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1.
Int J Mol Sci ; 25(7)2024 Mar 26.
Artigo em Inglês | MEDLINE | ID: mdl-38612490

RESUMO

Endometriosis is one of the leading pathologies of the reproductive system of women of fertile age, which shows changes in cell metabolism in the lesions. We conducted a study of the cellular respiration according to the polarography and the mRNA content of the main metabolic proteins using qRT-PCR of intraoperative endometrial biopsies from patients in the control group and with different localizations of endometriosis (adenomyosis, endometrioma, pelvic peritoneum). In biopsy samples of patients with endometriomas and pelvic peritoneum endometriotic lesions, the rate of oxygen absorption was significantly reduced, and, moreover, in the extragenital case, there was a shift to succinate utilization. The mRNA content of the cytochrome c, cytochrome c oxidase, and ATP synthase was also reduced, but hexokinase HK2 as well as pyruvate kinase were significantly higher than in the control. These oxidative phosphorylation and gene expression profiles suggest the Warburg effect and a shift in metabolism toward glycolysis. For adenomyosis, on the contrary, cellular respiration was significantly higher than in the control group due to the terminal region of the respiratory chain, ATP synthase, and its mRNA was increased as well. These data allow us to suggest that the therapeutic strategies of endometriosis based on modulation energy metabolism should take lesion localization into account.


Assuntos
Adenomiose , Endometriose , Humanos , Feminino , Endometriose/genética , Respiração Celular , Biópsia , RNA Mensageiro/genética , Trifosfato de Adenosina
2.
Proc Natl Acad Sci U S A ; 121(15): e2309636121, 2024 Apr 09.
Artigo em Inglês | MEDLINE | ID: mdl-38573964

RESUMO

Rates of microbial processes are fundamental to understanding the significance of microbial impacts on environmental chemical cycling. However, it is often difficult to quantify rates or to link processes to specific taxa or individual cells, especially in environments where there are few cultured representatives with known physiology. Here, we describe the use of the redox-enzyme-sensitive molecular probe RedoxSensor™ Green to measure rates of anaerobic electron transfer physiology (i.e., sulfate reduction and methanogenesis) in individual cells and link those measurements to genomic sequencing of the same single cells. We used this method to investigate microbial activity in hot, anoxic, low-biomass (~103 cells mL-1) groundwater of the Death Valley Regional Flow System, California. Combining this method with electron donor amendment experiments and metatranscriptomics confirmed that the abundant spore formers including Candidatus Desulforudis audaxviator were actively reducing sulfate in this environment, most likely with acetate and hydrogen as electron donors. Using this approach, we measured environmental sulfate reduction rates at 0.14 to 26.9 fmol cell-1 h-1. Scaled to volume, this equates to a bulk environmental rate of ~103 pmol sulfate L-1 d-1, similar to potential rates determined with radiotracer methods. Despite methane in the system, there was no evidence for active microbial methanogenesis at the time of sampling. Overall, this method is a powerful tool for estimating species-resolved, single-cell rates of anaerobic metabolism in low-biomass environments while simultaneously linking genomes to phenomes at the single-cell level. We reveal active elemental cycling conducted by several species, with a large portion attributable to Ca. Desulforudis audaxviator.


Assuntos
Ecossistema , Meio Ambiente , Transporte de Elétrons , Sulfatos/química , Respiração Celular
3.
Front Cell Infect Microbiol ; 14: 1287557, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38577619

RESUMO

Despite extensive knowledge of antibiotic-targeted bacterial cell death, deeper understanding of antibiotic tolerance mechanisms is necessary to combat multi-drug resistance in the global healthcare settings. Regulatory RNAs in bacteria control important cellular processes such as cell division, cellular respiration, metabolism, and virulence. Here, we investigated how exposing Escherichia coli to the moderately effective first-generation antibiotic cephalothin alters transcriptional and post-transcriptional dynamics. Bacteria switched from active aerobic respiration to anaerobic adaptation via an FnrS and Tp2 small RNA-mediated post-transcriptional regulatory circuit. From the early hours of antibiotic exposure, FnrS was involved in regulating reactive oxygen species levels, and delayed oxygen consumption in bacteria. We demonstrated that bacteria strive to maintain cellular homeostasis via sRNA-mediated sudden respiratory changes upon sublethal antibiotic exposure.


Assuntos
Antibacterianos , RNA , Antibacterianos/farmacologia , Anaerobiose , Respiração Celular , Bactérias , Respiração , Regulação Bacteriana da Expressão Gênica
4.
Methods Mol Biol ; 2761: 49-55, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38427228

RESUMO

High-resolution mitochondrial respirometry is a modern technique that enables to measure mitochondrial respiration in various cell types. It contains chambers with oxygen sensors that measure oxygen concentration via polarography and calculate its consumption. The chamber contains plastic stoppers with injection ports that allow the injection of samples and different substrates, inhibitors, and uncoupler substances to measure mitochondrial respiration with high efficiency. These substances act on the mitochondrial electron transport chain (ETC) and help to assess the mitochondrial ATP production capacity and oxidative phosphorylation. The respirograph obtained with the help of software represents the oxygen consumption in each stage after adding different reagents.


Assuntos
Respiração Celular , Roedores , Animais , Mitocôndrias/metabolismo , Fosforilação Oxidativa , Consumo de Oxigênio , Encéfalo/metabolismo , Oxigênio/metabolismo
5.
Cell Commun Signal ; 22(1): 120, 2024 02 12.
Artigo em Inglês | MEDLINE | ID: mdl-38347585

RESUMO

BACKGROUND: Breast cancer (BC) is the most frequent tumor entity in women worldwide with a high chance of therapeutic response in early- and non-metastatic disease stages. Among all BC subtypes, triple-negative BC (TNBC) is the most challenging cancer subtype lacking effective molecular targets due to the particular enrichment of cancer stem cells (CSCs), frequently leading to a chemoresistant phenotype and metastasis. The Ubiquitin Specific Peptidase 22 (USP22) is a deubiquitinase that has been frequently associated with a CSC-promoting function and intimately implicated in resistance to conventional therapies, tumor relapse, metastasis and overall poor survival in a broad range of cancer entities, including BC. To date, though, the role of USP22 in TNBC has been only superficially addressed. METHODS: The current study utilized the MMTV-cre, Usp22fl/fl transgenic mouse model to study the involvement of USP22 in the stem cell-like properties of the growing mammary tissue. Additionally, we combined high-throughput transcriptomic analyses with publicly available patient transcriptomic data and utilized TNBC culture models to decipher the functional role of USP22 in the CSC characteristics of this disease. RESULTS: Interestingly, we identified that USP22 promotes CSC properties and drug tolerance by supporting the oxidative phosphorylation program, known to be largely responsible for the poor response to conventional therapies in this particularly aggressive BC subtype. CONCLUSIONS: This study suggests a novel tumor-supportive role of USP22 in sustaining cellular respiration to facilitate the drug-tolerant behavior of HER2+-BC and TNBC cells. Therefore, we posit USP22 as a promising therapeutic target to optimize standard therapies and combat the aggressiveness of these malignancies. Video Abstract.


Assuntos
Neoplasias de Mama Triplo Negativas , Animais , Feminino , Humanos , Camundongos , Linhagem Celular Tumoral , Respiração Celular , Modelos Animais de Doenças , Recidiva Local de Neoplasia , Neoplasias de Mama Triplo Negativas/patologia , Ubiquitina Tiolesterase
6.
PLoS Biol ; 22(2): e3002508, 2024 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-38377076

RESUMO

Peroxisomes are organelles with crucial functions in oxidative metabolism. To correctly target to peroxisomes, proteins require specialized targeting signals. A mystery in the field is the sorting of proteins that carry a targeting signal for peroxisomes and as well as for other organelles, such as mitochondria or the endoplasmic reticulum (ER). Exploring several of these proteins in fungal model systems, we observed that they can act as tethers bridging organelles together to create contact sites. We show that in Saccharomyces cerevisiae this mode of tethering involves the peroxisome import machinery, the ER-mitochondria encounter structure (ERMES) at mitochondria and the guided entry of tail-anchored proteins (GET) pathway at the ER. Our findings introduce a previously unexplored concept of how dual affinity proteins can regulate organelle attachment and communication.


Assuntos
Mitocôndrias , Peroxissomos , Retículo Endoplasmático , Movimento Celular , Respiração Celular , Saccharomyces cerevisiae
7.
Nat Commun ; 15(1): 1598, 2024 Feb 21.
Artigo em Inglês | MEDLINE | ID: mdl-38383505

RESUMO

Organic electrochemical transistors (OECTs) are ideal devices for translating biological signals into electrical readouts and have applications in bioelectronics, biosensing, and neuromorphic computing. Despite their potential, developing programmable and modular methods for living systems to interface with OECTs has proven challenging. Here we describe hybrid OECTs containing the model electroactive bacterium Shewanella oneidensis that enable the transduction of biological computations to electrical responses. Specifically, we fabricated planar p-type OECTs and demonstrated that channel de-doping is driven by extracellular electron transfer (EET) from S. oneidensis. Leveraging this mechanistic understanding and our ability to control EET flux via transcriptional regulation, we used plasmid-based Boolean logic gates to translate biological computation into current changes within the OECT. Finally, we demonstrated EET-driven changes to OECT synaptic plasticity. This work enables fundamental EET studies and OECT-based biosensing and biocomputing systems with genetically controllable and modular design elements.


Assuntos
Respiração Celular , Eletricidade , Transporte de Elétrons
8.
Redox Biol ; 71: 103037, 2024 May.
Artigo em Inglês | MEDLINE | ID: mdl-38401291

RESUMO

Mitochondrial respiration extends beyond ATP generation, with the organelle participating in many cellular and physiological processes. Parallel changes in components of the mitochondrial electron transfer system with respiration render it an appropriate hub for coordinating cellular adaption to changes in oxygen levels. How changes in respiration under functional hypoxia (i.e., when intracellular O2 levels limit mitochondrial respiration) are relayed by the electron transfer system to impact mitochondrial adaption and remodeling after hypoxic exposure remains poorly defined. This is largely due to challenges integrating findings under controlled and defined O2 levels in studies connecting functions of isolated mitochondria to humans during physical exercise. Here we present experiments under conditions of hypoxia in isolated mitochondria, myotubes and exercising humans. Performing steady-state respirometry with isolated mitochondria we found that oxygen limitation of respiration reduced electron flow and oxidative phosphorylation, lowered the mitochondrial membrane potential difference, and decreased mitochondrial calcium influx. Similarly, in myotubes under functional hypoxia mitochondrial calcium uptake decreased in response to sarcoplasmic reticulum calcium release for contraction. In both myotubes and human skeletal muscle this blunted mitochondrial adaptive responses and remodeling upon contractions. Our results suggest that by regulating calcium uptake the mitochondrial electron transfer system is a hub for coordinating cellular adaption under functional hypoxia.


Assuntos
Cálcio , Consumo de Oxigênio , Humanos , Cálcio/metabolismo , Consumo de Oxigênio/fisiologia , Respiração Celular , Hipóxia/metabolismo , Músculo Esquelético/metabolismo , Oxigênio/metabolismo
9.
Chemosphere ; 350: 141090, 2024 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-38169199

RESUMO

Nitrate pollution in fresh water is becoming increasingly serious. In this study, the effects of temperature and graphene oxide materials on the potential functions of denitrification communities in lake sediments were investigated by metagenome. The addition of graphene oxide significantly affected the abundance of denitrification genes such as Nap, Nos, and enhanced the contribution of Pseudomonas, making low temperature and material addition conducive to the denitrification process. Module network implied that low temperature increased the centrality of denitrification in community functions. At low temperatures, graphene oxide enhanced community anabolism by stimulation organic carbon consumption and regulating the gene abundance in the citric acid cycle and the semi-phosphorylation Entner-Doudoroff, thus possibly stimulating extracellular polymeric substances (EPS) synthesis and secretion. In addition, graphene oxide may also regulate the transfer of reducing electrons from NADH to denitrifying enzymes by affecting the gene abundances of complex I and complex IV.


Assuntos
Desnitrificação , Grafite , Microbiota , Temperatura , Lagos , Nitratos/farmacologia , Respiração Celular , Nitrogênio/farmacologia
10.
J Appl Physiol (1985) ; 136(3): 472-481, 2024 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-38205552

RESUMO

The integrative response to exercise differs between sexes, with oxidative energy contribution purported as a potential mechanism. The present study investigated whether this difference was evident in the kinetics of oxygen uptake (V̇o2) and extraction (HHb + Mb) during exercise. Sixteen adults (8 males, 8 females, age: 27 ± 5 yr) completed three experimental visits. Incremental exercise testing was performed to obtain lactate threshold and V̇o2peak. Subsequent visits involved three 6-min cycling bouts at 80% of lactate threshold and one 30-min bout at a work rate of 30% between the lactate threshold and power at V̇o2peak. Pulmonary gas exchange and near-infrared spectroscopy of the vastus lateralis were used to continuously sample V̇o2 and HHb + Mb, respectively. The phase II V̇o2 kinetics were quantified using monoexponential curves during moderate and heavy exercise. Slow component amplitudes were also quantified for the heavy-intensity domain. Relative V̇o2peak values were not different between sexes (P = 0.111). Males achieved ∼30% greater power outputs (P = 0.002). In the moderate- and heavy-intensity domains, the relative amplitude of the phase II transition was not different between sexes for V̇o2 (∼24 and ∼40% V̇o2peak, P ≥ 0.179) and HHb + Mb (∼20 and ∼32% ischemia, P ≥ 0.193). Similarly, there were no sex differences in the time constants for V̇o2 (∼28 s, P ≥ 0.385) or HHb + Mb (∼10 s, P ≥ 0.274). In the heavy-intensity domain, neither V̇o2 (P ≥ 0.686) or HHb + Mb (P ≥ 0.432) slow component amplitudes were different between sexes. The oxidative response to moderate- and heavy-intensity exercises did not differ between males and females, suggesting similar dynamic responses of oxidative metabolism during intensity-matched exercise.NEW & NOTEWORTHY This study demonstrated no sex differences in the oxidative response to moderate- and heavy-intensity cycling exercise. The change in oxygen uptake and deoxyhemoglobin were modeled with monoexponential curve fitting, which revealed no differences in the rate of oxidative energy provision between sexes. This provides insight into previously reported sex differences in the integrative response to exercise.


Assuntos
Ácido Láctico , Caracteres Sexuais , Adulto , Humanos , Feminino , Masculino , Adulto Jovem , Ciclismo , Respiração Celular , Oxigênio
11.
Sci Rep ; 14(1): 755, 2024 01 08.
Artigo em Inglês | MEDLINE | ID: mdl-38191891

RESUMO

Skeletal muscle is one of the largest metabolic tissues in mammals and is composed of four different types of muscle fibers (types 1, 2A, 2X, and 2B); however, type 2B is absent in humans. Given that slow-twitch fibers are superior to fast-twitch fibers in terms of oxidative metabolism and are rich in mitochondria, shift of muscle fiber types in direction towards slower fiber types improves metabolic disorders and endurance capacity. We previously had reported that oleic acid supplementation increases type 1 fiber formation in C2C12 myotubes; however, its function still remains unclear. This study aimed to determine the effect of oleic acid on the muscle fiber types and endurance capacity. An in vivo mouse model was used, and mice were fed a 10% oleic acid diet for 4 weeks. Two different skeletal muscles, slow soleus muscle with the predominance of slow-twitch fibers and fast extensor digitorum longus (EDL) muscle with the predominance of fast-twitch fibers, were used. We found that dietary oleic acid intake improved running endurance and altered fiber type composition of muscles, the proportion of type 1 and 2X fibers increased in the soleus muscle and type 2X increased in the EDL muscle. The fiber type shift in the EDL muscle was accompanied by an increased muscle TAG content. In addition, blood triacylglycerol (TAG) and non-esterified fatty acid levels decreased during exercise. These changes suggested that lipid utilization as an energy substrate was enhanced by oleic acid. Increased proliferator-activated receptor γ coactivator-1ß protein levels were observed in the EDL muscle, which potentially enhanced the fiber type transitions towards type 2X and muscle TAG content. In conclusion, dietary oleic acid intake improved running endurance with the changes of muscle fiber type shares in mice. This study elucidated a novel functionality of oleic acid in skeletal muscle fiber types. Further studies are required to elucidate the underlying mechanisms. Our findings have the potential to contribute to the field of health and sports science through nutritional approaches, such as the development of supplements aimed at improving muscle function.


Assuntos
Fibras Musculares Esqueléticas , Ácido Oleico , Humanos , Animais , Camundongos , Ácido Oleico/farmacologia , Músculo Esquelético , Respiração Celular , Suplementos Nutricionais , Mamíferos
12.
New Phytol ; 241(2): 578-591, 2024 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-37897087

RESUMO

Leaf dark respiration (Rd ) acclimates to environmental changes. However, the magnitude, controls and time scales of acclimation remain unclear and are inconsistently treated in ecosystem models. We hypothesized that Rd and Rubisco carboxylation capacity (Vcmax ) at 25°C (Rd,25 , Vcmax,25 ) are coordinated so that Rd,25 variations support Vcmax,25 at a level allowing full light use, with Vcmax,25 reflecting daytime conditions (for photosynthesis), and Rd,25 /Vcmax,25 reflecting night-time conditions (for starch degradation and sucrose export). We tested this hypothesis temporally using a 5-yr warming experiment, and spatially using an extensive field-measurement data set. We compared the results to three published alternatives: Rd,25 declines linearly with daily average prior temperature; Rd at average prior night temperatures tends towards a constant value; and Rd,25 /Vcmax,25 is constant. Our hypothesis accounted for more variation in observed Rd,25 over time (R2 = 0.74) and space (R2 = 0.68) than the alternatives. Night-time temperature dominated the seasonal time-course of Rd , with an apparent response time scale of c. 2 wk. Vcmax dominated the spatial patterns. Our acclimation hypothesis results in a smaller increase in global Rd in response to rising CO2 and warming than is projected by the two of three alternative hypotheses, and by current models.


Assuntos
Respiração Celular , Ecossistema , Fotossíntese , Folhas de Planta , Aclimatação/fisiologia , Dióxido de Carbono/metabolismo , Fotossíntese/fisiologia , Folhas de Planta/fisiologia , Plantas/metabolismo , Temperatura , Fenômenos Fisiológicos Vegetais
13.
Plant Cell Environ ; 47(2): 408-415, 2024 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-37927244

RESUMO

Establishing the temperature dependence of respiration is critical for accurate predictions of the global carbon cycle under climate change. Diurnal temperature fluctuations, or changes in substrate availability, lead to variations in leaf respiration. Additionally, recent studies hint that the thermal sensitivity of respiration could be time-dependent. However, the role for endogenous processes, independent from substrate availability, as drivers of temporal changes in the sensitivity of respiration to temperature across phylogenies has not yet been addressed. Here, we examined the diurnal variation in the response of respiration to temperatures (R-T relationship) for different lycophyte, fern, gymnosperm and angiosperm species. We tested whether time-dependent changes in the R-T relationship would impact leaf level respiration modelling. We hypothesized that interactions between endogenous processes, like the circadian clock, and leaf respiration would be independent from changes in substrate availability. Overall, we observed a time-dependent sensitivity in the R-T relationship across phylogenies, independent of temperature, that affected modelling parameters. These results are compatible with circadian gating of respiration, but further studies should analyse the possible involvement of the clock. Our results indicate time-dependent regulation of respiration might be widespread across phylogenies, and that endogenous regulation of respiration is likely affecting leaf-level respiration fluxes.


Assuntos
Aclimatação , Respiração Celular , Respiração Celular/fisiologia , Aclimatação/fisiologia , Plantas , Temperatura , Respiração , Folhas de Planta/fisiologia
14.
New Phytol ; 241(1): 52-58, 2024 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-37858976

RESUMO

Leaf day respiration (Rd ) strongly influences carbon-use efficiencies of whole plants and the global terrestrial biosphere. It has long been thought that Rd is slower than respiration in the dark at a given temperature, but measuring Rd by gas exchange remains a challenge because leaves in the light are also photosynthesizing. The Kok method and the Laisk method are widely used to estimate Rd . We highlight theoretical limitations of these popular methods, and recent progress toward their improvement by using additional information from chlorophyll fluorescence and by accounting for the photosynthetic reassimilation of respired CO2 . The latest evidence for daytime CO2 and energy release from the oxidative pentose phosphate pathway in chloroplasts appears to be important to understanding Rd .


Assuntos
Dióxido de Carbono , Respiração Celular , Dióxido de Carbono/metabolismo , Fotossíntese , Folhas de Planta/metabolismo , Respiração
15.
Cell Metab ; 36(1): 36-47, 2024 01 02.
Artigo em Inglês | MEDLINE | ID: mdl-38128528

RESUMO

Contrary to their well-known functions in nutrient breakdown, mitochondria are also important biosynthetic hubs and express an evolutionarily conserved mitochondrial fatty acid synthesis (mtFAS) pathway. mtFAS builds lipoic acid and longer saturated fatty acids, but its exact products, their ultimate destination in cells, and the cellular significance of the pathway are all active research questions. Moreover, why mitochondria need mtFAS despite their well-defined ability to import fatty acids is still unclear. The identification of patients with inborn errors of metabolism in mtFAS genes has sparked fresh research interest in the pathway. New mammalian models have provided insights into how mtFAS coordinates many aspects of oxidative mitochondrial metabolism and raise questions about its role in diseases such as obesity, diabetes, and heart failure. In this review, we discuss the products of mtFAS, their function, and the consequences of mtFAS impairment across models and in metabolic disease.


Assuntos
Mitocôndrias , Ácido Tióctico , Animais , Humanos , Mitocôndrias/metabolismo , Respiração Celular , Ácidos Graxos/metabolismo , Ácido Tióctico/metabolismo , Estresse Oxidativo , Mamíferos/metabolismo
17.
Elife ; 122023 Dec 11.
Artigo em Inglês | MEDLINE | ID: mdl-38079477

RESUMO

Based on studies with a fluorescent reporter dye, Mito Thermo Yellow (MTY), and the genetically encoded gTEMP ratiometric fluorescent temperature indicator targeted to mitochondria, the temperature of active mitochondria in four mammalian and one insect cell line was estimated to be up to 15°C above that of the external environment to which the cells were exposed. High mitochondrial temperature was maintained in the face of a variety of metabolic stresses, including substrate starvation or modification, decreased ATP demand due to inhibition of cytosolic protein synthesis, inhibition of the mitochondrial adenine nucleotide transporter and, if an auxiliary pathway for electron transfer was available via the alternative oxidase, even respiratory poisons acting downstream of oxidative phosphorylation (OXPHOS) complex I. We propose that the high temperature of active mitochondria is an inescapable consequence of the biochemistry of OXPHOS and is homeostatically maintained as a primary feature of mitochondrial metabolism.


Assuntos
Respiração Celular , Mitocôndrias , Animais , Temperatura , Mitocôndrias/metabolismo , Fosforilação Oxidativa , Regulação da Temperatura Corporal , Estresse Fisiológico , Mamíferos
18.
Biochem Soc Trans ; 51(6): 1989-2004, 2023 12 20.
Artigo em Inglês | MEDLINE | ID: mdl-38108469

RESUMO

SLC25A51 is the primary mitochondrial NAD+ transporter in humans and controls many local reactions by mediating the influx of oxidized NAD+. Intriguingly, SLC25A51 lacks several key features compared with other members in the mitochondrial carrier family, thus its molecular mechanism has been unclear. A deeper understanding would shed light on the control of cellular respiration, the citric acid cycle, and free NAD+ concentrations in mammalian mitochondria. This review discusses recent insights into the transport mechanism of SLC25A51, and in the process highlights a multitiered regulation that governs NAD+ transport. The aspects regulating SLC25A51 import activity can be categorized as contributions from (1) structural characteristics of the transporter itself, (2) its microenvironment, and (3) distinctive properties of the transported ligand. These unique mechanisms further evoke compelling new ideas for modulating the activity of this transporter, as well as new mechanistic models for the mitochondrial carrier family.


Assuntos
Mitocôndrias , NAD , Animais , Humanos , Transporte Biológico , Respiração Celular , Mamíferos/metabolismo , Mitocôndrias/metabolismo , Proteínas de Transporte da Membrana Mitocondrial/metabolismo , NAD/metabolismo
19.
Nutrients ; 15(21)2023 Oct 25.
Artigo em Inglês | MEDLINE | ID: mdl-37960182

RESUMO

PDIA3 is an endoplasmic reticulum disulfide isomerase, which is involved in the folding and trafficking of newly synthesized proteins. PDIA3 was also described as an alternative receptor for the active form of vitamin D (1,25(OH)2D3). Here, we investigated an impact of PDIA3 in mitochondrial morphology and bioenergetics in squamous cell carcinoma line A431 treated with 1,25(OH)2D3. It was observed that PDIA3 deletion resulted in changes in the morphology of mitochondria including a decrease in the percentage of mitochondrial section area, maximal diameter, and perimeter. The 1,25(OH)2D3 treatment of A431∆PDIA3 cells partially reversed the effect of PDIA3 deletion increasing aforementioned parameters; meanwhile, in A431WT cells, only an increase in mitochondrial section area was observed. Moreover, PDIA3 knockout affected mitochondrial bioenergetics and modulated STAT3 signaling. Oxygen consumption rate (OCR) was significantly increased, with no visible effect of 1,25(OH)2D3 treatment in A431∆PDIA3 cells. In the case of Extracellular Acidification Rate (ECAR), an increase was observed for glycolysis and glycolytic capacity parameters in the case of non-treated A431WT cells versus A431∆PDIA3 cells. The 1,25(OH)2D3 treatment had no significant effect on glycolytic parameters. Taken together, the presented results suggest that PDIA3 is strongly involved in the regulation of mitochondrial bioenergetics in cancerous cells and modulation of its response to 1,25(OH)2D3, possibly through STAT3.


Assuntos
Isomerases de Dissulfetos de Proteínas , Vitamina D , Vitamina D/farmacologia , Isomerases de Dissulfetos de Proteínas/genética , Isomerases de Dissulfetos de Proteínas/metabolismo , Proteínas/metabolismo , Vitaminas , Respiração Celular , Glicólise
20.
Nat Commun ; 14(1): 7525, 2023 11 18.
Artigo em Inglês | MEDLINE | ID: mdl-37980339

RESUMO

The inability to inspect metabolic activities within distinct subcellular compartments has been a major barrier to our understanding of eukaryotic cell metabolism. Previous work addressed this challenge by analyzing metabolism in isolated organelles, which grossly bias metabolic activity. Here, we describe a method for inferring physiological metabolic fluxes and metabolite concentrations in mitochondria and cytosol based on isotope tracing experiments performed with intact cells. This is made possible by computational deconvolution of metabolite isotopic labeling patterns and concentrations into cytosolic and mitochondrial counterparts, coupled with metabolic and thermodynamic modelling. Our approach lowers the uncertainty regarding compartmentalized fluxes and concentrations by one and three orders of magnitude compared to existing modelling approaches, respectively. We derive a quantitative view of mitochondrial and cytosolic metabolic activities in central carbon metabolism across cultured cell lines without performing cell fractionation, finding major variability in compartmentalized malate-aspartate shuttle fluxes. We expect our approach for inferring metabolism at a subcellular resolution to be instrumental for a variety of studies of metabolic dysfunction in human disease and for bioengineering.


Assuntos
Respiração Celular , Mitocôndrias , Humanos , Citosol/metabolismo , Mitocôndrias/metabolismo , Linhagem Celular , Isótopos/metabolismo , Marcação por Isótopo
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