Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 43.042
Filtrar
Mais filtros

Intervalo de ano de publicação
1.
Nat Immunol ; 24(1): 162-173, 2023 01.
Artigo em Inglês | MEDLINE | ID: mdl-36471170

RESUMO

Amino acid metabolism is essential for cell survival, while the byproduct ammonia is toxic and can injure cellular longevity. Here we show that CD8+ memory T (TM) cells mobilize the carbamoyl phosphate (CP) metabolic pathway to clear ammonia, thus promoting memory development. CD8+ TM cells use ß-hydroxybutyrylation to upregulate CP synthetase 1 and trigger the CP metabolic cascade to form arginine in the cytosol. This cytosolic arginine is then translocated into the mitochondria where it is split by arginase 2 to urea and ornithine. Cytosolic arginine is also converted to nitric oxide and citrulline by nitric oxide synthases. Thus, both the urea and citrulline cycles are employed by CD8+ T cells to clear ammonia and enable memory development. This ammonia clearance machinery might be targeted to improve T cell-based cancer immunotherapies.


Assuntos
Amônia , Citrulina , Citrulina/metabolismo , Amônia/metabolismo , Ureia/metabolismo , Linfócitos T CD8-Positivos/metabolismo , Óxido Nítrico , Arginina/metabolismo , Arginase/metabolismo
2.
Cell ; 174(6): 1344-1346, 2018 09 06.
Artigo em Inglês | MEDLINE | ID: mdl-30193108

RESUMO

Altered cell metabolism is ubiquitous in cancer cells; however, it remains challenging to exploit these alterations for cancer therapy. A new study reveals that metabolic alterations to the urea cycle promote tumor growth but unexpectedly also trigger mutations that mark cancer cells for recognition by immunotherapy.


Assuntos
Imunoterapia , Neoplasias , Genômica , Humanos , Ureia
3.
Cell ; 174(6): 1559-1570.e22, 2018 09 06.
Artigo em Inglês | MEDLINE | ID: mdl-30100185

RESUMO

The urea cycle (UC) is the main pathway by which mammals dispose of waste nitrogen. We find that specific alterations in the expression of most UC enzymes occur in many tumors, leading to a general metabolic hallmark termed "UC dysregulation" (UCD). UCD elicits nitrogen diversion toward carbamoyl-phosphate synthetase2, aspartate transcarbamylase, and dihydrooratase (CAD) activation and enhances pyrimidine synthesis, resulting in detectable changes in nitrogen metabolites in both patient tumors and their bio-fluids. The accompanying excess of pyrimidine versus purine nucleotides results in a genomic signature consisting of transversion mutations at the DNA, RNA, and protein levels. This mutational bias is associated with increased numbers of hydrophobic tumor antigens and a better response to immune checkpoint inhibitors independent of mutational load. Taken together, our findings demonstrate that UCD is a common feature of tumors that profoundly affects carcinogenesis, mutagenesis, and immunotherapy response.


Assuntos
Genômica , Metabolômica , Neoplasias/patologia , Ureia/metabolismo , Sistemas de Transporte de Aminoácidos Básicos/metabolismo , Animais , Aspartato Carbamoiltransferase/genética , Aspartato Carbamoiltransferase/metabolismo , Carbamoil Fosfato Sintase (Glutamina-Hidrolizante)/genética , Carbamoil Fosfato Sintase (Glutamina-Hidrolizante)/metabolismo , Linhagem Celular Tumoral , Di-Hidro-Orotase/genética , Di-Hidro-Orotase/metabolismo , Feminino , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos SCID , Proteínas de Transporte da Membrana Mitocondrial , Neoplasias/metabolismo , Ornitina Carbamoiltransferase/antagonistas & inibidores , Ornitina Carbamoiltransferase/genética , Ornitina Carbamoiltransferase/metabolismo , Fosforilação/efeitos dos fármacos , Pirimidinas/biossíntese , Pirimidinas/química , Interferência de RNA , RNA Interferente Pequeno/metabolismo , Sirolimo/farmacologia , Serina-Treonina Quinases TOR/antagonistas & inibidores , Serina-Treonina Quinases TOR/metabolismo
4.
Nature ; 619(7971): 749-754, 2023 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-37380782

RESUMO

Proton transfer is one of the most fundamental events in aqueous-phase chemistry and an emblematic case of coupled ultrafast electronic and structural dynamics1,2. Disentangling electronic and nuclear dynamics on the femtosecond timescales remains a formidable challenge, especially in the liquid phase, the natural environment of biochemical processes. Here we exploit the unique features of table-top water-window X-ray absorption spectroscopy3-6 to reveal femtosecond proton-transfer dynamics in ionized urea dimers in aqueous solution. Harnessing the element specificity and the site selectivity of X-ray absorption spectroscopy with the aid of ab initio quantum-mechanical and molecular-mechanics calculations, we show how, in addition to the proton transfer, the subsequent rearrangement of the urea dimer and the associated change of the electronic structure can be identified with site selectivity. These results establish the considerable potential of flat-jet, table-top X-ray absorption spectroscopy7,8 in elucidating solution-phase ultrafast dynamics in biomolecular systems.


Assuntos
Prótons , Ureia , Ureia/química , Soluções/química , Água/química , Espectroscopia por Absorção de Raios X , Teoria Quântica , Fatores de Tempo
5.
Nature ; 621(7979): 511-515, 2023 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-37553075

RESUMO

Plywood is widely used in construction, such as for flooring and interior walls, as well as in the manufacture of household items such as furniture and cabinets. Such items are made of wood veneers that are bonded together with adhesives such as urea-formaldehyde and phenol-formaldehyde resins1,2. Researchers in academia and industry have long aimed to synthesize lignin-phenol-formaldehyde resin adhesives using biomass-derived lignin, a phenolic polymer that can be used to substitute the petroleum-derived phenol3-6. However, lignin-phenol-formaldehyde resin adhesives are less attractive to plywood manufacturers than urea-formaldehyde and phenol-formaldehyde resins owing to their appearance and cost. Here we report a simple and practical strategy for preparing lignin-based wood adhesives from lignocellulosic biomass. Our strategy involves separation of uncondensed or slightly condensed lignins from biomass followed by direct application of a suspension of the lignin and water as an adhesive on wood veneers. Plywood products with superior performances could be prepared with such lignin adhesives at a wide range of hot-pressing temperatures, enabling the use of these adhesives as promising alternatives to traditional wood adhesives in different market segments. Mechanistic studies indicate that the adhesion mechanism of such lignin adhesives may involve softening of lignin by water, filling of vessels with softened lignin and crosslinking of lignins in adhesives with those in the cell wall.


Assuntos
Adesivos , Lignina , Madeira , Adesivos/química , Formaldeído/química , Lignina/química , Fenóis/química , Ureia/química , Água/química , Madeira/química , Biomassa , Temperatura Alta
6.
Mol Cell ; 81(18): 3749-3759, 2021 09 16.
Artigo em Inglês | MEDLINE | ID: mdl-34469752

RESUMO

The expression of the urea cycle (UC) proteins is dysregulated in multiple cancers, providing metabolic benefits to tumor survival, proliferation, and growth. Here, we review the main changes described in the expression of UC enzymes and metabolites in different cancers at various stages and suggest that these changes are dynamic and should hence be viewed in a context-specific manner. Understanding the evolvability in the activity of the UC pathway in cancer has implications for cancer-immune cell interactions and for cancer diagnosis and therapy.


Assuntos
Carcinogênese/metabolismo , Transformação Celular Neoplásica/metabolismo , Ureia/metabolismo , Amônia/metabolismo , Linhagem Celular Tumoral , Proliferação de Células , Expressão Gênica/genética , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Distúrbios Congênitos do Ciclo da Ureia/metabolismo , Distúrbios Congênitos do Ciclo da Ureia/fisiopatologia
7.
Immunity ; 51(6): 975-977, 2019 12 17.
Artigo em Inglês | MEDLINE | ID: mdl-31951542

RESUMO

Integrating transcriptomic, proteomic, and metabolomic data, Lercher et al. show in a mouse model of LCMV infection that type I interferon alters the expression and function of key enzymes of the urea cycle in hepatocytes. This results in altered systemic metabolism, attenuating antiviral T cell responses and ameliorating liver injury.


Assuntos
Antivirais , Interferon Tipo I , Animais , Fígado , Vírus da Coriomeningite Linfocítica , Camundongos , Proteômica , Linfócitos T , Ureia
8.
Immunity ; 51(6): 1074-1087.e9, 2019 12 17.
Artigo em Inglês | MEDLINE | ID: mdl-31784108

RESUMO

Infections induce complex host responses linked to antiviral defense, inflammation, and tissue damage and repair. We hypothesized that the liver, as a central metabolic hub, may orchestrate systemic metabolic changes during infection. We infected mice with chronic lymphocytic choriomeningitis virus (LCMV), performed RNA sequencing and proteomics of liver tissue, and integrated these data with serum metabolomics at different infection phases. Widespread reprogramming of liver metabolism occurred early after infection, correlating with type I interferon (IFN-I) responses. Viral infection induced metabolic alterations of the liver that depended on the interferon alpha/beta receptor (IFNAR1). Hepatocyte-intrinsic IFNAR1 repressed the transcription of metabolic genes, including Otc and Ass1, which encode urea cycle enzymes. This led to decreased arginine and increased ornithine concentrations in the circulation, resulting in suppressed virus-specific CD8+ T cell responses and ameliorated liver pathology. These findings establish IFN-I-induced modulation of hepatic metabolism and the urea cycle as an endogenous mechanism of immunoregulation. VIDEO ABSTRACT.


Assuntos
Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Interferon Tipo I/imunologia , Fígado/metabolismo , Vírus da Coriomeningite Linfocítica/imunologia , Receptor de Interferon alfa e beta/metabolismo , Animais , Arginina/sangue , Linhagem Celular , Chlorocebus aethiops , Cricetinae , Feminino , Hepatócitos/metabolismo , Fígado/imunologia , Fígado/virologia , Coriomeningite Linfocítica/imunologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Ornitina/sangue , Ornitina Carbamoiltransferase/genética , Transdução de Sinais/imunologia , Ureia/metabolismo , Células Vero
9.
Am J Hum Genet ; 111(4): 714-728, 2024 04 04.
Artigo em Inglês | MEDLINE | ID: mdl-38579669

RESUMO

Argininosuccinate lyase deficiency (ASLD) is a recessive metabolic disorder caused by variants in ASL. In an essential step in urea synthesis, ASL breaks down argininosuccinate (ASA), a pathognomonic ASLD biomarker. The severe disease forms lead to hyperammonemia, neurological injury, and even early death. The current treatments are unsatisfactory, involving a strict low-protein diet, arginine supplementation, nitrogen scavenging, and in some cases, liver transplantation. An unmet need exists for improved, efficient therapies. Here, we show the potential of a lipid nanoparticle-mediated CRISPR approach using adenine base editors (ABEs) for ASLD treatment. To model ASLD, we first generated human-induced pluripotent stem cells (hiPSCs) from biopsies of individuals homozygous for the Finnish founder variant (c.1153C>T [p.Arg385Cys]) and edited this variant using the ABE. We then differentiated the hiPSCs into hepatocyte-like cells that showed a 1,000-fold decrease in ASA levels compared to those of isogenic non-edited cells. Lastly, we tested three different FDA-approved lipid nanoparticle formulations to deliver the ABE-encoding RNA and the sgRNA targeting the ASL variant. This approach efficiently edited the ASL variant in fibroblasts with no apparent cell toxicity and minimal off-target effects. Further, the treatment resulted in a significant decrease in ASA, to levels of healthy donors, indicating restoration of the urea cycle. Our work describes a highly efficient approach to editing the disease-causing ASL variant and restoring the function of the urea cycle. This method relies on RNA delivered by lipid nanoparticles, which is compatible with clinical applications, improves its safety profile, and allows for scalable production.


Assuntos
Argininossuccinato Liase , Acidúria Argininossuccínica , Humanos , Argininossuccinato Liase/genética , Acidúria Argininossuccínica/genética , Acidúria Argininossuccínica/terapia , Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas , RNA Guia de Sistemas CRISPR-Cas , Ureia , Edição de Genes/métodos
10.
Proc Natl Acad Sci U S A ; 121(14): e2317825121, 2024 Apr 02.
Artigo em Inglês | MEDLINE | ID: mdl-38536756

RESUMO

Trimethylamine-N-oxide (TMAO) and urea are metabolites that are used by some marine animals to maintain their cell volume in a saline environment. Urea is a well-known denaturant, and TMAO is a protective osmolyte that counteracts urea-induced protein denaturation. TMAO also has a general protein-protective effect, for example, it counters pressure-induced protein denaturation in deep-sea fish. These opposing effects on protein stability have been linked to the spatial relationship of TMAO, urea, and protein molecules. It is generally accepted that urea-induced denaturation proceeds through the accumulation of urea at the protein surface and their subsequent interaction. In contrast, it has been suggested that TMAO's protein-stabilizing effects stem from its exclusion from the protein surface, and its ability to deplete urea from protein surfaces; however, these spatial relationships are uncertain. We used neutron diffraction, coupled with structural refinement modeling, to study the spatial associations of TMAO and urea with the tripeptide derivative glycine-proline-glycinamide in aqueous urea, aqueous TMAO, and aqueous urea-TMAO (in the mole ratio 1:2 TMAO:urea). We found that TMAO depleted urea from the peptide's surface and that while TMAO was not excluded from the tripeptide's surface, strong atomic interactions between the peptide and TMAO were limited to hydrogen bond donating peptide groups. We found that the repartition of urea, by TMAO, was associated with preferential TMAO-urea bonding and enhanced urea-water hydrogen bonding, thereby anchoring urea in the bulk solution and depleting urea from the peptide surface.


Assuntos
Peptídeos , Ureia , Animais , Ureia/química , Peptídeos/química , Água/química , Metilaminas/química , Proteínas de Membrana
11.
Proc Natl Acad Sci U S A ; 121(10): e2312652121, 2024 Mar 05.
Artigo em Inglês | MEDLINE | ID: mdl-38408229

RESUMO

Metformin is the first-line treatment for type II diabetes patients and a pervasive pollutant with more than 180 million kg ingested globally and entering wastewater. The drug's direct mode of action is currently unknown but is linked to effects on gut microbiomes and may involve specific gut microbial reactions to the drug. In wastewater treatment plants, metformin is known to be transformed by microbes to guanylurea, although genes encoding this metabolism had not been elucidated. In the present study, we revealed the function of two genes responsible for metformin decomposition (mfmA and mfmB) found in isolated bacteria from activated sludge. MfmA and MfmB form an active heterocomplex (MfmAB) and are members of the ureohydrolase protein superfamily with binuclear metal-dependent activity. MfmAB is nickel-dependent and catalyzes the hydrolysis of metformin to dimethylamine and guanylurea with a catalytic efficiency (kcat/KM) of 9.6 × 103 M-1s-1 and KM for metformin of 0.82 mM. MfmAB shows preferential activity for metformin, being able to discriminate other close substrates by several orders of magnitude. Crystal structures of MfmAB show coordination of binuclear nickel bound in the active site of the MfmA subunit but not MfmB subunits, indicating that MfmA is the active site for the MfmAB complex. Mutagenesis of residues conserved in the MfmA active site revealed those critical to metformin hydrolase activity and its small substrate binding pocket allowed for modeling of bound metformin. This study characterizes the products of the mfmAB genes identified in wastewater treatment plants on three continents, suggesting that metformin hydrolase is widespread globally in wastewater.


Assuntos
Diabetes Mellitus Tipo 2 , Guanidina/análogos & derivados , Metformina , Microbiota , Ureia/análogos & derivados , Humanos , Metformina/metabolismo , Águas Residuárias , Níquel , Hidrolases/genética , Preparações Farmacêuticas
12.
Nature ; 581(7808): 339-343, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32433613

RESUMO

Cholesterol is an essential component of mammalian cell membranes, constituting up to 50% of plasma membrane lipids. By contrast, it accounts for only 5% of lipids in the endoplasmic reticulum (ER)1. The ER enzyme sterol O-acyltransferase 1 (also named acyl-coenzyme A:cholesterol acyltransferase, ACAT1) transfers a long-chain fatty acid to cholesterol to form cholesteryl esters that coalesce into cytosolic lipid droplets. Under conditions of cholesterol overload, ACAT1 maintains the low cholesterol concentration of the ER and thereby has an essential role in cholesterol homeostasis2,3. ACAT1 has also been implicated in Alzheimer's disease4, atherosclerosis5 and cancers6. Here we report a cryo-electron microscopy structure of human ACAT1 in complex with nevanimibe7, an inhibitor that is in clinical trials for the treatment of congenital adrenal hyperplasia. The ACAT1 holoenzyme is a tetramer that consists of two homodimers. Each monomer contains nine transmembrane helices (TMs), six of which (TM4-TM9) form a cavity that accommodates nevanimibe and an endogenous acyl-coenzyme A. This cavity also contains a histidine that has previously been identified as essential for catalytic activity8. Our structural data and biochemical analyses provide a physical model to explain the process of cholesterol esterification, as well as details of the interaction between nevanimibe and ACAT1, which may help to accelerate the development of ACAT1 inhibitors to treat related diseases.


Assuntos
Microscopia Crioeletrônica , Esterol O-Aciltransferase/química , Esterol O-Aciltransferase/ultraestrutura , Ureia/análogos & derivados , Colesterol/química , Colesterol/metabolismo , Histidina/química , Histidina/metabolismo , Holoenzimas/química , Holoenzimas/ultraestrutura , Humanos , Ligantes , Modelos Moleculares , Multimerização Proteica , Eletricidade Estática , Ureia/química
13.
J Biol Chem ; 300(8): 107495, 2024 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-38925327

RESUMO

Transthyretin (TTR) is an homotetrameric protein involved in the transport of thyroxine. More than 150 different mutations have been described in the TTR gene, several of them associated with familial amyloid cardiomyopathy. Recently, our group described a new variant of TTR in Brazil, namely A39D-TTR, which causes a severe cardiac condition. Position 39 is in the AB loop, a region of the protein that is located within the thyroxine-binding channels and is involved in tetramer formation. In the present study, we solved the structure and characterize the thermodynamic stability of this new variant of TTR using urea and high hydrostatic pressure. Interestingly, during the process of purification, A39D-TTR turned out to be a dimer and not a tetramer, a variation that might be explained by the close contact of the four aspartic acids at position 39, where they face each other inside the thyroxine channel. In the presence of subdenaturing concentrations of urea, bis-ANS binding and dynamic light scattering revealed A39D-TTR in the form of a molten-globule dimer. Co-expression of A39D and WT isoforms in the same bacterial cell did not produce heterodimers or heterotetramers, suggesting that somehow a negative charge at the AB loop precludes tetramer formation. A39D-TTR proved to be highly amyloidogenic, even at mildly acidic pH values where WT-TTR does not aggregate. Interestingly, despite being a dimer, aggregation of A39D-TTR was inhibited by diclofenac, which binds to the thyroxine channel in the tetramer, suggesting the existence of other pockets in A39D-TTR able to accommodate this molecule.


Assuntos
Cardiomiopatias , Pré-Albumina , Multimerização Proteica , Termodinâmica , Pré-Albumina/genética , Pré-Albumina/química , Pré-Albumina/metabolismo , Humanos , Cardiomiopatias/metabolismo , Cardiomiopatias/genética , Tiroxina/metabolismo , Tiroxina/química , Mutação de Sentido Incorreto , Amiloide/metabolismo , Amiloide/química , Amiloide/genética , Substituição de Aminoácidos , Ureia/química , Ureia/metabolismo
14.
Physiol Rev ; 98(2): 641-665, 2018 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-29412048

RESUMO

The arginase enzyme developed in early life forms and was maintained during evolution. As the last step in the urea cycle, arginase cleaves l-arginine to form urea and l-ornithine. The urea cycle provides protection against excess ammonia, while l-ornithine is needed for cell proliferation, collagen formation, and other physiological functions. In mammals, increases in arginase activity have been linked to dysfunction and pathologies of the cardiovascular system, kidney, and central nervous system and also to dysfunction of the immune system and cancer. Two important aspects of the excessive activity of arginase may be involved in diseases. First, overly active arginase can reduce the supply of l-arginine needed for the production of nitric oxide (NO) by NO synthase. Second, too much l-ornithine can lead to structural problems in the vasculature, neuronal toxicity, and abnormal growth of tumor cells. Seminal studies have demonstrated that increased formation of reactive oxygen species and key inflammatory mediators promote this pathological elevation of arginase activity. Here, we review the involvement of arginase in diseases affecting the cardiovascular, renal, and central nervous system and cancer and discuss the value of therapies targeting the elevated activity of arginase.


Assuntos
Arginase/metabolismo , Endotélio Vascular/metabolismo , Óxido Nítrico/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Ureia/metabolismo , Animais , Arginina/metabolismo , Endotélio Vascular/fisiopatologia , Humanos
15.
Hum Mol Genet ; 32(11): 1922-1931, 2023 05 18.
Artigo em Inglês | MEDLINE | ID: mdl-36881658

RESUMO

Citrin deficiency (CD) is an inborn error of metabolism caused by loss-of-function of the mitochondrial aspartate/glutamate transporter, CITRIN, which is involved in both the urea cycle and malate-aspartate shuttle. Patients with CD develop hepatosteatosis and hyperammonemia but there is no effective therapy for CD. Currently, there are no animal models that faithfully recapitulate the human CD phenotype. Accordingly, we generated a CITRIN knockout HepG2 cell line using Clustered Regularly Interspaced Short Palindromic Repeats/Cas 9 genome editing technology to study metabolic and cell signaling defects in CD. CITRIN KO cells showed increased ammonia accumulation, higher cytosolic ratio of reduced versus oxidized form of nicotinamide adenine dinucleotide (NAD) and reduced glycolysis. Surprisingly, these cells showed impaired fatty acid metabolism and mitochondrial activity. CITRIN KO cells also displayed increased cholesterol and bile acid metabolism resembling those observed in CD patients. Remarkably, normalizing cytosolic NADH:NAD+ ratio by nicotinamide riboside increased glycolysis and fatty acid oxidation but had no effect on the hyperammonemia suggesting the urea cycle defect was independent of the aspartate/malate shuttle defect of CD. The correction of glycolysis and fatty acid metabolism defects in CITRIN KO cells by reducing cytoplasmic NADH:NAD+ levels suggests this may be a novel strategy to treat some of the metabolic defects of CD and other mitochondrial diseases.


Assuntos
Citrulinemia , Hiperamonemia , Humanos , Citrulinemia/genética , Citrulinemia/metabolismo , NAD/metabolismo , Malatos , Ácido Aspártico/metabolismo , Hiperamonemia/genética , Proteínas de Transporte da Membrana Mitocondrial/genética , Hepatócitos/metabolismo , Glicólise , Ureia/metabolismo , Ácidos Graxos
16.
Development ; 149(23)2022 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-36355069

RESUMO

Upon WNT/ß-catenin pathway activation, stabilized ß-catenin travels to the nucleus where it associates with the TCF/LEF transcription factors, constitutively bound to genomic Wnt-responsive elements (WREs), to activate target gene transcription. Discovering the binding profile of ß-catenin is therefore required to unambiguously assign direct targets of WNT signaling. Cleavage under targets and release using nuclease (CUT&RUN) has emerged as prime technique for mapping the binding profile of DNA-interacting proteins. Here, we present a modified version of CUT&RUN, named LoV-U (low volume and urea), that enables the robust and reproducible generation of ß-catenin binding profiles, uncovering direct WNT/ß-catenin target genes in human cells, as well as in cells isolated from developing mouse tissues. CUT&RUN-LoV-U outperforms original CUT&RUN when targeting co-factors that do not bind the DNA, can profile all classes of chromatin regulators and is well suited for simultaneous processing of several samples. We believe that the application of our protocol will allow the detection of the complex system of tissue-specific WNT/ß-catenin target genes, together with other non-DNA-binding transcriptional regulators that act downstream of ontogenetically fundamental signaling cascades.


Assuntos
Fatores de Transcrição , beta Catenina , Humanos , Camundongos , Animais , beta Catenina/genética , beta Catenina/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Fatores de Transcrição TCF/metabolismo , Via de Sinalização Wnt/genética , Endonucleases/metabolismo , Genômica , Ureia , Ativação Transcricional
17.
Nature ; 573(7772): 102-107, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31485055

RESUMO

Amides and related carbonyl derivatives are of central importance across the physical and life sciences1,2. As a key biological building block, the stability and conformation of amides affect the structures of peptides and proteins as well as their biological function. In addition, amide-bond formation is one of the most frequently used chemical transformations3,4. Given their ubiquity, a technology that is capable of modifying the fundamental properties of amides without compromising on stability may have considerable potential in pharmaceutical, agrochemical and materials science. In order to influence the physical properties of organic molecules-such as solubility, lipophilicity, conformation, pKa and (metabolic) stability-fluorination approaches have been widely adopted5-7. Similarly, site-specific modification with isosteres and peptidomimetics8, or in particular by N-methylation9, has been used to improve the stability, physical properties, bioactivities and cellular permeabilities of compounds. However, the N-trifluoromethyl carbonyl motif-which combines both N-methylation and fluorination approaches-has not yet been explored, owing to a lack of efficient methodology to synthesize it. Here we report a straightforward method to access N-trifluoromethyl analogues of amides and related carbonyl compounds. The strategy relies on the operationally simple preparation of bench-stable carbamoyl fluoride building blocks, which can be readily diversified to the corresponding N-CF3 amides, carbamates, thiocarbamates and ureas. This method tolerates rich functionality and stereochemistry, and we present numerous examples of highly functionalized compounds-including analogues of widely used drugs, antibiotics, hormones and polymer units.


Assuntos
Amidas/química , Carbamatos/química , Ureia/análogos & derivados , Ureia/química , Fluoretos/química , Isotiocianatos/química , Preparações Farmacêuticas/síntese química , Preparações Farmacêuticas/química
18.
Nature ; 567(7747): 253-256, 2019 03.
Artigo em Inglês | MEDLINE | ID: mdl-30842655

RESUMO

Cancer cells exhibit altered and usually increased metabolic processes to meet their high biogenetic demands1,2. Under these conditions, ammonia is concomitantly produced by the increased metabolic processing. However, it is unclear how tumour cells dispose of excess ammonia and what outcomes might be caused by the accumulation of ammonia. Here we report that the tumour suppressor p53, the most frequently mutated gene in human tumours, regulates ammonia metabolism by repressing the urea cycle. Through transcriptional downregulation of CPS1, OTC and ARG1, p53 suppresses ureagenesis and elimination of ammonia in vitro and in vivo, leading to the inhibition of tumour growth. Conversely, downregulation of these genes reciprocally activates p53 by MDM2-mediated mechanism(s). Furthermore, the accumulation of ammonia causes a significant decline in mRNA translation of the polyamine biosynthetic rate-limiting enzyme ODC, thereby inhibiting the biosynthesis of polyamine and cell proliferation. Together, these findings link p53 to ureagenesis and ammonia metabolism, and further reveal a role for ammonia in controlling polyamine biosynthesis and cell proliferation.


Assuntos
Amônia/metabolismo , Regulação da Expressão Gênica/genética , Poliaminas/metabolismo , Proteína Supressora de Tumor p53/metabolismo , Ureia/metabolismo , Arginase/genética , Carbamoil-Fosfato Sintase (Amônia)/genética , Proliferação de Células , Humanos , Neoplasias/genética , Neoplasias/patologia , Ornitina Carbamoiltransferase/genética , Ornitina Descarboxilase/biossíntese , Ornitina Descarboxilase/genética , Proteínas Proto-Oncogênicas c-mdm2/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Transcrição Gênica/genética
19.
Mol Cell ; 68(1): 198-209.e6, 2017 Oct 05.
Artigo em Inglês | MEDLINE | ID: mdl-28985504

RESUMO

In addition to responding to environmental entrainment with diurnal variation, metabolism is also tightly controlled by cell-autonomous circadian clock. Extensive studies have revealed key roles of transcription in circadian control. Post-transcriptional regulation for the rhythmic gating of metabolic enzymes remains elusive. Here, we show that arginine biosynthesis and subsequent ureagenesis are collectively regulated by CLOCK (circadian locomotor output cycles kaput) in circadian rhythms. Facilitated by BMAL1 (brain and muscle Arnt-like protein), CLOCK directly acetylates K165 and K176 of argininosuccinate synthase (ASS1) to inactivate ASS1, which catalyzes the rate-limiting step of arginine biosynthesis. ASS1 acetylation by CLOCK exhibits circadian oscillation in human cells and mouse liver, possibly caused by rhythmic interaction between CLOCK and ASS1, leading to the circadian regulation of ASS1 and ureagenesis. Furthermore, we also identified NADH dehydrogenase [ubiquinone] 1 alpha subcomplex subunit 9 (NDUFA9) and inosine-5'-monophosphate dehydrogenase 2 (IMPDH2) as acetylation substrates of CLOCK. Taken together, CLOCK modulates metabolic rhythmicity by acting as a rhythmic acetyl-transferase for metabolic enzymes.


Assuntos
Fatores de Transcrição ARNTL/genética , Argininossuccinato Sintase/genética , Proteínas CLOCK/genética , Ritmo Circadiano/genética , Processamento de Proteína Pós-Traducional , Ureia/metabolismo , Fatores de Transcrição ARNTL/metabolismo , Acetilação , Animais , Arginina/biossíntese , Argininossuccinato Sintase/metabolismo , Proteínas CLOCK/metabolismo , Linhagem Celular Tumoral , Relógios Circadianos , Complexo I de Transporte de Elétrons/genética , Complexo I de Transporte de Elétrons/metabolismo , Células HEK293 , Hepatócitos/citologia , Hepatócitos/metabolismo , Humanos , IMP Desidrogenase/genética , IMP Desidrogenase/metabolismo , Masculino , Camundongos , Camundongos Knockout , Osteoblastos/metabolismo , Osteoblastos/patologia , Transdução de Sinais
20.
Nucleic Acids Res ; 51(8): 3754-3769, 2023 05 08.
Artigo em Inglês | MEDLINE | ID: mdl-37014002

RESUMO

The N-(2-deoxy-d-erythro-pentofuranosyl)-urea DNA lesion forms following hydrolytic fragmentation of cis-5R,6S- and trans-5R,6R-dihydroxy-5,6-dihydrothymidine (thymine glycol, Tg) or from oxidation of 7,8-dihydro-8-oxo-deoxyguanosine (8-oxodG) and subsequent hydrolysis. It interconverts between α and ß deoxyribose anomers. Synthetic oligodeoxynucleotides containing this adduct are efficiently incised by unedited (K242) and edited (R242) forms of the hNEIL1 glycosylase. The structure of a complex between the active site unedited mutant CΔ100 P2G hNEIL1 (K242) glycosylase and double-stranded (ds) DNA containing a urea lesion reveals a pre-cleavage intermediate, in which the Gly2 N-terminal amine forms a conjugate with the deoxyribose C1' of the lesion, with the urea moiety remaining intact. This structure supports a proposed catalytic mechanism in which Glu3-mediated protonation of O4' facilitates attack at deoxyribose C1'. The deoxyribose is in the ring-opened configuration with the O4' oxygen protonated. The electron density of Lys242 suggests the 'residue 242-in conformation' associated with catalysis. This complex likely arises because the proton transfer steps involving Glu6 and Lys242 are hindered due to Glu6-mediated H-bonding with the Gly2 and the urea lesion. Consistent with crystallographic data, biochemical analyses show that the CΔ100 P2G hNEIL1 (K242) glycosylase exhibits a residual activity against urea-containing dsDNA.


Assuntos
DNA Glicosilases , Reparo do DNA , Desoxirribose , Ureia , Desoxirribose/química , DNA/química , Dano ao DNA , DNA Glicosilases/metabolismo , Humanos
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA