RESUMO
ATP, NAD+, and nucleic acids are abundant purines that, in addition to having critical intracellular functions, have evolved extracellular roles as danger signals released in response to cell lysis, apoptosis, degranulation, or membrane pore formation. In general ATP and NAD+ have excitatory and adenosine has anti-inflammatory effects on immune cells. This review focuses on recent advances in our understanding of purine release mechanisms, ectoenzymes that metabolize purines (CD38, CD39, CD73, ENPP1, and ENPP2/autotaxin), and signaling by key P2 purinergic receptors (P2X7, P2Y2, and P2Y12). In addition to metabolizing ATP or NAD+, some purinergic ectoenzymes metabolize other inflammatory modulators, notably lysophosphatidic acid and cyclic GMP-AMP (cGAMP). Also discussed are extracellular signaling effects of NAD+ mediated by ADP-ribosylation, and epigenetic effects of intracellular adenosine mediated by modification of S-adenosylmethionine-dependent DNA methylation.
Assuntos
Inflamação/imunologia , Purinas/metabolismo , Receptores Purinérgicos/metabolismo , ADP-Ribosilação , Trifosfato de Adenosina/metabolismo , Animais , Metilação de DNA , Humanos , Inflamação/genética , Inflamação/metabolismo , Lisofosfolipídeos/metabolismo , Transdução de SinaisRESUMO
Purine nucleotides are vital for RNA and DNA synthesis, signaling, metabolism, and energy homeostasis. To synthesize purines, cells use two principal routes: the de novo and salvage pathways. Traditionally, it is believed that proliferating cells predominantly rely on de novo synthesis, whereas differentiated tissues favor the salvage pathway. Unexpectedly, we find that adenine and inosine are the most effective circulating precursors for supplying purine nucleotides to tissues and tumors, while hypoxanthine is rapidly catabolized and poorly salvaged in vivo. Quantitative metabolic analysis demonstrates comparative contribution from de novo synthesis and salvage pathways in maintaining purine nucleotide pools in tumors. Notably, feeding mice nucleotides accelerates tumor growth, while inhibiting purine salvage slows down tumor progression, revealing a crucial role of the salvage pathway in tumor metabolism. These findings provide fundamental insights into how normal tissues and tumors maintain purine nucleotides and highlight the significance of purine salvage in cancer.
Assuntos
Neoplasias , Nucleotídeos de Purina , Purinas , Animais , Camundongos , Purinas/metabolismo , Purinas/biossíntese , Neoplasias/metabolismo , Neoplasias/patologia , Nucleotídeos de Purina/metabolismo , Humanos , Inosina/metabolismo , Hipoxantina/metabolismo , Camundongos Endogâmicos C57BL , Adenina/metabolismo , Linhagem Celular Tumoral , FemininoRESUMO
Over the past fifteen years, we have unveiled a new mechanism by which cells achieve greater efficiency in de novo purine biosynthesis. This mechanism relies on the compartmentalization of de novo purine biosynthetic enzymes into a dynamic complex called the purinosome. In this review, we highlight our current understanding of the purinosome with emphasis on its biophysical properties and function and on the cellular mechanisms that regulate its assembly. We propose a model for functional purinosomes in which they consist of at least ten enzymes that localize near mitochondria and carry out de novo purine biosynthesis by metabolic channeling. We conclude by discussing challenges and opportunities associated with studying the purinosome and analogous metabolons.
Assuntos
Mitocôndrias , Purinas , Animais , Mamíferos/metabolismo , Mitocôndrias/genética , Mitocôndrias/metabolismo , Purinas/metabolismoRESUMO
Upon stress, eukaryotes typically reprogram their translatome through GCN2-mediated phosphorylation of the eukaryotic translation initiation factor, eIF2α, to inhibit general translation initiation while selectively translating essential stress regulators. Unexpectedly, in plants, pattern-triggered immunity (PTI) and response to other environmental stresses occur independently of the GCN2/eIF2α pathway. Here, we show that while PTI induces mRNA decapping to inhibit general translation, defense mRNAs with a purine-rich element ("R-motif") are selectively translated using R-motif as an internal ribosome entry site (IRES). R-motif-dependent translation is executed by poly(A)-binding proteins (PABPs) through preferential association with the PTI-activating eIFiso4G over the repressive eIF4G. Phosphorylation by PTI regulators mitogen-activated protein kinase 3 and 6 (MPK3/6) inhibits eIF4G's activity while enhancing PABP binding to the R-motif and promoting eIFiso4G-mediated defense mRNA translation, establishing a link between PTI signaling and protein synthesis. Given its prevalence in both plants and animals, the PABP/R-motif translation initiation module may have a broader role in reprogramming the stress translatome.
Assuntos
Fator de Iniciação Eucariótico 4G , Proteínas de Ligação a Poli(A) , Animais , Fator de Iniciação Eucariótico 4G/genética , Fator de Iniciação Eucariótico 4G/metabolismo , Fatores de Iniciação em Eucariotos/metabolismo , Proteínas de Ligação a Poli(A)/metabolismo , Biossíntese de Proteínas , Purinas , RNA Mensageiro/metabolismoRESUMO
Mitochondrial DNA (mtDNA) editing paves the way for disease modeling of mitochondrial genetic disorders in cell lines and animals and also for the treatment of these diseases in the future. Bacterial cytidine deaminase DddA-derived cytosine base editors (DdCBEs) enabling mtDNA editing, however, are largely limited to C-to-T conversions in the 5'-TC context (e.g., TC-to-TT conversions), suitable for generating merely 1/8 of all possible transition (purine-to-purine and pyrimidine-to-pyrimidine) mutations. Here, we present transcription-activator-like effector (TALE)-linked deaminases (TALEDs), composed of custom-designed TALE DNA-binding arrays, a catalytically impaired, full-length DddA variant or split DddA originated from Burkholderia cenocepacia, and an engineered deoxyadenosine deaminase derived from the E. coli TadA protein, which induce targeted A-to-G editing in human mitochondria. Custom-designed TALEDs were highly efficient in human cells, catalyzing A-to-G conversions at a total of 17 target sites in various mitochondrial genes with editing frequencies of up to 49%.
Assuntos
DNA Mitocondrial , Doenças Mitocondriais , Animais , Sistemas CRISPR-Cas , Citosina/metabolismo , DNA Mitocondrial/genética , Escherichia coli/genética , Escherichia coli/metabolismo , Edição de Genes , Humanos , Mitocôndrias/genética , Mitocôndrias/metabolismo , Doenças Mitocondriais/genética , PurinasRESUMO
I endeavor to share how various choices-some deliberate, some unconscious-and the unmistakable influence of many others shaped my scientific pursuits. I am fascinated by how two long-term, major streams of my research, DNA replication and purine biosynthesis, have merged with unexpected interconnections. If I have imparted to many of the talented individuals who have passed through my lab a degree of my passion for uncloaking the mysteries hidden in scientific research and an understanding of the honesty and rigor it demands and its impact on the world community, then my mentorship has been successful.
Assuntos
Bioquímica/história , Replicação do DNA , Enzimas , Purinas/biossíntese , Anti-Infecciosos/química , Anti-Infecciosos/farmacologia , Anticorpos Catalíticos/química , Anticorpos Catalíticos/metabolismo , Enzimas/química , Enzimas/metabolismo , História do Século XX , História do Século XXI , Humanos , Masculino , Estados UnidosRESUMO
SARS-CoV-2-induced hypercytokinemia and inflammation are critically associated with COVID-19 severity. Baricitinib, a clinically approved JAK1/JAK2 inhibitor, is currently being investigated in COVID-19 clinical trials. Here, we investigated the immunologic and virologic efficacy of baricitinib in a rhesus macaque model of SARS-CoV-2 infection. Viral shedding measured from nasal and throat swabs, bronchoalveolar lavages, and tissues was not reduced with baricitinib. Type I interferon (IFN) antiviral responses and SARS-CoV-2-specific T cell responses remained similar between the two groups. Animals treated with baricitinib showed reduced inflammation, decreased lung infiltration of inflammatory cells, reduced NETosis activity, and more limited lung pathology. Importantly, baricitinib-treated animals had a rapid and remarkably potent suppression of lung macrophage production of cytokines and chemokines responsible for inflammation and neutrophil recruitment. These data support a beneficial role for, and elucidate the immunological mechanisms underlying, the use of baricitinib as a frontline treatment for inflammation induced by SARS-CoV-2 infection.
Assuntos
Anti-Inflamatórios/administração & dosagem , Azetidinas/administração & dosagem , Tratamento Farmacológico da COVID-19 , COVID-19/imunologia , Macaca mulatta , Infiltração de Neutrófilos/efeitos dos fármacos , Purinas/administração & dosagem , Pirazóis/administração & dosagem , Sulfonamidas/administração & dosagem , Animais , COVID-19/fisiopatologia , Morte Celular/efeitos dos fármacos , Degranulação Celular/efeitos dos fármacos , Modelos Animais de Doenças , Inflamação/tratamento farmacológico , Inflamação/genética , Inflamação/imunologia , Janus Quinases/antagonistas & inibidores , Pulmão/efeitos dos fármacos , Pulmão/imunologia , Pulmão/patologia , Ativação Linfocitária/efeitos dos fármacos , Macrófagos Alveolares/imunologia , SARS-CoV-2/fisiologia , Índice de Gravidade de Doença , Linfócitos T/imunologia , Replicação Viral/efeitos dos fármacosRESUMO
Mutations in FAMIN cause arthritis and inflammatory bowel disease in early childhood, and a common genetic variant increases the risk for Crohn's disease and leprosy. We developed an unbiased liquid chromatography-mass spectrometry screen for enzymatic activity of this orphan protein. We report that FAMIN phosphorolytically cleaves adenosine into adenine and ribose-1-phosphate. Such activity was considered absent from eukaryotic metabolism. FAMIN and its prokaryotic orthologs additionally have adenosine deaminase, purine nucleoside phosphorylase, and S-methyl-5'-thioadenosine phosphorylase activity, hence, combine activities of the namesake enzymes of central purine metabolism. FAMIN enables in macrophages a purine nucleotide cycle (PNC) between adenosine and inosine monophosphate and adenylosuccinate, which consumes aspartate and releases fumarate in a manner involving fatty acid oxidation and ATP-citrate lyase activity. This macrophage PNC synchronizes mitochondrial activity with glycolysis by balancing electron transfer to mitochondria, thereby supporting glycolytic activity and promoting oxidative phosphorylation and mitochondrial H+ and phosphate recycling.
Assuntos
Peptídeos e Proteínas de Sinalização Intracelular/genética , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Adenina/metabolismo , Adenosina/metabolismo , Adenosina Desaminase/metabolismo , Cromatografia Líquida/métodos , Células HEK293 , Células Hep G2 , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/fisiologia , Espectrometria de Massas/métodos , Enzimas Multifuncionais/genética , Fosforilação , Proteínas/genética , Nucleotídeos de Purina/metabolismo , Purinas/metabolismoRESUMO
The gut microbiome has been implicated in multiple human chronic gastrointestinal (GI) disorders. Determining its mechanistic role in disease has been difficult due to apparent disconnects between animal and human studies and lack of an integrated multi-omics view of disease-specific physiological changes. We integrated longitudinal multi-omics data from the gut microbiome, metabolome, host epigenome, and transcriptome in the context of irritable bowel syndrome (IBS) host physiology. We identified IBS subtype-specific and symptom-related variation in microbial composition and function. A subset of identified changes in microbial metabolites correspond to host physiological mechanisms that are relevant to IBS. By integrating multiple data layers, we identified purine metabolism as a novel host-microbial metabolic pathway in IBS with translational potential. Our study highlights the importance of longitudinal sampling and integrating complementary multi-omics data to identify functional mechanisms that can serve as therapeutic targets in a comprehensive treatment strategy for chronic GI diseases. VIDEO ABSTRACT.
Assuntos
Microbioma Gastrointestinal/genética , Regulação da Expressão Gênica/genética , Síndrome do Intestino Irritável/metabolismo , Metaboloma , Purinas/metabolismo , Transcriptoma/genética , Animais , Ácidos e Sais Biliares/metabolismo , Biópsia , Butiratos/metabolismo , Cromatografia Líquida , Estudos Transversais , Epigenômica , Fezes/microbiologia , Feminino , Microbioma Gastrointestinal/fisiologia , Regulação da Expressão Gênica/fisiologia , Interações entre Hospedeiro e Microrganismos/genética , Humanos , Hipoxantina/metabolismo , Síndrome do Intestino Irritável/genética , Síndrome do Intestino Irritável/microbiologia , Estudos Longitudinais , Masculino , Metaboloma/fisiologia , Camundongos , Estudos Observacionais como Assunto , Estudos Prospectivos , Software , Espectrometria de Massas em Tandem , Transcriptoma/fisiologiaRESUMO
TLR8 is among the highest-expressed pattern-recognition receptors in the human myeloid compartment, yet its mode of action is poorly understood. TLR8 engages two distinct ligand binding sites to sense RNA degradation products, although it remains unclear how these ligands are formed in cellulo in the context of complex RNA molecule sensing. Here, we identified the lysosomal endoribonuclease RNase T2 as a non-redundant upstream component of TLR8-dependent RNA recognition. RNase T2 activity is required for rendering complex single-stranded, exogenous RNA molecules detectable for TLR8. This is due to RNase T2's preferential cleavage of single-stranded RNA molecules between purine and uridine residues, which critically contributes to the supply of catabolic uridine and the generation of purine-2',3'-cyclophosphate-terminated oligoribonucleotides. Thus-generated molecules constitute agonistic ligands for the first and second binding pocket of TLR8. Together, these results establish the identity and origin of the RNA-derived molecular pattern sensed by TLR8.
Assuntos
Endorribonucleases/metabolismo , Proteólise , Receptor 8 Toll-Like/metabolismo , Motivos de Aminoácidos , Sequência de Bases , Linhagem Celular , Endorribonucleases/deficiência , Humanos , Modelos Moleculares , Monócitos/metabolismo , Células Mieloides/metabolismo , Isótopos de Nitrogênio , Oligonucleotídeos/metabolismo , Purinas/metabolismo , RNA/metabolismo , Staphylococcus aureus/metabolismo , Receptor 7 Toll-Like/metabolismo , Receptor 8 Toll-Like/agonistas , Receptor 8 Toll-Like/química , Uridina/metabolismoRESUMO
Current machine learning techniques enable robust association of biological signals with measured phenotypes, but these approaches are incapable of identifying causal relationships. Here, we develop an integrated "white-box" biochemical screening, network modeling, and machine learning approach for revealing causal mechanisms and apply this approach to understanding antibiotic efficacy. We counter-screen diverse metabolites against bactericidal antibiotics in Escherichia coli and simulate their corresponding metabolic states using a genome-scale metabolic network model. Regression of the measured screening data on model simulations reveals that purine biosynthesis participates in antibiotic lethality, which we validate experimentally. We show that antibiotic-induced adenine limitation increases ATP demand, which elevates central carbon metabolism activity and oxygen consumption, enhancing the killing effects of antibiotics. This work demonstrates how prospective network modeling can couple with machine learning to identify complex causal mechanisms underlying drug efficacy.
Assuntos
Antibacterianos/metabolismo , Antibacterianos/farmacologia , Redes e Vias Metabólicas/efeitos dos fármacos , Adenina/metabolismo , Biologia Computacional/métodos , Avaliação Pré-Clínica de Medicamentos/métodos , Escherichia coli/metabolismo , Aprendizado de Máquina , Redes e Vias Metabólicas/imunologia , Modelos Teóricos , Purinas/metabolismoRESUMO
In two recent studies appearing in Cell1 and Cell Metabolism,2 Tran et al. and Wu et al. describe underappreciated nuance in organismal and cellular purine nucleotide salvage pathways and identify purine salvage as a metabolic limitation for tumor growth.
Assuntos
Purinas , Purinas/metabolismo , Humanos , Animais , Neoplasias/metabolismo , Neoplasias/genética , Neoplasias/patologia , Nucleotídeos de Purina/metabolismoRESUMO
The cyclic guanosine monophosphate (GMP)-AMP synthase (cGAS)-stimulator of interferon genes (STING) pathway plays a pivotal role in innate immune responses to viral infection and inhibition of autoimmunity. Recent studies have suggested that micronuclei formed by genotoxic stress can activate innate immune signaling via the cGAS-STING pathway. Here, we investigated cGAS localization, activation, and downstream signaling from micronuclei induced by ionizing radiation, replication stress, and chromosome segregation errors. Although cGAS localized to ruptured micronuclei via binding to self-DNA, we failed to observe cGAS activation; cGAMP production; downstream phosphorylation of STING, TBK1, or IRF3; nuclear accumulation of IRF3; or expression of interferon-stimulated genes. Failure to activate the cGAS-STING pathway was observed across primary and immortalized cell lines, which retained the ability to activate the cGAS-STING pathway in response to dsDNA or modified vaccinia virus infection. We provide evidence that micronuclei formed by genotoxic insults contain histone-bound self-DNA, which we show is inhibitory to cGAS activation in cells.
Assuntos
Micronúcleos com Defeito Cromossômico , Nucleotidiltransferases , Transdução de Sinais , Humanos , Células HeLa , Radiação , Replicação do DNA , Dano ao DNA , Fator Regulador 3 de Interferon/metabolismo , Transcrição Gênica , Fatores Reguladores de Interferon/metabolismo , Técnicas de Inativação de Genes , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Nucleotidiltransferases/genética , Nucleotidiltransferases/metabolismo , Cinética , Transfecção , Nucleossomos/metabolismo , Morfolinas , Purinas , Hidroxiureia , Linhagem Celular Tumoral , Vaccinia virus/fisiologia , Vacínia/imunologia , Vacínia/metabolismoRESUMO
Antigenic stimulation promotes T cell metabolic reprogramming to meet increased biosynthetic, bioenergetic, and signaling demands. We show that the one-carbon (1C) metabolism enzyme methylenetetrahydrofolate dehydrogenase 2 (MTHFD2) regulates de novo purine synthesis and signaling in activated T cells to promote proliferation and inflammatory cytokine production. In pathogenic T helper-17 (Th17) cells, MTHFD2 prevented aberrant upregulation of the transcription factor FoxP3 along with inappropriate gain of suppressive capacity. MTHFD2 deficiency also promoted regulatory T (Treg) cell differentiation. Mechanistically, MTHFD2 inhibition led to depletion of purine pools, accumulation of purine biosynthetic intermediates, and decreased nutrient sensor mTORC1 signaling. MTHFD2 was also critical to regulate DNA and histone methylation in Th17 cells. Importantly, MTHFD2 deficiency reduced disease severity in multiple in vivo inflammatory disease models. MTHFD2 is thus a metabolic checkpoint to integrate purine metabolism with pathogenic effector cell signaling and is a potential therapeutic target within 1C metabolism pathways.
Assuntos
Inflamação/imunologia , Alvo Mecanístico do Complexo 1 de Rapamicina/metabolismo , Metilenotetra-Hidrofolato Desidrogenase (NADP)/metabolismo , Purinas/biossíntese , Linfócitos T Reguladores/imunologia , Células Th17/imunologia , Animais , Diferenciação Celular , Citocinas/metabolismo , Metilação de DNA , Modelos Animais de Doenças , Humanos , Mediadores da Inflamação/metabolismo , Ativação Linfocitária , Metilenotetra-Hidrofolato Desidrogenase (NADP)/genética , Camundongos , Camundongos Transgênicos , Mutação/genética , Transdução de SinaisRESUMO
With the elucidation of myriad anabolic and catabolic enzyme-catalyzed cellular pathways crisscrossing each other, an obvious question arose: how could these networks operate with maximal catalytic efficiency and minimal interference? A logical answer was the postulate of metabolic channeling, which in its simplest embodiment assumes that the product generated by one enzyme passes directly to a second without diffusion into the surrounding medium. This tight coupling of activities might increase a pathway's metabolic flux and/or serve to sequester unstable/toxic/reactive intermediates as well as prevent their access to other networks. Here, we present evidence for this concept, commencing with enzymes that feature a physical molecular tunnel, to multi-enzyme complexes that retain pathway substrates through electrostatics or enclosures, and finally to metabolons that feature collections of enzymes assembled into clusters with variable stoichiometric composition. Lastly, we discuss the advantages of reversibly assembled metabolons in the context of the purinosome, the purine biosynthesis metabolon.
Assuntos
Redes e Vias Metabólicas/fisiologia , Metabolismo/fisiologia , Metaboloma/fisiologia , Animais , Humanos , Complexos Multienzimáticos/metabolismo , Mapas de Interação de Proteínas/fisiologia , Purinas/metabolismoRESUMO
Purine biosynthesis and metabolism, conserved in all living organisms, is essential for cellular energy homeostasis and nucleic acid synthesis. The de novo synthesis of purine precursors is under tight negative feedback regulation mediated by adenosine and guanine nucleotides. We describe a distinct early-onset neurodegenerative condition resulting from mutations in the adenosine monophosphate deaminase 2 gene (AMPD2). Patients have characteristic brain imaging features of pontocerebellar hypoplasia (PCH) due to loss of brainstem and cerebellar parenchyma. We found that AMPD2 plays an evolutionary conserved role in the maintenance of cellular guanine nucleotide pools by regulating the feedback inhibition of adenosine derivatives on de novo purine synthesis. AMPD2 deficiency results in defective GTP-dependent initiation of protein translation, which can be rescued by administration of purine precursors. These data suggest AMPD2-related PCH as a potentially treatable early-onset neurodegenerative disease.
Assuntos
AMP Desaminase/metabolismo , Atrofias Olivopontocerebelares/metabolismo , Purinas/biossíntese , AMP Desaminase/química , AMP Desaminase/genética , Animais , Tronco Encefálico/patologia , Cerebelo/patologia , Criança , Feminino , Guanosina Trifosfato/metabolismo , Humanos , Masculino , Camundongos , Camundongos Knockout , Mutação , Células-Tronco Neurais/metabolismo , Atrofias Olivopontocerebelares/genética , Atrofias Olivopontocerebelares/patologia , Biossíntese de Proteínas , Saccharomyces cerevisiae/enzimologia , Saccharomyces cerevisiae/metabolismoRESUMO
The RAS-ERK/MAPK (RAS-extracellular signal-regulated kinase/mitogen-activated protein kinase) pathway integrates growth-promoting signals to stimulate cell growth and proliferation, at least in part, through alterations in metabolic gene expression. However, examples of direct and rapid regulation of the metabolic pathways by the RAS-ERK pathway remain elusive. We find that physiological and oncogenic ERK signaling activation leads to acute metabolic flux stimulation through the de novo purine synthesis pathway, thereby increasing building block availability for RNA and DNA synthesis, which is required for cell growth and proliferation. We demonstrate that ERK2, but not ERK1, phosphorylates the purine synthesis enzyme PFAS (phosphoribosylformylglycinamidine synthase) at T619 in cells to stimulate de novo purine synthesis. The expression of nonphosphorylatable PFAS (T619A) decreases purine synthesis, RAS-dependent cancer cell-colony formation, and tumor growth. Thus, ERK2-mediated PFAS phosphorylation facilitates the increase in nucleic acid synthesis required for anabolic cell growth and proliferation.
Assuntos
Carbono-Nitrogênio Ligases com Glutamina como Doadora de N-Amida/metabolismo , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Purinas/biossíntese , Células A549 , Animais , Carbono-Nitrogênio Ligases com Glutamina como Doadora de N-Amida/genética , Ciclo Celular/fisiologia , Linhagem Celular Tumoral , Proliferação de Células/genética , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Células HeLa , Humanos , Sistema de Sinalização das MAP Quinases/fisiologia , Fosforilação , Purinas/metabolismo , Transdução de Sinais/fisiologia , Proteínas ras/metabolismoRESUMO
(p)ppGpp is a nucleotide messenger universally produced in bacteria following nutrient starvation. In E. coli, ppGpp inhibits purine nucleotide synthesis by targeting several different enzymes, but the physiological significance of their inhibition is unknown. Here, we report the structural basis of inhibition for one target, Gsk, the inosine-guanosine kinase. Gsk creates an unprecedented, allosteric binding pocket for ppGpp by restructuring terminal sequences, which restrains conformational dynamics necessary for catalysis. Guided by this structure, we generated a chromosomal mutation that abolishes Gsk regulation by ppGpp. This mutant strain accumulates abnormally high levels of purine nucleotides following amino-acid starvation, compromising cellular fitness. We demonstrate that this unrestricted increase in purine nucleotides is detrimental because it severely depletes pRpp and essential, pRpp-derived metabolites, including UTP, histidine, and tryptophan. Thus, our results reveal the significance of ppGpp's regulation of purine nucleotide synthesis and a critical mechanism by which E. coli coordinates biosynthetic processes during starvation.
Assuntos
Aminoácidos/biossíntese , Escherichia coli/metabolismo , Guanosina Tetrafosfato/metabolismo , Nucleotídeos/biossíntese , Proteínas de Escherichia coli/química , Proteínas de Escherichia coli/metabolismo , Retroalimentação Fisiológica , Guanosina Difosfato/metabolismo , Modelos Moleculares , Conformação Proteica , Multimerização Proteica , Purinas/biossíntese , Pirimidinas/biossínteseRESUMO
BACKGROUND: Ribociclib has been shown to have a significant overall survival benefit in patients with hormone receptor (HR)-positive, human epidermal growth factor receptor 2 (HER2)-negative advanced breast cancer. Whether this benefit in advanced breast cancer extends to early breast cancer is unclear. METHODS: In this international, open-label, randomized, phase 3 trial, we randomly assigned patients with HR-positive, HER2-negative early breast cancer in a 1:1 ratio to receive ribociclib (at a dose of 400 mg per day for 3 weeks, followed by 1 week off, for 3 years) plus a nonsteroidal aromatase inhibitor (NSAI; letrozole at a dose of 2.5 mg per day or anastrozole at a dose of 1 mg per day for ≥5 years) or an NSAI alone. Premenopausal women and men also received goserelin every 28 days. Eligible patients had anatomical stage II or III breast cancer. Here we report the results of a prespecified interim analysis of invasive disease-free survival, the primary end point; other efficacy and safety results are also reported. Invasive disease-free survival was evaluated with the use of the Kaplan-Meier method. The statistical comparison was made with the use of a stratified log-rank test, with a protocol-specified stopping boundary of a one-sided P-value threshold of 0.0128 for superior efficacy. RESULTS: As of the data-cutoff date for this prespecified interim analysis (January 11, 2023), a total of 426 patients had had invasive disease, recurrence, or death. A significant invasive disease-free survival benefit was seen with ribociclib plus an NSAI as compared with an NSAI alone. At 3 years, invasive disease-free survival was 90.4% with ribociclib plus an NSAI and 87.1% with an NSAI alone (hazard ratio for invasive disease, recurrence, or death, 0.75; 95% confidence interval, 0.62 to 0.91; P = 0.003). Secondary end points - distant disease-free survival and recurrence-free survival - also favored ribociclib plus an NSAI. The 3-year regimen of ribociclib at a 400-mg starting dose plus an NSAI was not associated with any new safety signals. CONCLUSIONS: Ribociclib plus an NSAI significantly improved invasive disease-free survival among patients with HR-positive, HER2-negative stage II or III early breast cancer. (Funded by Novartis; NATALEE ClinicalTrials.gov number, NCT03701334.).
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica , Inibidores da Aromatase , Neoplasias da Mama , Letrozol , Feminino , Humanos , Aminopiridinas/administração & dosagem , Aminopiridinas/efeitos adversos , Aminopiridinas/uso terapêutico , Protocolos de Quimioterapia Combinada Antineoplásica/administração & dosagem , Protocolos de Quimioterapia Combinada Antineoplásica/efeitos adversos , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/metabolismo , Neoplasias da Mama/mortalidade , Neoplasias da Mama/patologia , Letrozol/administração & dosagem , Letrozol/efeitos adversos , Letrozol/uso terapêutico , Purinas/administração & dosagem , Purinas/efeitos adversos , Purinas/uso terapêutico , Receptor ErbB-2/metabolismo , Inibidores da Aromatase/administração & dosagem , Inibidores da Aromatase/efeitos adversos , Inibidores da Aromatase/uso terapêutico , Receptores de Estrogênio , Receptores de Progesterona , Gosserrelina/administração & dosagem , Gosserrelina/efeitos adversos , Gosserrelina/uso terapêutico , Antineoplásicos Hormonais , MasculinoRESUMO
Mitochondrial uncoupling protein 1 (UCP1) is responsible for nonshivering thermogenesis in brown adipose tissue (BAT). Upon activation by long-chain fatty acids (LCFAs), UCP1 increases the conductance of the inner mitochondrial membrane (IMM) to make BAT mitochondria generate heat rather than ATP. Despite being a member of the family of mitochondrial anion carriers (SLC25), UCP1 is believed to transport H(+) by an unusual mechanism that has long remained unresolved. Here, we achieved direct patch-clamp measurements of UCP1 currents from the IMM of BAT mitochondria. We show that UCP1 is an LCFA anion/H(+) symporter. However, the LCFA anions cannot dissociate from UCP1 due to hydrophobic interactions established by their hydrophobic tails, and UCP1 effectively operates as an H(+) carrier activated by LCFA. A similar LCFA-dependent mechanism of transmembrane H(+) transport may be employed by other SLC25 members and be responsible for mitochondrial uncoupling and regulation of metabolic efficiency in various tissues.