RESUMO
O-GlcNAc is a dynamic post-translational modification (PTM) that regulates protein functions. In studying the regulatory roles of O-GlcNAc, a major roadblock is the inability to change O-GlcNAcylation on a single protein at a time. Herein, we developed a dual RNA-aptamer-based approach that simultaneously targeted O-GlcNAc transferase (OGT) and ß-catenin, the key transcription factor of the Wnt signaling pathway, to selectively increase O-GlcNAcylation of the latter without affecting other OGT substrates. Using the OGT/ß-catenin dual-specificity aptamers, we found that O-GlcNAcylation of ß-catenin stabilizes the protein by inhibiting its interaction with ß-TrCP. O-GlcNAc also increases ß-catenin's interaction with EZH2, recruits EZH2 to promoters, and dramatically alters the transcriptome. Further, by coupling riboswitches or an inducible expression system to aptamers, we enabled inducible regulation of protein-specific O-GlcNAcylation. Together, our findings demonstrate the efficacy and versatility of dual-specificity aptamers for regulating O-GlcNAcylation on individual proteins.
Assuntos
Aptâmeros de Nucleotídeos , beta Catenina/metabolismo , Processamento de Proteína Pós-Traducional , Via de Sinalização Wnt , N-Acetilglucosaminiltransferases/genética , N-Acetilglucosaminiltransferases/metabolismo , Acetilglucosamina/metabolismoRESUMO
Located at the inner leaflet of the plasma membrane (PM), phosphatidyl-inositol 4,5-bisphosphate [PI(4,5)P2] composes only 1-2 mol% of total PM lipids. With its synthesis and turnover both spatially and temporally regulated, PI(4,5)P2 recruits and interacts with hundreds of cellular proteins to support a broad spectrum of cellular functions. Several factors contribute to the versatile and dynamic distribution of PI(4,5)P2 in membranes. Physiological multivalent cations such as Ca2+ and Mg2+ can bridge between PI(4,5)P2 headgroups, forming nanoscopic PI(4,5)P2-cation clusters. The distinct lipid environment surrounding PI(4,5)P2 affects the degree of PI(4,5)P2 clustering. In addition, diverse cellular proteins interacting with PI(4,5)P2 can further regulate PI(4,5)P2 lateral distribution and accessibility. This review summarizes the current understanding of PI(4,5)P2 behavior in both cells and model membranes, with emphasis on both multivalent cation- and protein-induced PI(4,5)P2 clustering. Understanding the nature of spatially separated pools of PI(4,5)P2 is fundamental to cell biology.
Assuntos
Interações Hospedeiro-Patógeno/fisiologia , Fosfatidilinositol 4,5-Difosfato/metabolismo , Replicação Viral/fisiologia , Animais , Membrana Celular/metabolismo , Humanos , Micelas , Fosfatidilinositol 4,5-Difosfato/química , Proteínas Virais/metabolismoRESUMO
Cells bend their plasma membranes into highly curved forms to interact with the local environment, but how shape generation is regulated is not fully resolved. Here, we report a synergy between shape-generating processes in the cell interior and the external organization and composition of the cell-surface glycocalyx. Mucin biopolymers and long-chain polysaccharides within the glycocalyx can generate entropic forces that favor or disfavor the projection of spherical and finger-like extensions from the cell surface. A polymer brush model of the glycocalyx successfully predicts the effects of polymer size and cell-surface density on membrane morphologies. Specific glycocalyx compositions can also induce plasma membrane instabilities to generate more exotic undulating and pearled membrane structures and drive secretion of extracellular vesicles. Together, our results suggest a fundamental role for the glycocalyx in regulating curved membrane features that serve in communication between cells and with the extracellular matrix.
Assuntos
Forma Celular , Matriz Extracelular/metabolismo , Glicocálix/metabolismo , Glicoproteínas de Membrana/metabolismo , Mucinas/metabolismo , Animais , Linhagem Celular , Matriz Extracelular/genética , Glicocálix/genética , Cavalos , Humanos , Glicoproteínas de Membrana/genética , Mucinas/genéticaRESUMO
Hundreds of different mutations in genes encoding a few dozen sarcomeric proteins cause two reciprocal human disease phenotypes, hypertrophic or dilated cardiomyopathy. How molecular dysfunction evokes different patterns of cardiac remodeling is unclear. Davis et al. describe a biophysical metric of cardiomyocyte function, the force-time integral, which predicts disease phenotype.
Assuntos
Cardiomiopatia Dilatada/genética , Mutação , Coração , Humanos , FenótipoRESUMO
Whether the human fetus and the prenatal intrauterine environment (amniotic fluid and placenta) are stably colonized by microbial communities in a healthy pregnancy remains a subject of debate. Here we evaluate recent studies that characterized microbial populations in human fetuses from the perspectives of reproductive biology, microbial ecology, bioinformatics, immunology, clinical microbiology and gnotobiology, and assess possible mechanisms by which the fetus might interact with microorganisms. Our analysis indicates that the detected microbial signals are likely the result of contamination during the clinical procedures to obtain fetal samples or during DNA extraction and DNA sequencing. Furthermore, the existence of live and replicating microbial populations in healthy fetal tissues is not compatible with fundamental concepts of immunology, clinical microbiology and the derivation of germ-free mammals. These conclusions are important to our understanding of human immune development and illustrate common pitfalls in the microbial analyses of many other low-biomass environments. The pursuit of a fetal microbiome serves as a cautionary example of the challenges of sequence-based microbiome studies when biomass is low or absent, and emphasizes the need for a trans-disciplinary approach that goes beyond contamination controls by also incorporating biological, ecological and mechanistic concepts.
Assuntos
Biomassa , Contaminação por DNA , Feto , Microbiota , Animais , Feminino , Humanos , Gravidez , Líquido Amniótico/imunologia , Líquido Amniótico/microbiologia , Mamíferos , Microbiota/genética , Placenta/imunologia , Placenta/microbiologia , Feto/imunologia , Feto/microbiologia , Reprodutibilidade dos TestesRESUMO
In eukaryotes, gene expression is performed by three RNA polymerases that are targeted to promoters by molecular complexes. A unique common factor, the TATA-box binding protein (TBP), is thought to serve as a platform to assemble pre-initiation complexes competent for transcription. Here, we describe a novel molecular mechanism of nutrient regulation of gene transcription by dynamic O-GlcNAcylation of TBP. We show that O-GlcNAcylation at T114 of TBP blocks its interaction with BTAF1, hence the formation of the B-TFIID complex, and its dynamic cycling on and off of DNA. Transcriptomic and metabolomic analyses of TBPT114A CRISPR/Cas9-edited cells showed that loss of O-GlcNAcylation at T114 increases TBP binding to BTAF1 and directly impacts expression of 408 genes. Lack of O-GlcNAcylation at T114 is associated with a striking reprogramming of cellular metabolism induced by a profound modification of the transcriptome, leading to gross alterations in lipid storage.
Assuntos
Glucose/metabolismo , Gotículas Lipídicas/metabolismo , Metabolismo dos Lipídeos , Fatores Associados à Proteína de Ligação a TATA/metabolismo , Proteína de Ligação a TATA-Box/metabolismo , Fator de Transcrição TFIID/metabolismo , Animais , Cromatina/genética , Cromatina/metabolismo , Diabetes Mellitus Experimental/genética , Diabetes Mellitus Experimental/metabolismo , Regulação da Expressão Gênica , Glicosilação , Células HEK293 , Células HeLa , Humanos , Metabolismo dos Lipídeos/genética , Masculino , Complexos Multiproteicos , Ratos Sprague-Dawley , Transdução de Sinais , Fatores Associados à Proteína de Ligação a TATA/genética , Proteína de Ligação a TATA-Box/genética , Fatores de Tempo , Fator de Transcrição TFIID/genética , Transcrição Gênica , TranscriptomaRESUMO
O-GlcNAcylation is the addition of ß-D-N-acetylglucosamine to serine or threonine residues of nuclear and cytoplasmic proteins. O-linked N-acetylglucosamine (O-GlcNAc) was not discovered until the early 1980s and still remains difficult to detect and quantify. Nonetheless, O-GlcNAc is highly abundant and cycles on proteins with a timescale similar to protein phosphorylation. O-GlcNAc occurs in organisms ranging from some bacteria to protozoans and metazoans, including plants and nematodes up the evolutionary tree to man. O-GlcNAcylation is mostly on nuclear proteins, but it occurs in all intracellular compartments, including mitochondria. Recent glycomic analyses have shown that O-GlcNAcylation has surprisingly extensive cross talk with phosphorylation, where it serves as a nutrient/stress sensor to modulate signaling, transcription, and cytoskeletal functions. Abnormal amounts of O-GlcNAcylation underlie the etiology of insulin resistance and glucose toxicity in diabetes, and this type of modification plays a direct role in neurodegenerative disease. Many oncogenic proteins and tumor suppressor proteins are also regulated by O-GlcNAcylation. Current data justify extensive efforts toward a better understanding of this invisible, yet abundant, modification. As tools for the study of O-GlcNAc become more facile and available, exponential growth in this area of research will eventually take place.
Assuntos
Acetilglucosamina/metabolismo , Doença Crônica , Transdução de Sinais/fisiologia , Transcrição Gênica , Acetilglucosamina/química , Animais , Diabetes Mellitus/fisiopatologia , Glicosilação , Humanos , Modelos Moleculares , Estrutura Molecular , N-Acetilglucosaminiltransferases/metabolismo , Neoplasias/fisiopatologia , Doenças Neurodegenerativas/fisiopatologia , Fosforilação , Conformação ProteicaRESUMO
The central functions fulfilled by mitochondria as both energy generators essential for tissue homeostasis and gateways to programmed apoptotic and necrotic cell death mandate tight control over the quality and quantity of these ubiquitous endosymbiotic organelles. Mitophagy, the targeted engulfment and destruction of mitochondria by the cellular autophagy apparatus, has conventionally been considered as the mechanism primarily responsible for mitochondrial quality control. However, our understanding of how, why, and under what specific conditions mitophagy is activated has grown tremendously over the past decade. Evidence is accumulating that nonmitophagic mitochondrial quality control mechanisms are more important to maintaining normal tissue homeostasis whereas mitophagy is an acute tissue stress response. Moreover, previously unrecognized mitophagic regulation of mitochondrial quantity control, metabolic reprogramming, and cell differentiation suggests that the mechanisms linking genetic or acquired defects in mitophagy to neurodegenerative and cardiovascular diseases or cancer are more complex than simple failure of normal mitochondrial quality control. Here, we provide a comprehensive overview of mitophagy in cellular homeostasis and disease and examine the most revolutionary concepts in these areas. In this context, we discuss evidence that atypical mitophagy and nonmitophagic pathways play central roles in mitochondrial quality control, functioning that was previously considered to be the primary domain of mitophagy.
Assuntos
Autofagia/fisiologia , Homeostase/fisiologia , Mitocôndrias/metabolismo , Mitofagia/fisiologia , Animais , Diferenciação Celular/fisiologia , Humanos , Neoplasias/metabolismoRESUMO
BACKGROUND: In a previous phase 3 trial, treatment with trifluridine-tipiracil (FTD-TPI) prolonged overall survival among patients with metastatic colorectal cancer. Preliminary data from single-group and randomized phase 2 trials suggest that treatment with FTD-TPI in addition to bevacizumab has the potential to extend survival. METHODS: We randomly assigned, in a 1:1 ratio, adult patients who had received no more than two previous chemotherapy regimens for the treatment of advanced colorectal cancer to receive FTD-TPI plus bevacizumab (combination group) or FTD-TPI alone (FTD-TPI group). The primary end point was overall survival. Secondary end points were progression-free survival and safety, including the time to worsening of the Eastern Cooperative Oncology Group (ECOG) performance-status score from 0 or 1 to 2 or more (on a scale from 0 to 5, with higher scores indicating greater disability). RESULTS: A total of 246 patients were assigned to each group. The median overall survival was 10.8 months in the combination group and 7.5 months in the FTD-TPI group (hazard ratio for death, 0.61; 95% confidence interval [CI], 0.49 to 0.77; P<0.001). The median progression-free survival was 5.6 months in the combination group and 2.4 months in the FTD-TPI group (hazard ratio for disease progression or death, 0.44; 95% CI, 0.36 to 0.54; P<0.001). The most common adverse events in both groups were neutropenia, nausea, and anemia. No treatment-related deaths were reported. The median time to worsening of the ECOG performance-status score from 0 or 1 to 2 or more was 9.3 months in the combination group and 6.3 months in the FTD-TPI group (hazard ratio, 0.54; 95% CI, 0.43 to 0.67). CONCLUSIONS: Among patients with refractory metastatic colorectal cancer, treatment with FTD-TPI plus bevacizumab resulted in longer overall survival than FTD-TPI alone. (Funded by Servier and Taiho Oncology; SUNLIGHT ClinicalTrials.gov number, NCT04737187; EudraCT number, 2020-001976-14.).
Assuntos
Neoplasias do Colo , Neoplasias Colorretais , Neoplasias Retais , Adulto , Humanos , Protocolos de Quimioterapia Combinada Antineoplásica/efeitos adversos , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Bevacizumab/efeitos adversos , Bevacizumab/uso terapêutico , Neoplasias do Colo/tratamento farmacológico , Neoplasias Colorretais/tratamento farmacológico , Combinação de Medicamentos , Pirrolidinas/efeitos adversos , Pirrolidinas/uso terapêutico , Neoplasias Retais/tratamento farmacológico , Trifluridina/efeitos adversos , Trifluridina/uso terapêutico , UracilaRESUMO
Mitochondrial involvement in neurodegenerative diseases is widespread and multifactorial. Targeting mitochondrial pathology is therefore of interest. The recent development of bioactive molecules that modulate mitochondrial dynamics (fusion, fission and motility) offers a new therapeutic approach for neurodegenerative diseases with either indirect or direct mitochondrial involvement. Here, we asked: (1) Can enhanced mitochondrial fusion and motility improve secondary mitochondrial pathology in superoxide dismutase1 (SOD1) mutant amyotrophic lateral sclerosis (ALS)? And: (2) What is the impact of enhancing mitochondria fitness on in vivo manifestations of SOD1 mutant ALS? We observed that small molecule mitofusin activators corrected mitochondrial fragmentation, depolarization and dysmotility in genetically diverse ALS patient reprogrammed motor neurons and fibroblasts, and in motor neurons, sensory neurons and fibroblasts from SOD1 G93A mice. Continuous, but not intermittent, pharmacologic mitofusin activation delayed phenotype progression and lethality in SOD1 G93A mice, reducing neuron loss and improving neuromuscular connectivity. Mechanistically, mitofusin activation increased mitochondrial motility, fitness and residency within neuromuscular synapses; reduced mitochondrial reactive oxygen species production; and diminished apoptosis in SOD1 mutant neurons. These benefits were accompanied by improved mitochondrial respiratory coupling, despite characteristic SOD1 mutant ALS-associated downregulation of mitochondrial respiratory complexes. Targeting mitochondrial dysdynamism is a promising approach to alleviate pathology caused by secondary mitochondrial dysfunction in some neurodegenerative diseases.
Assuntos
Esclerose Lateral Amiotrófica , Camundongos , Animais , Esclerose Lateral Amiotrófica/tratamento farmacológico , Esclerose Lateral Amiotrófica/genética , Esclerose Lateral Amiotrófica/patologia , Superóxido Dismutase-1/genética , Superóxido Dismutase-1/metabolismo , Superóxidos/metabolismo , Superóxido Dismutase/genética , Superóxido Dismutase/metabolismo , Camundongos Transgênicos , Neurônios Motores/metabolismo , Mitocôndrias/genética , Mitocôndrias/patologia , Progressão da Doença , Modelos Animais de DoençasRESUMO
BACKGROUND & AIMS: More than half of pancreatic ductal adenocarcinomas (PDACs) recur within 12 months after curative-intent resection. This systematic review and meta-analysis was conducted to identify all reported prognostic factors for early recurrence in resected PDACs. METHODS: After a systematic literature search, a meta-analysis was conducted using a random effects model. Separate analyses were performed for adjusted vs unadjusted effect estimates as well as reported odds ratios (ORs) and hazard ratios (HRs). Risk of bias was assessed using the Quality in Prognostic Studies tool, and evidence was rated according to Grading of Recommendations Assessment, Development and Evaluation recommendations. RESULTS: After 2,903 abstracts were screened, 65 studies were included. Of these, 28 studies (43.1%) defined early recurrence as evidence of recurrence within 6 months, whereas 34 (52.3%) defined it as evidence of recurrence within 12 months after surgery. Other definitions were uncommon. Analysis of unadjusted ORs and HRs revealed 41 and 5 prognostic factors for early recurrence within 6 months, respectively. When exclusively considering adjusted data, we identified 25 and 10 prognostic factors based on OR and HR, respectively. Using a 12-month definition, we identified 38 (OR) and 15 (HR) prognostic factors from unadjusted data and 38 (OR) and 30 (HR) prognostic factors from adjusted data, respectively. On the basis of frequency counts of adjusted data, preoperative carbohydrate antigen 9-9, N status, nondelivery of adjuvant therapy, grading, and tumor size based on imaging were identified as key prognostic factors for early recurrence. CONCLUSIONS: Reported prognostic factors of early recurrence vary considerably. Identified key prognostic factors could aid in the development of a risk stratification framework for early recurrence. However, prospective validation is necessary.
RESUMO
BACKGROUND: Cardiac contractile function requires high energy from mitochondria, and Ca2+ from the sarcoplasmic reticulum (SR). Via local Ca2+ transfer at close mitochondria-SR contacts, cardiac excitation feedforward regulates mitochondrial ATP production to match surges in demand (excitation-bioenergetics coupling). However, pathological stresses may cause mitochondrial Ca2+ overload, excessive reactive oxygen species production and permeability transition, risking homeostatic collapse and myocyte loss. Excitation-bioenergetics coupling involves mitochondria-SR tethers but the role of tethering in cardiac physiology/pathology is debated. Endogenous tether proteins are multifunctional; therefore, nonselective targets to scrutinize interorganelle linkage. Here, we assessed the physiological/pathological relevance of selective chronic enhancement of cardiac mitochondria-SR tethering. METHODS: We introduced to mice a cardiac muscle-specific engineered tether (linker) transgene with a fluorescent protein core and deployed 2D/3D electron microscopy, biochemical approaches, fluorescence imaging, in vivo and ex vivo cardiac performance monitoring and stress challenges to characterize the linker phenotype. RESULTS: Expressed in the mature cardiomyocytes, the linker expanded and tightened individual mitochondria-junctional SR contacts; but also evoked a marked remodeling with large dense mitochondrial clusters that excluded dyads. Yet, excitation-bioenergetics coupling remained well-preserved, likely due to more longitudinal mitochondria-dyad contacts and nanotunnelling between mitochondria exposed to junctional SR and those sealed away from junctional SR. Remarkably, the linker decreased female vulnerability to acute massive ß-adrenergic stress. It also reduced myocyte death and mitochondrial calcium-overload-associated myocardial impairment in ex vivo ischemia/reperfusion injury. CONCLUSIONS: We propose that mitochondria-SR/endoplasmic reticulum contacts operate at a structural optimum. Although acute changes in tethering may cause dysfunction, upon chronic enhancement of contacts from early life, adaptive remodeling of the organelles shifts the system to a new, stable structural optimum. This remodeling balances the individually enhanced mitochondrion-junctional SR crosstalk and excitation-bioenergetics coupling, by increasing the connected mitochondrial pool and, presumably, Ca2+/reactive oxygen species capacity, which then improves the resilience to stresses associated with dysregulated hyperactive Ca2+ signaling.
Assuntos
Sinalização do Cálcio , Retículo Sarcoplasmático , Feminino , Camundongos , Animais , Retículo Sarcoplasmático/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Miocárdio/metabolismo , Miócitos Cardíacos/metabolismo , Mitocôndrias Cardíacas/metabolismo , Cálcio/metabolismoRESUMO
Cells run on carbohydrates. Glycans, sequences of carbohydrates conjugated to proteins and lipids, are arguably the most abundant and structurally diverse class of molecules in nature. Recent advances in glycomics reveal the scope and scale of their functional roles and their impact on human disease.
Assuntos
Glicômica , Animais , Biomarcadores Tumorais/análise , Glicolipídeos/análise , Glicosilação , Glicosiltransferases/genética , Glicosiltransferases/metabolismo , Humanos , Neoplasias/diagnóstico , Polissacarídeos/análise , Polissacarídeos/metabolismo , Processamento de Proteína Pós-Traducional , Proteoglicanas/análiseRESUMO
Management of hospitalized patients with type 2 diabetes mellitus (T2DM) presents unique challenges. Two recently released guidelines, one from the American Diabetes Association and the other from the Endocrine Society, provide useful recommendations and evidence review to inform the care of medical inpatients with T2DM. These guidelines mostly agree, although there are slight differences in their recommendations. In these rounds, 2 expert diabetologists discuss their approach to inpatient management of T2DM, specifically regarding inpatient glycemic goals on the medical ward, the use of noninsulin antihyperglycemic medications, and patient safety strategies for patients receiving long-acting insulin. They conclude with recommendations for Mr. D, a real patient with T2DM admitted with a recurrent foot infection.
RESUMO
Nearly 15% of U.S. adults have diabetes; type 2 diabetes (T2D) accounts for more than 90% of cases. Approximately one third of all patients with diabetes will develop chronic kidney disease (CKD). All patients with T2D should be screened annually for CKD with both a urine albumin-creatinine ratio and an estimated glomerular filtration rate. Research into strategies to slow the worsening of CKD and reduce renal and cardiovascular morbidity in patients with T2D and CKD has evolved substantially. In 2022, a consensus statement from the American Diabetes Association and the Kidney Disease: Improving Global Outcomes recommended prioritizing the use of sodium-glucose cotransporter-2 inhibitors and metformin and included guidance for add-on therapy with glucagon-like peptide 1 receptors agonists for most patients whose first-line therapy failed. It also recommended nonsteroidal mineralocorticoid receptor antagonists for patients with hypertension that is not adequately controlled with angiotensin-converting enzyme inhibitors or angiotensin-receptor blockers. Here, an endocrinologist and a nephrologist discuss the care of patients with T2D and CKD and how they would apply the consensus statement to the care of an individual patient with T2D who is unaware that he has CKD.
Assuntos
Diabetes Mellitus Tipo 2 , Insuficiência Renal Crônica , Inibidores do Transportador 2 de Sódio-Glicose , Humanos , Diabetes Mellitus Tipo 2/complicações , Diabetes Mellitus Tipo 2/tratamento farmacológico , Insuficiência Renal Crônica/complicações , Insuficiência Renal Crônica/tratamento farmacológico , Inibidores do Transportador 2 de Sódio-Glicose/uso terapêutico , Hipoglicemiantes/uso terapêutico , Antagonistas de Receptores de Mineralocorticoides/uso terapêutico , Visitas de Preceptoria , Nefropatias Diabéticas/tratamento farmacológico , Metformina/uso terapêutico , Receptor do Peptídeo Semelhante ao Glucagon 1/agonistas , Taxa de Filtração Glomerular , MasculinoRESUMO
Elevation of intracellular Ca2+ concentration ([Ca2+]i) activates Ca2+/calmodulin-dependent kinases (CaMK) and promotes gene transcription. This signaling pathway is referred to as excitationtranscription (E-T) coupling. Although vascular myocytes can exhibit E-T coupling, the molecular mechanisms and physiological/pathological roles are unknown. Multiscale analysis spanning from single molecules to whole organisms has revealed essential steps in mouse vascular myocyte E-T coupling. Upon a depolarizing stimulus, Ca2+ influx through Cav1.2 voltage-dependent Ca2+ channels activates CaMKK2 and CaMK1a, resulting in intranuclear CREB phosphorylation. Within caveolae, the formation of a molecular complex of Cav1.2/CaMKK2/CaMK1a is promoted in vascular myocytes. Live imaging using a genetically encoded Ca2+ indicator revealed direct activation of CaMKK2 by Ca2+ influx through Cav1.2 localized to caveolae. CaMK1a is phosphorylated by CaMKK2 at caveolae and translocated to the nucleus upon membrane depolarization. In addition, sustained depolarization of a mesenteric artery preparation induced genes related to chemotaxis, leukocyte adhesion, and inflammation, and these changes were reversed by inhibitors of Cav1.2, CaMKK2, and CaMK, or disruption of caveolae. In the context of pathophysiology, when the mesenteric artery was loaded by high pressure in vivo, we observed CREB phosphorylation in myocytes, macrophage accumulation at adventitia, and an increase in thickness and cross-sectional area of the tunica media. These changes were reduced in caveolin1-knockout mice or in mice treated with the CaMKK2 inhibitor STO609. In summary, E-T coupling depends on Cav1.2/CaMKK2/CaMK1a localized to caveolae, and this complex converts [Ca2+]i changes into gene transcription. This ultimately leads to macrophage accumulation and media remodeling for adaptation to increased circumferential stretch.
Assuntos
Canais de Cálcio Tipo L , Quinase da Proteína Quinase Dependente de Cálcio-Calmodulina , Proteína Quinase Tipo 1 Dependente de Cálcio-Calmodulina , Cavéolas , Transcrição Gênica , Remodelação Vascular , Animais , Cálcio/metabolismo , Canais de Cálcio Tipo L/metabolismo , Quinase da Proteína Quinase Dependente de Cálcio-Calmodulina/metabolismo , Proteína Quinase Tipo 1 Dependente de Cálcio-Calmodulina/metabolismo , Cavéolas/metabolismo , Caveolina 1/genética , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Acoplamento Excitação-Contração , Camundongos , Camundongos Knockout , Músculo Liso Vascular/metabolismo , Músculo Liso Vascular/fisiologia , Miócitos de Músculo Liso/metabolismo , Miócitos de Músculo Liso/fisiologia , Neurônios/metabolismo , FosforilaçãoRESUMO
The ionotropic purinergic P2X7 receptor responds to extracellular ATP and can trigger proinflammatory immune signaling in macrophages. Caveolin-1 (Cav-1) is known to modulate functions of macrophages and innate immunity. However, it is unknown how Cav-1 modulates P2X7 receptor activity in macrophages. We herein examined P2X7 receptor activity and macrophage functions using bone marrow-derived macrophages (BMDMs) from wild-type (WT) and Cav-1 knockout (KO) mice. ATP (1 mM) application caused biphasic increase in cytosolic [Ca2+] and sustained decrease in cytosolic [K+]. A specific P2X7 receptor blocker, A-740003, inhibited the maintained cytosolic [Ca2+] increase and cytosolic [K+] decrease. Total internal reflection fluorescent imaging and proximity ligation assays revealed a novel molecular complex formation between P2X7 receptors and Cav-1 in WT BMDMs that were stimulated with lipopolysaccharides. This molecular coupling was increased by ATP application. Specifically, the ATP-induced Ca2+ influx and K+ efflux through P2X7 receptors were increased in Cav-1 KO BMDMs, even though the total and surface protein levels of P2X7 receptors in WT and Cav-1 KO BMDMs were unchanged. Cell-impermeable dye (TO-PRO3) uptake analysis revealed that macropore formation of P2X7 receptors was enhanced in Cav-1 KO BMDMs. Cav-1 KO BMDMs increased ATP-induced IL-1ß secretion, reactive oxygen species production, Gasdermin D (GSDMD) cleavage, and lactate dehydrogenase release indicating pyroptosis. A-740003 completely prevented ATP-induced pyroptosis. In combination, these datasets show that Cav-1 has a negative effect on P2X7 receptor activity in BMDMs and that Cav-1 in macrophages may contribute to finely tuned immune responses by preventing excessive IL-1ß secretion and pyroptosis.NEW & NOTEWORTHY In bone marrow-derived macrophages, Cav-1 suppresses the macropore formation of P2X7 receptors through their direct or indirect interactions, resulting in reduced membrane permeability of cations (Ca2+ and K+) and large cell-impermeable dye (TO-PRO3) induced by ATP. Cav-1 also inhibits ATP-induced IL-1ß secretion, ROS production, GSDMD cleavage, and pyroptosis. Cav-1 contributes to the maintenance of proper immune responses by finely tuning IL-1ß secretion and cell death in macrophages.
Assuntos
Caveolina 1 , Receptores Purinérgicos P2X7 , Animais , Camundongos , Trifosfato de Adenosina/farmacologia , Trifosfato de Adenosina/metabolismo , Caveolina 1/genética , Caveolina 1/metabolismo , Interleucina-1beta/metabolismo , Macrófagos/metabolismo , Receptores Purinérgicos P2X7/metabolismoRESUMO
T-type calcium channels perform crucial physiological roles across a wide spectrum of tissues, spanning both neuronal and non-neuronal system. For instance, they serve as pivotal regulators of neuronal excitability, contribute to cardiac pacemaking, and mediate the secretion of hormones. These functions significantly hinge upon the intricate interplay of T-type channels with interacting proteins that modulate their expression and function at the plasma membrane. In this review, we offer a panoramic exploration of the current knowledge surrounding these T-type channel interactors, and spotlight certain aspects of their potential for drug-based therapeutic intervention.
Assuntos
Canais de Cálcio Tipo T , Cálcio , Cálcio/metabolismo , Bloqueadores dos Canais de Cálcio , Canais de Cálcio Tipo T/metabolismo , Neurônios/metabolismoRESUMO
Whether or not populations diverge with respect to the genetic contribution to risk of specific complex diseases is relevant to understanding the evolution of susceptibility and origins of health disparities. Here, we describe a large-scale whole-genome sequencing study of inflammatory bowel disease encompassing 1,774 affected individuals and 1,644 healthy control Americans with African ancestry (African Americans). Although no new loci for inflammatory bowel disease are discovered at genome-wide significance levels, we identify numerous instances of differential effect sizes in combination with divergent allele frequencies. For example, the major effect at PTGER4 fine maps to a single credible interval of 22 SNPs corresponding to one of four independent associations at the locus in European ancestry individuals but with an elevated odds ratio for Crohn disease in African Americans. A rare variant aggregate analysis implicates Ca2+-binding neuro-immunomodulator CALB2 in ulcerative colitis. Highly significant overall overlap of common variant risk for inflammatory bowel disease susceptibility between individuals with African and European ancestries was observed, with 41 of 241 previously known lead variants replicated and overall correlations in effect sizes of 0.68 for combined inflammatory bowel disease. Nevertheless, subtle differences influence the performance of polygenic risk scores, and we show that ancestry-appropriate weights significantly improve polygenic prediction in the highest percentiles of risk. The median amount of variance explained per locus remains the same in African and European cohorts, providing evidence for compensation of effect sizes as allele frequencies diverge, as expected under a highly polygenic model of disease.
Assuntos
Calbindina 2/genética , Predisposição Genética para Doença , Doenças Inflamatórias Intestinais/genética , Receptores de Prostaglandina E Subtipo EP4/genética , Negro ou Afro-Americano/genética , Idoso , Idoso de 80 Anos ou mais , Colite Ulcerativa/genética , Colite Ulcerativa/patologia , Doença de Crohn/genética , Doença de Crohn/patologia , Feminino , Frequência do Gene , Estudo de Associação Genômica Ampla , Humanos , Doenças Inflamatórias Intestinais/patologia , Masculino , Herança Multifatorial/genética , Polimorfismo de Nucleotídeo Único/genética , População Branca/genética , Sequenciamento Completo do GenomaRESUMO
Gut microbiota function has numerous effects on humans and the diet humans consume has emerged as a pivotal determinant of gut microbiota function. Here, a new concept that gut microbiota can be trained by diet-derived exosome-like nanoparticles (ELNs) to release healthy outer membrane vesicles (OMVs) is introduced. Specifically, OMVs released from garlic ELN (GaELNs) trained human gut Akkermansia muciniphila (A. muciniphila) can reverse high-fat diet-induced type 2 diabetes (T2DM) in mice. Oral administration of OMVs released from GaELNs trained A. muciniphila can traffick to the brain where they are taken up by microglial cells, resulting in inhibition of high-fat diet-induced brain inflammation. GaELNs treatment increases the levels of OMV Amuc-1100, P9, and phosphatidylcholines. Increasing the levels of Amuc-1100 and P9 leads to increasing the GLP-1 plasma level. Increasing the levels of phosphatidylcholines is required for inhibition of cGas and STING-mediated inflammation and GLP-1R crosstalk with the insulin pathway that leads to increasing expression of Insulin Receptor Substrate (IRS1 and IRS2) on OMV targeted cells. These findings reveal a molecular mechanism whereby OMVs from plant nanoparticle-trained gut bacteria regulate genes expressed in the brain, and have implications for the treatment of brain dysfunction caused by a metabolic syndrome.