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1.
bioRxiv ; 2023 Apr 29.
Artigo em Inglês | MEDLINE | ID: mdl-37162991

RESUMO

5-fluorouracil (5-FU) is a successful and broadly used anti-cancer therapeutic. A major mechanism of action of 5-FU is thought to be through thymidylate synthase (TYMS) inhibition resulting in dTTP depletion and activation of the DNA damage response. This suggests that 5-FU should synergize with other DNA damaging agents. However, we found that combinations of 5-FU and oxaliplatin or irinotecan failed to display any evidence of synergy in clinical trials, and resulted in sub-additive killing in a panel of colorectal cancer (CRC) cell lines. In seeking to understand this antagonism, we unexpectedly found that an RNA damage response during ribosome biogenesis dominates the drug's efficacy in tumor types for which 5-FU shows clinical benefit. 5-FU has an inherent bias for RNA incorporation, and blocking this greatly reduced drug-induced lethality, indicating that accumulation of damaged RNA is more deleterious than the lack of new RNA synthesis. Using 5-FU metabolites that specifically incorporate into either RNA or DNA revealed that CRC cell lines and patient-derived colorectal cancer organoids are inherently more sensitive to RNA damage. This difference held true in cell lines from other tissues in which 5-FU has shown clinical utility, whereas cell lines from tumor tissues that lack clinical 5-FU responsiveness typically showed greater sensitivity to the drug's DNA damage effects. Analysis of changes in the phosphoproteome and ubiquitinome shows RNA damage triggers the selective ubiquitination of multiple ribosomal proteins leading to autophagy-dependent rRNA catabolism and proteasome-dependent degradation of ubiquitinated ribosome proteins. Further, RNA damage response to 5-FU is selectively enhanced by compounds that promote ribosome biogenesis, such as KDM2A inhibitors. These results demonstrate the presence of a strong RNA damage response linked to apoptotic cell death, with clear utility of combinatorially targeting this response in cancer therapy.

2.
Nat Metab ; 4(12): 1792-1811, 2022 12.
Artigo em Inglês | MEDLINE | ID: mdl-36536136

RESUMO

The mechanistic target of rapamycin complex 1 (mTORC1) senses and relays environmental signals from growth factors and nutrients to metabolic networks and adaptive cellular systems to control the synthesis and breakdown of macromolecules; however, beyond inducing de novo lipid synthesis, the role of mTORC1 in controlling cellular lipid content remains poorly understood. Here we show that inhibition of mTORC1 via small molecule inhibitors or nutrient deprivation leads to the accumulation of intracellular triglycerides in both cultured cells and a mouse tumor model. The elevated triglyceride pool following mTORC1 inhibition stems from the lysosome-dependent, but autophagy-independent, hydrolysis of phospholipid fatty acids. The liberated fatty acids are available for either triglyceride synthesis or ß-oxidation. Distinct from the established role of mTORC1 activation in promoting de novo lipid synthesis, our data indicate that mTORC1 inhibition triggers membrane phospholipid trafficking to the lysosome for catabolism and an adaptive shift in the use of constituent fatty acids for storage or energy production.


Assuntos
Ácidos Graxos , Lisossomos , Camundongos , Animais , Alvo Mecanístico do Complexo 1 de Rapamicina/metabolismo , Lisossomos/metabolismo , Triglicerídeos/metabolismo , Ácidos Graxos/metabolismo , Fosfolipídeos/metabolismo
3.
iScience ; 25(11): 105458, 2022 Nov 18.
Artigo em Inglês | MEDLINE | ID: mdl-36388985

RESUMO

mTORC1 is aberrantly activated in cancer and in the genetic tumor syndrome tuberous sclerosis complex (TSC), which is caused by loss-of-function mutations in the TSC complex, a negative regulator of mTORC1. Clinically approved mTORC1 inhibitors, such as rapamycin, elicit a cytostatic effect that fails to eliminate tumors and is rapidly reversible. We sought to determine the effects of mTORC1 on the core regulators of intrinsic apoptosis. In TSC2-deficient cells and tumors, we find that mTORC1 inhibitors shift cellular dependence from MCL-1 to BCL-2 and BCL-XL for survival, thereby altering susceptibility to BH3 mimetics that target specific pro-survival BCL-2 proteins. The BCL-2/BCL-XL inhibitor ABT-263 synergizes with rapamycin to induce apoptosis in TSC-deficient cells and in a mouse tumor model of TSC, resulting in a more complete and durable response. These data expose a therapeutic vulnerability in regulation of the apoptotic machinery downstream of mTORC1 that promotes a cytotoxic response to rapamycin.

4.
Nat Metab ; 4(6): 711-723, 2022 06.
Artigo em Inglês | MEDLINE | ID: mdl-35739397

RESUMO

Production of oxidized biomass, which requires regeneration of the cofactor NAD+, can be a proliferation bottleneck that is influenced by environmental conditions. However, a comprehensive quantitative understanding of metabolic processes that may be affected by NAD+ deficiency is currently missing. Here, we show that de novo lipid biosynthesis can impose a substantial NAD+ consumption cost in proliferating cancer cells. When electron acceptors are limited, environmental lipids become crucial for proliferation because NAD+ is required to generate precursors for fatty acid biosynthesis. We find that both oxidative and even net reductive pathways for lipogenic citrate synthesis are gated by reactions that depend on NAD+ availability. We also show that access to acetate can relieve lipid auxotrophy by bypassing the NAD+ consuming reactions. Gene expression analysis demonstrates that lipid biosynthesis strongly anti-correlates with expression of hypoxia markers across tumor types. Overall, our results define a requirement for oxidative metabolism to support biosynthetic reactions and provide a mechanistic explanation for cancer cell dependence on lipid uptake in electron acceptor-limited conditions, such as hypoxia.


Assuntos
NAD , Neoplasias , Proliferação de Células , Elétrons , Humanos , Hipóxia , Lipídeos , NAD/metabolismo
5.
Cell Rep ; 39(7): 110824, 2022 05 17.
Artigo em Inglês | MEDLINE | ID: mdl-35584673

RESUMO

The tuberous sclerosis complex (TSC) 1 and 2 proteins associate with TBC1D7 to form the TSC complex, which is an essential suppressor of mTOR complex 1 (mTORC1), a ubiquitous driver of cell and tissue growth. Loss-of-function mutations in TSC1 or TSC2, but not TBC1D7, give rise to TSC, a pleiotropic disorder with aberrant activation of mTORC1 in various tissues. Here, we characterize mice with genetic deletion of Tbc1d7, which are viable with normal growth and development. Consistent with partial loss of function of the TSC complex, Tbc1d7 knockout (KO) mice display variable increases in tissue mTORC1 signaling with increased muscle fiber size but with strength and motor defects. Their most pronounced phenotype is brain overgrowth due to thickening of the cerebral cortex, with enhanced neuron-intrinsic mTORC1 signaling and growth. Thus, TBC1D7 is required for full TSC complex function in tissues, and the brain is particularly sensitive to its growth-suppressing activities.


Assuntos
Encéfalo , Peptídeos e Proteínas de Sinalização Intracelular , Alvo Mecanístico do Complexo 1 de Rapamicina , Neurônios , Proteína 1 do Complexo Esclerose Tuberosa , Esclerose Tuberosa , Proteínas Supressoras de Tumor , Animais , Encéfalo/crescimento & desenvolvimento , Encéfalo/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Alvo Mecanístico do Complexo 1 de Rapamicina/metabolismo , Camundongos , Camundongos Knockout , Neurônios/citologia , Neurônios/metabolismo , Esclerose Tuberosa/metabolismo , Esclerose Tuberosa/patologia , Proteína 1 do Complexo Esclerose Tuberosa/metabolismo , Proteína 2 do Complexo Esclerose Tuberosa/metabolismo , Proteínas Supressoras de Tumor/genética , Proteínas Supressoras de Tumor/metabolismo
6.
Cancer Res ; 81(19): 4896-4898, 2021 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-34598998

RESUMO

The Warburg effect, the propensity of some cells to metabolize glucose to lactate in the presence of oxygen (also known as aerobic glycolysis), has long been observed in cancer and other contexts of cell proliferation, but only in the past two decades have significant gains been made in understanding how and why this metabolic transformation occurs. In 2004, Cancer Research published a study by Elstrom and colleagues that provided one of the first connections between a specific oncogene and aerobic glycolysis. Studying hematopoietic and glioblastoma cell lines, they demonstrated that constitutive activation of AKT promotes an increased glycolytic rate without altering proliferation or oxygen consumption in culture. They proposed that it is this effect that allows constitutive AKT activation to transform cells and found that it sensitizes cells to glucose deprivation. In the years since, mechanistic understanding of oncogenic control of metabolism, and glycolysis specifically, has deepened substantially. Current work seeks to understand the benefits and liabilities associated with glycolytic metabolism and to identify inhibitors that might be of clinical benefit to target glycolytic cancer cells.See related article by Elstrom and colleagues, Cancer Res 2004;64:3892-9.


Assuntos
Glioblastoma , Proteínas Proto-Oncogênicas c-akt , Proliferação de Células , Ciclo do Ácido Cítrico , Glioblastoma/genética , Glucose , Glicólise , Humanos , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de Sinais
7.
FEBS J ; 288(19): 5629-5649, 2021 10.
Artigo em Inglês | MEDLINE | ID: mdl-33811729

RESUMO

Many metabolic phenotypes in cancer cells are also characteristic of proliferating nontransformed mammalian cells, and attempts to distinguish between phenotypes resulting from oncogenic perturbation from those associated with increased proliferation are limited. Here, we examined the extent to which metabolic changes corresponding to oncogenic KRAS expression differed from those corresponding to epidermal growth factor (EGF)-driven proliferation in human mammary epithelial cells (HMECs). Removal of EGF from culture medium reduced growth rates and glucose/glutamine consumption in control HMECs despite limited changes in respiration and fatty acid synthesis, while the relative contribution of branched-chain amino acids to the TCA cycle and lipogenesis increased in the near-quiescent conditions. Most metabolic phenotypes measured in HMECs expressing mutant KRAS were similar to those observed in EGF-stimulated control HMECs that were growing at comparable rates. However, glucose and glutamine consumption as well as lactate and glutamate production were lower in KRAS-expressing cells cultured in media without added EGF, and these changes correlated with reduced sensitivity to GLUT1 inhibitor and phenformin treatment. Our results demonstrate the strong dependence of metabolic behavior on growth rate and provide a model to distinguish the metabolic influences of oncogenic mutations and nononcogenic growth.


Assuntos
Neoplasias da Mama/genética , Carcinogênese/genética , Fator de Crescimento Epidérmico/genética , Transportador de Glucose Tipo 1/genética , Proteínas Proto-Oncogênicas p21(ras)/genética , Animais , Mama/crescimento & desenvolvimento , Mama/patologia , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Proliferação de Células/genética , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Feminino , Regulação Neoplásica da Expressão Gênica/genética , Glucose/metabolismo , Transportador de Glucose Tipo 1/antagonistas & inibidores , Ácido Glutâmico/metabolismo , Glutamina/metabolismo , Humanos , Ácido Láctico/metabolismo , Glândulas Mamárias Humanas/crescimento & desenvolvimento , Glândulas Mamárias Humanas/patologia , Células Tumorais Cultivadas
8.
Elife ; 102021 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-33646118

RESUMO

The mechanistic target of rapamycin complex 1 (mTORC1) stimulates a coordinated anabolic program in response to growth-promoting signals. Paradoxically, recent studies indicate that mTORC1 can activate the transcription factor ATF4 through mechanisms distinct from its canonical induction by the integrated stress response (ISR). However, its broader roles as a downstream target of mTORC1 are unknown. Therefore, we directly compared ATF4-dependent transcriptional changes induced upon insulin-stimulated mTORC1 signaling to those activated by the ISR. In multiple mouse embryo fibroblast and human cancer cell lines, the mTORC1-ATF4 pathway stimulated expression of only a subset of the ATF4 target genes induced by the ISR, including genes involved in amino acid uptake, synthesis, and tRNA charging. We demonstrate that ATF4 is a metabolic effector of mTORC1 involved in both its established role in promoting protein synthesis and in a previously unappreciated function for mTORC1 in stimulating cellular cystine uptake and glutathione synthesis.


When building healthy tissue, the human body must carefully control the growth of new cells to prevent them from becoming cancerous. A core component of this regulation is the protein mTORC1, which responds to various growth-stimulating factors and nutrients, and activates the chemical reactions cells need to grow. Part of this process involves controlling 'nutrient-sensing transcription factors' ­ proteins that regulate the activity of specific genes based on the availability of different nutrients. One of these nutrient-sensing transcription factors, ATF4, has recently been shown to be involved in some of the processes triggered by mTORC1. The role this factor plays in how cells respond to stress ­ such as when specific nutrients are depleted, protein folding is disrupted or toxins are present ­ is well-studied. But how it reacts to the activation of mTORC1 is less clear. To bridge this gap, Torrence et al. studied mouse embryonic cells and human prostate cancer cells grown in the laboratory, to see whether mTORC1 influenced the behavior of ATF4 differently than cellular stress. Cells were treated either with insulin, which activates mTORC1, or an antibiotic that sparks the stress response. The cells were then analyzed using a molecular tool to see which genes were switched on by ATF4 following treatment. This revealed that less than 10% of the genes activated by ATF4 during cellular stress are also activated in response to mTORC1-driven growth. Many of the genes activated in both scenarios were involved in synthesizing and preparing the building blocks that make up proteins. This was consistent with the discovery that ATF4 helps mTORC1 stimulate growth by promoting protein synthesis. Torrence et al. also found that mTORC1's regulation of ATF4 stimulated the synthesis of glutathione, the most abundant antioxidant in cells. The central role mTORC1 plays in controlling cell growth means it is important to understand how it works and how it can lead to uncontrolled growth in human diseases. mTORC1 is activated in many overgrowth syndromes and the majority of human cancers. These new findings could provide insight into how tumors coordinate their drive for growth while adapting to cellular stress, and reveal new drug targets for cancer treatment.


Assuntos
Fator 4 Ativador da Transcrição/metabolismo , Glutationa/biossíntese , Alvo Mecanístico do Complexo 1 de Rapamicina/efeitos dos fármacos , Fator 4 Ativador da Transcrição/genética , Animais , Linhagem Celular , Linhagem Celular Tumoral , Embrião de Mamíferos , Fibroblastos , Humanos , Insulina/farmacologia , Alvo Mecanístico do Complexo 1 de Rapamicina/genética , Alvo Mecanístico do Complexo 1 de Rapamicina/metabolismo , Camundongos , Transdução de Sinais
9.
Sci Rep ; 10(1): 12054, 2020 Jul 21.
Artigo em Inglês | MEDLINE | ID: mdl-32694612

RESUMO

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

10.
FEBS Lett ; 594(4): 646-664, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-31642061

RESUMO

Mammalian pyruvate kinase catalyzes the final step of glycolysis, and its M2 isoform (PKM2) is widely expressed in proliferative tissues. Mutations in PKM2 are found in some human cancers; however, the effects of these mutations on enzyme activity and regulation are unknown. Here, we characterized five cancer-associated PKM2 mutations, occurring at various locations on the enzyme, with respect to substrate kinetics and activation by the allosteric activator fructose-1,6-bisphosphate (FBP). The mutants exhibit reduced maximal velocity, reduced substrate affinity, and/or altered activation by FBP. The kinetic parameters of five additional PKM2 mutants that have been used to study enzyme function or regulation also demonstrate the deleterious effects of mutations on PKM2 function. Our findings indicate that PKM2 is sensitive to many amino acid changes and support the hypothesis that decreased PKM2 activity is selected for in rapidly proliferating cells.


Assuntos
Proteínas de Transporte/genética , Proteínas de Membrana/genética , Mutação , Neoplasias/genética , Hormônios Tireóideos/genética , Proteínas de Transporte/química , Proteínas de Transporte/metabolismo , Humanos , Cinética , Proteínas de Membrana/química , Proteínas de Membrana/metabolismo , Neoplasias/enzimologia , Multimerização Proteica/genética , Estrutura Quaternária de Proteína , Hormônios Tireóideos/química , Hormônios Tireóideos/metabolismo , Proteínas de Ligação a Hormônio da Tireoide
11.
Nat Commun ; 10(1): 5604, 2019 12 06.
Artigo em Inglês | MEDLINE | ID: mdl-31811141

RESUMO

Increased glucose uptake and metabolism is a prominent phenotype of most cancers, but efforts to clinically target this metabolic alteration have been challenging. Here, we present evidence that lactoylglutathione (LGSH), a byproduct of methylglyoxal detoxification, is elevated in both human and murine non-small cell lung cancers (NSCLC). Methylglyoxal is a reactive metabolite byproduct of glycolysis that reacts non-enzymatically with nucleophiles in cells, including basic amino acids, and reduces cellular fitness. Detoxification of methylglyoxal requires reduced glutathione (GSH), which accumulates to high levels in NSCLC relative to normal lung. Ablation of the methylglyoxal detoxification enzyme glyoxalase I (Glo1) potentiates methylglyoxal sensitivity and reduces tumor growth in mice, arguing that targeting pathways involved in detoxification of reactive metabolites is an approach to exploit the consequences of increased glucose metabolism in cancer.


Assuntos
Glucose/metabolismo , Glicólise , Neoplasias Pulmonares/metabolismo , Animais , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Linhagem Celular Tumoral , Glutationa/metabolismo , Humanos , Inativação Metabólica , Lactoilglutationa Liase/metabolismo , Pulmão/metabolismo , Masculino , Metabolômica , Camundongos , Camundongos Endogâmicos C57BL , Aldeído Pirúvico/metabolismo , Aldeído Pirúvico/toxicidade
12.
J Biol Chem ; 293(52): 20051-20061, 2018 12 28.
Artigo em Inglês | MEDLINE | ID: mdl-30381394

RESUMO

Monoallelic point mutations in the gene encoding the cytosolic, NADP+-dependent enzyme isocitrate dehydrogenase 1 (IDH1) cause increased production of the oncometabolite 2-hydroxyglutarate (2-HG) in multiple cancers. Most IDH1 mutant tumors retain one wildtype (WT) IDH1 allele. Several studies have proposed that retention of this WT allele is protumorigenic by facilitating substrate channeling through a WT-mutant IDH1 heterodimer, with the WT subunit generating a local supply of α-ketoglutarate and NADPH that is then consumed by the mutant subunit to produce 2-HG. Here, we confirmed that coexpression of WT and mutant IDH1 subunits leads to formation of WT-mutant hetero-oligomers and increases 2-HG production. An analysis of a recently reported crystal structure of the WT-R132H IDH1 heterodimer and of in vitro kinetic parameters for 2-HG production, however, indicated that substrate channeling between the subunits is biophysically implausible. We also found that putative carbon-substrate flux between WT and mutant IDH1 subunits is inconsistent with the results of isotope tracing experiments in cancer cells harboring an endogenous monoallelic IDH1 mutation. Finally, using a mathematical model of WT-mutant IDH1 heterodimers, we estimated that the NADPH:NADP+ ratio is higher in the cytosol than in the mitochondria, suggesting that NADPH is unlikely to be limiting for 2-HG production in the cytosol. These findings argue against supply of either substrate being limiting for 2-HG production by a cytosolic IDH1 mutant and suggest that the retention of a WT allele in IDH1 mutant tumors is not due to a requirement for carbon or cofactor flux between WT and mutant IDH1.


Assuntos
Hidroxibutiratos/metabolismo , Isocitrato Desidrogenase , Modelos Biológicos , Mutação , Proteínas de Neoplasias , Neoplasias , Linhagem Celular Tumoral , Células HEK293 , Humanos , Isocitrato Desidrogenase/genética , Isocitrato Desidrogenase/metabolismo , NADP/genética , NADP/metabolismo , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Neoplasias/genética , Neoplasias/metabolismo , Neoplasias/patologia , Multimerização Proteica
13.
EMBO J ; 37(22)2018 11 15.
Artigo em Inglês | MEDLINE | ID: mdl-30348863

RESUMO

The Hippo pathway and its nuclear effector Yap regulate organ size and cancer formation. While many modulators of Hippo activity have been identified, little is known about the Yap target genes that mediate these growth effects. Here, we show that yap-/- mutant zebrafish exhibit defects in hepatic progenitor potential and liver growth due to impaired glucose transport and nucleotide biosynthesis. Transcriptomic and metabolomic analyses reveal that Yap regulates expression of glucose transporter glut1, causing decreased glucose uptake and use for nucleotide biosynthesis in yap-/- mutants, and impaired glucose tolerance in adults. Nucleotide supplementation improves Yap deficiency phenotypes, indicating functional importance of glucose-fueled nucleotide biosynthesis. Yap-regulated glut1 expression and glucose uptake are conserved in mammals, suggesting that stimulation of anabolic glucose metabolism is an evolutionarily conserved mechanism by which the Hippo pathway controls organ growth. Together, our results reveal a central role for Hippo signaling in glucose metabolic homeostasis.


Assuntos
Glucose/metabolismo , Fígado/embriologia , Nucleotídeos/biossíntese , Transdução de Sinais/fisiologia , Transativadores/metabolismo , Proteínas de Peixe-Zebra/metabolismo , Peixe-Zebra/embriologia , Animais , Glucose/genética , Transportador de Glucose Tipo 1/genética , Transportador de Glucose Tipo 1/metabolismo , Camundongos , Nucleotídeos/genética , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Serina-Treonina Quinase 3 , Transativadores/genética , Proteínas de Sinalização YAP , Peixe-Zebra/genética , Proteínas de Peixe-Zebra/genética
15.
Nat Cell Biol ; 20(7): 782-788, 2018 07.
Artigo em Inglês | MEDLINE | ID: mdl-29941931

RESUMO

Defining the metabolic limitations of tumour growth will help to develop cancer therapies1. Cancer cells proliferate slower in tumours than in standard culture conditions, indicating that a metabolic limitation may restrict cell proliferation in vivo. Aspartate synthesis can limit cancer cell proliferation when respiration is impaired2-4; however, whether acquiring aspartate is endogenously limiting for tumour growth is unknown. We confirm that aspartate has poor cell permeability, which prevents environmental acquisition, whereas the related amino acid asparagine is available to cells in tumours, but cancer cells lack asparaginase activity to convert asparagine to aspartate. Heterologous expression of guinea pig asparaginase 1 (gpASNase1), an enzyme that produces aspartate from asparagine5, confers the ability to use asparagine to supply intracellular aspartate to cancer cells in vivo. Tumours expressing gpASNase1 grow at a faster rate, indicating that aspartate acquisition is an endogenous metabolic limitation for the growth of some tumours. Tumours expressing gpASNase1 are also refractory to the growth suppressive effects of metformin, suggesting that metformin inhibits tumour growth by depleting aspartate. These findings suggest that therapeutic aspartate suppression could be effective to treat cancer.


Assuntos
Ácido Aspártico/metabolismo , Proliferação de Células , Metabolismo Energético , Neoplasias/metabolismo , Animais , Antineoplásicos/farmacologia , Asparaginase/genética , Asparaginase/metabolismo , Proliferação de Células/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos , Cobaias , Células HCT116 , Células HEK293 , Células HeLa , Humanos , Masculino , Metabolômica/métodos , Metformina/farmacologia , Camundongos Nus , Camundongos Transgênicos , Neoplasias/tratamento farmacológico , Neoplasias/genética , Neoplasias/patologia , Transdução de Sinais , Fatores de Tempo , Carga Tumoral , Microambiente Tumoral , Ensaios Antitumorais Modelo de Xenoenxerto
16.
J Biol Chem ; 293(20): 7490-7498, 2018 05 18.
Artigo em Inglês | MEDLINE | ID: mdl-29339555

RESUMO

Cell growth and division require nutrients, and proliferating cells use a variety of sources to acquire the amino acids, lipids, and nucleotides that support macromolecule synthesis. Lipids are more reduced than other nutrients, whereas nucleotides and amino acids are typically more oxidized. Cells must therefore generate reducing and oxidizing (redox) equivalents to convert consumed nutrients into biosynthetic precursors. To that end, redox cofactor metabolism plays a central role in meeting cellular redox requirements. In this Minireview, we highlight the biosynthetic pathways that involve redox reactions and discuss their integration with metabolism in proliferating mammalian cells.


Assuntos
Fenômenos Fisiológicos Celulares , Proliferação de Células , Metabolismo Energético , Mitocôndrias/metabolismo , Transdução de Sinais , Animais , Humanos , Mamíferos , Oxirredução
17.
Bio Protoc ; 8(11): e2876, 2018 Jun 05.
Artigo em Inglês | MEDLINE | ID: mdl-34285990

RESUMO

Studying lipid metabolism in cultured cells is complicated by the fact that cells are typically cultured in the presence of animal serum, which contains a wide, variable, and undefined variety of lipid species. Lipid metabolism can impact cell physiology, signaling, and proliferation, and the ability to culture cells in the absence of exogenous lipids can reveal the importance of lipid biosynthesis pathways and facilitate the generation of media with defined lipid species. We have adapted a protocol to remove lipids from serum without eliminating its ability to support the proliferation of cells in culture. This method requires di-isopropyl ether and butanol and can be used to generate small batches of lipid-stripped serum in four days. The resulting serum supports proliferation of many cell lines in culture and can be used to compare the metabolism of cells in lipid replete and depleted conditions.

18.
Dev Cell ; 40(2): 118-119, 2017 01 23.
Artigo em Inglês | MEDLINE | ID: mdl-28118598

RESUMO

Acetyl-CoA has diverse fates in metabolism and can be derived from a variety of nutrients. In a recent study published in Nature, Wong et al. (2016) show that endothelial cells oxidize fatty acids to produce acetyl-CoA for epigenetic modifications critical to lymphangiogenesis.


Assuntos
Acetilcoenzima A/metabolismo , Linfangiogênese , Células Endoteliais/metabolismo , Ácidos Graxos/metabolismo , Humanos , Oxirredução
19.
Cell Metab ; 24(5): 716-727, 2016 11 08.
Artigo em Inglês | MEDLINE | ID: mdl-27746050

RESUMO

Metformin use is associated with reduced cancer mortality, but how metformin impacts cancer outcomes is controversial. Although metformin can act on cells autonomously to inhibit tumor growth, the doses of metformin that inhibit proliferation in tissue culture are much higher than what has been described in vivo. Here, we show that the environment drastically alters sensitivity to metformin and other complex I inhibitors. We find that complex I supports proliferation by regenerating nicotinamide adenine dinucleotide (NAD)+, and metformin's anti-proliferative effect is due to loss of NAD+/NADH homeostasis and inhibition of aspartate biosynthesis. However, complex I is only one of many inputs that determines the cellular NAD+/NADH ratio, and dependency on complex I is dictated by the activity of other pathways that affect NAD+ regeneration and aspartate levels. This suggests that cancer drug sensitivity and resistance are not intrinsic properties of cancer cells, and demonstrates that the environment can dictate sensitivity to therapies that impact cell metabolism.


Assuntos
Ácido Aspártico/biossíntese , Complexo I de Transporte de Elétrons/metabolismo , Metformina/farmacologia , Mitocôndrias/metabolismo , NAD/metabolismo , Neoplasias/patologia , Microambiente Tumoral/efeitos dos fármacos , Animais , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Homeostase/efeitos dos fármacos , Humanos , Camundongos Nus , Mitocôndrias/efeitos dos fármacos , Ácido Pirúvico/farmacologia
20.
Science ; 353(6304): 1161-5, 2016 09 09.
Artigo em Inglês | MEDLINE | ID: mdl-27609895

RESUMO

Tumor genetics guides patient selection for many new therapies, and cell culture studies have demonstrated that specific mutations can promote metabolic phenotypes. However, whether tissue context defines cancer dependence on specific metabolic pathways is unknown. Kras activation and Trp53 deletion in the pancreas or the lung result in pancreatic ductal adenocarinoma (PDAC) or non-small cell lung carcinoma (NSCLC), respectively, but despite the same initiating events, these tumors use branched-chain amino acids (BCAAs) differently. NSCLC tumors incorporate free BCAAs into tissue protein and use BCAAs as a nitrogen source, whereas PDAC tumors have decreased BCAA uptake. These differences are reflected in expression levels of BCAA catabolic enzymes in both mice and humans. Loss of Bcat1 and Bcat2, the enzymes responsible for BCAA use, impairs NSCLC tumor formation, but these enzymes are not required for PDAC tumor formation, arguing that tissue of origin is an important determinant of how cancers satisfy their metabolic requirements.


Assuntos
Aminoácidos de Cadeia Ramificada/metabolismo , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Carcinoma Ductal Pancreático/genética , Carcinoma Ductal Pancreático/metabolismo , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/metabolismo , Proteínas Proto-Oncogênicas p21(ras)/genética , Animais , Regulação Neoplásica da Expressão Gênica , Humanos , Masculino , Redes e Vias Metabólicas , Camundongos , Camundongos Endogâmicos C57BL , Antígenos de Histocompatibilidade Menor/genética , Mutação , Nitrogênio/metabolismo , Especificidade de Órgãos , Proteínas da Gravidez/genética , Transaminases/genética
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