stable isotope tracing with U-¹³C-glucose and U-¹³Cglutamine showed that CDK12 inhibition disrupted mitochondrial respiration, resulting in energy depletion and apoptotic cell death characterized by caspase activation and Noxa induction. Mechanistically, we identified a direct interaction between CDK12 and GSK3β. CDK12 inhibition activated GSK3β, leading to downregulation of PPARD, a transcriptional regulator of oxidative metabolism. This CDK12–GSK3β–PPARD axis was required for glioblastoma cell proliferation and metabolic homeostasis. In vivo, CDK12 inhibition significantly extended survival without overt toxicity and induced complete tumor regression in a subset of animals. Strikingly, combined CDK12 inhibition and temozolomide treatment led to complete tumor eradication in all animals tested. These findings establish CDK12 as a key regulator of glioblastoma metabolism and survival, and provide strong preclinical rationale for its therapeutic targeting in combination with standard-of-care treatments.
Authors
Jeong-Yeon Mun, Chang Shu, Qiuqiang Gao, Zhe Zhu, Hasan O. Akman, Mike-Andrew Westhoff, Georg Karpel-Massler, Markus D. Siegelin
Abstract
Recent experimental and epidemiologic data have strongly associated air pollution in the pathogenesis of insulin resistance and type 2 diabetes mellitus. We explored the effect of inhalational exposure to concentrated ambient particulate matter smaller than 2.5 μm (PM2.5), or filtered air, using a whole-body inhalation system (6 hours/day, 5 days/week) for 24 weeks on metabolism and brown adipose tissue (BAT) function. Mechanistic evaluation of insulin resistance, glucose uptake with 18F-fluorodeoxyglucose positron emission tomography, alongside evaluation for differentially methylated regions, chromatin accessibility, and differential expression of genes was performed. PM2.5 exposure impaired metabolism through changes in key BAT transcriptional programs involved in redox stress, lipid deposition, fibrosis, and altered thermogenesis. Significant differential methylation and widespread chromatin remodeling was noted in BAT with PM2.5. Integrated analysis uncovered a role for the histone deacetylase HDAC9 and histone demethylase KDM2B. The latter demethylates Lys-4 and Lys-36 of histone H3. Specifically, studies using ChIP combined with quantitative PCR confirmed HDAC9 and KDM2B occupancy and reduced H3K36me2 on the promoter of target BAT genes in PM2.5 mice, while Hdac9/Kdm2b knockdown and overexpression increased and reduced BAT metabolism, respectively. Collectively, our results provide insights into air pollution exposure and changes in BAT and metabolism.
Authors
Rengasamy Palanivel, Jean-Eudes Dazard, Bongsoo Park, Sarah Costantino, Skanda T. Moorthy, Armando Vergara-Martel, Elaine Ann Cara, Jonnelle Edwards-Glenn, Shyam Biswal, Lung Chi Chen, Mukesh K. Jain, Francesco Paneni, Sanjay Rajagopalan
No posts were found with this tag.
