Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 1.117
Filtrar
1.
Int J Mol Sci ; 25(5)2024 Mar 06.
Artigo em Inglês | MEDLINE | ID: mdl-38474278

RESUMO

The small GTPase RAS acts as a plasma membrane-anchored intracellular neurotrophin counteracting neuronal degeneration in the brain, but the underlying molecular mechanisms are largely unknown. In transgenic mice expressing constitutively activated V12-Ha-RAS selectively in neurons, proteome analysis uncovered a 70% decrease in voltage-dependent anion channel-1 (VDAC-1) in the cortex and hippocampus. We observed a corresponding reduction in the levels of mRNA splicing variant coding for plasma membrane-targeted VDAC-1 (pl-VDAC-1) while mRNA levels for mitochondrial membrane VDAC-1 (mt-VDAC-1) remained constant. In primary cortical neurons derived from V12-Ha-RAS animals, a decrease in pl-VDAC-1 mRNA levels was observed, accompanied by a concomitant reduction in the ferricyanide reductase activity associated with VDAC-1 protein. Application of MEK inhibitor U0126 to transgenic cortical neurons reconstituted pl-VDAC-1 mRNA to reach wild-type levels. Excitotoxic glutamate-induced cell death was strongly attenuated in transgenic V12-Ha-RAS overexpressing cortical cultures. Consistently, a neuroprotective effect could also be achieved in wild-type cortical cultures by the extracellular application of channel-blocking antibody targeting the N-terminus of VDAC-1. These results may encourage novel therapeutic approaches toward blocking pl-VDAC-1 by monoclonal antibody targeting for complementary treatments in transplantation and neurodegenerative disease.


Assuntos
Doenças Neurodegenerativas , Canais de Ânion Dependentes de Voltagem , Camundongos , Animais , Canais de Ânion Dependentes de Voltagem/metabolismo , Neuroproteção , Doenças Neurodegenerativas/metabolismo , Proteínas ras/metabolismo , Regulação para Baixo , Canal de Ânion 1 Dependente de Voltagem/metabolismo , Membrana Celular/metabolismo , Camundongos Transgênicos , RNA Mensageiro/metabolismo , Canal de Ânion 2 Dependente de Voltagem/metabolismo
2.
Arch Biochem Biophys ; 753: 109914, 2024 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-38290597

RESUMO

The mitochondrial outer membrane protein porin 1 (Por1), the yeast orthologue of mammalian voltage-dependent anion channel (VDAC), is the major permeability pathway for the flux of metabolites and ions between cytosol and mitochondria. In yeast, several Por1 phosphorylation sites have been identified. Protein phosphorylation is a major modification regulating a variety of biological activities, but the potential biological roles of Por1 phosphorylation remains unaddressed. In this work, we analysed 10 experimentally observed phosphorylation sites in yeast Por1 using bioinformatics tools. Two of the residues, T100 and S133, predicted to reduce and increase pore permeability, respectively, were validated using biological assays. In accordance, Por1T100D reduced mitochondrial respiration, while Por1S133E phosphomimetic mutant increased it. Por1T100A expression also improved respiratory growth, while Por1S133A caused defects in all growth conditions tested, notably in fermenting media. In conclusion, we found phosphorylation has the potential to modulate Por1, causing a marked effect on mitochondrial function. It can also impact on cell morphology and growth both in respiratory and, unpredictably, also in fermenting conditions, expanding our knowledge on the role of Por1 in cell physiology.


Assuntos
Proteínas de Saccharomyces cerevisiae , Saccharomyces cerevisiae , Animais , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Fosforilação , Canais de Ânion Dependentes de Voltagem/metabolismo , Mitocôndrias/metabolismo , Proteínas de Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/metabolismo , Mamíferos/metabolismo
3.
J Biol Chem ; 300(2): 105632, 2024 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-38199573

RESUMO

We previously reported that bakuchiol, a phenolic isoprenoid anticancer compound, and its analogs exert anti-influenza activity. However, the proteins targeted by bakuchiol remain unclear. Here, we investigated the chemical structures responsible for the anti-influenza activity of bakuchiol and found that all functional groups and C6 chirality of bakuchiol were required for its anti-influenza activity. Based on these results, we synthesized a molecular probe containing a biotin tag bound to the C1 position of bakuchiol. With this probe, we performed a pulldown assay for Madin-Darby canine kidney cell lysates and purified the specific bakuchiol-binding proteins with SDS-PAGE. Using nanoLC-MS/MS analysis, we identified prohibitin (PHB) 2, voltage-dependent anion channel (VDAC) 1, and VDAC2 as binding proteins of bakuchiol. We confirmed the binding of bakuchiol to PHB1, PHB2, and VDAC2 in vitro using Western blot analysis. Immunofluorescence analysis showed that bakuchiol was bound to PHBs and VDAC2 in cells and colocalized in the mitochondria. The knockdown of PHBs or VDAC2 by transfection with specific siRNAs, along with bakuchiol cotreatment, led to significantly reduced influenza nucleoprotein expression levels and viral titers in the conditioned medium of virus-infected Madin-Darby canine kidney cells, compared to the levels observed with transfection or treatment alone. These findings indicate that reducing PHBs or VDAC2 protein, combined with bakuchiol treatment, additively suppressed the growth of influenza virus. Our findings indicate that bakuchiol exerts anti-influenza activity via a novel mechanism involving these mitochondrial proteins, providing new insight for developing anti-influenza agents.


Assuntos
Antivirais , Influenza Humana , Fenóis , Animais , Cães , Humanos , Antivirais/farmacologia , Antivirais/química , Proteínas Mitocondriais/metabolismo , Proibitinas , Espectrometria de Massas em Tandem , Canal de Ânion 1 Dependente de Voltagem , Canal de Ânion 2 Dependente de Voltagem/metabolismo , Canais de Ânion Dependentes de Voltagem , Linhagem Celular
4.
Plant Physiol ; 194(2): 1041-1058, 2024 Jan 31.
Artigo em Inglês | MEDLINE | ID: mdl-37772952

RESUMO

In Arabidopsis (Arabidopsis thaliana), stomatal closure mediated by abscisic acid (ABA) is redundantly controlled by ABA receptor family proteins (PYRABACTIN RESISTANCE 1 [PYR1]/PYR1-LIKE [PYLs]) and subclass III SUCROSE NONFERMENTING 1 (SNF1)-RELATED PROTEIN KINASES 2 (SnRK2s). Among these proteins, the roles of PYR1, PYL2, and SnRK2.6 are more dominant. A recent discovery showed that ABA-induced accumulation of reactive oxygen species (ROS) in mitochondria promotes stomatal closure. By analyzing stomatal movements in an array of single and higher order mutants, we revealed that the mitochondrial protein VOLTAGE-DEPENDENT ANION CHANNEL 3 (VDAC3) jointly regulates ABA-mediated stomatal closure with a specialized set of PYLs and SnRK2s by affecting cellular and mitochondrial ROS accumulation. VDAC3 interacted with 9 PYLs and all 3 subclass III SnRK2s. Single mutation in VDAC3, PYLs (except PYR1 and PYL2), or SnRK2.2/2.3 had little effect on ABA-mediated stomatal closure. However, knocking out PYR1, PYL1/2/4/8, or SnRK2.2/2.3 in vdac3 mutants resulted in significantly delayed or attenuated ABA-mediated stomatal closure, despite the presence of other PYLs or SnRK2s conferring redundant functions. We found that cellular and mitochondrial accumulation of ROS induced by ABA was altered in vdac3pyl1 mutants. Moreover, H2O2 treatment restored ABA-induced stomatal closure in mutants with decreased stomatal sensitivity to ABA. Our work reveals that VDAC3 ensures redundant control of ABA-mediated stomatal closure by canonical ABA signaling components.


Assuntos
Proteínas de Arabidopsis , Arabidopsis , Ácido Abscísico/farmacologia , Ácido Abscísico/metabolismo , Proteínas de Arabidopsis/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Peróxido de Hidrogênio/metabolismo , Estômatos de Plantas/metabolismo , Arabidopsis/metabolismo , Canais de Ânion Dependentes de Voltagem/metabolismo , Mitocôndrias/metabolismo
5.
Plant Physiol Biochem ; 206: 108237, 2024 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-38109831

RESUMO

Pathogen severely affects plant mitochondrial processes including respiration, however, the roles and mechanism of mitochondrial protein during the immune response remain largely unexplored. The interplay of plant hormone signaling during defense is an outcome of plant pathogen interaction. We recently discovered that the Arabidopsis calcineurin B-like interacting protein kinase 9 (AtCIPK9) interacts with the voltage-dependent anion channel 3 (AtVDAC3) and inhibits MV-induced oxidative damage. Here we report the characterization of AtVDAC3 in an antagonistic interaction pathway between abscisic acid (ABA) and salicylic acid (SA) signaling in Pseudomonas syringae -Arabidopsis interaction. In this study, we observed that mutants of AtVDAC3 were highly susceptible to Pseudomonas syringae infection as compared to the wild type (WT) Arabidopsis plants. Transcripts of VDAC3 and CIPK9 were inducible upon ABA application. Following pathogen exposure, expression analyses of ABA and SA biosynthesis genes indicated that the function of VDAC3 is required for isochorisimate synthase 1 (ICS1) expression but not for Nine-cis-epoxycaotenoid dioxygenase 3 (NCED3) expression. Despite the fact that vdac3 mutants had increased NCED3 expression in response to pathogen challenge, transcripts of ABA sensitive genes such as AtRD22 and AtRAB18 were downregulated even after exogenous ABA application. VDAC3 is required for ABA responsive genes expression upon exogenous ABA application. We also found that Pseudomonas syringae-induced SA signaling is downregulated in vdac3 mutants since overexpression of VDAC3 resulted in hyperaccumulation of Pathogenesis related gene1 (PR1) transcript. Interestingly, ABA application prior to P. syringae inoculation resulted in the upregulation of ABA responsive genes like Responsive to ABA18 (RAB18) and Responsive to dehydration 22 (RD22). Intriguingly, in the absence of AtVDAC3, Pst challenge can dramatically increase ABA-induced RD22 and RAB18 expression. Altogether our results reveal a novel Pathogen-SA-ABA interaction pathway in plants. Our findings show that ABA plays a significant role in modifying plant-pathogen interactions, owing to cross-talk with the biotic stress signaling pathways of ABA and SA.


Assuntos
Proteínas de Arabidopsis , Arabidopsis , Dioxigenases , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Ácido Salicílico/farmacologia , Ácido Salicílico/metabolismo , Reguladores de Crescimento de Plantas/metabolismo , Ácido Abscísico/metabolismo , Dioxigenases/genética , Canais de Ânion Dependentes de Voltagem/genética , Canais de Ânion Dependentes de Voltagem/metabolismo , Pseudomonas syringae/fisiologia , Doenças das Plantas/genética , Regulação da Expressão Gênica de Plantas , Proteínas Serina-Treonina Quinases/metabolismo
6.
J Mol Biol ; 436(4): 168432, 2024 02 15.
Artigo em Inglês | MEDLINE | ID: mdl-38161000

RESUMO

Helicobacter pylori colonizes the stomach in about half of the human population, leading to an increased risk of peptic ulcer disease and gastric cancer. H. pylori secretes an 88 kDa VacA toxin that contributes to pathogenesis. VacA assembles into oligomeric complexes in solution and forms anion-selective channels in cell membranes. Cryo-electron microscopy (cryo-EM) analyses of VacA oligomers in solution provided insights into VacA oligomerization but failed to reveal the structure of the hydrophobic N-terminal region predicted to be a pore-forming domain. In this study, we incubated VacA with liposomes and used single particle cryo-EM to analyze detergent-extracted VacA oligomers. A 3D structure of detergent-solubilized VacA hexamers revealed the presence of six α-helices extending from the center of the oligomers, a feature not observed in previous studies of water-soluble VacA oligomers. Cryo-electron tomography analysis and 2D averages of VacA associated with liposomes confirmed that central regions of the membrane-associated VacA oligomers can insert into the lipid bilayer. However, insertion is heterogenous, with some membrane-associated oligomers appearing only partially inserted and others sitting on top of the bilayer. These studies indicate that VacA undergoes a conformational change when contacting the membrane and reveal an α-helical region positioned to extend into the membrane. Although the reported VacA 3D structure does not represent a selective anion channel, our combined single particle 3D analysis, cryo-electron tomography, and modeling allow us to propose a model for the structural organization of the VacA N-terminus in the context of a hexamer as it inserts into the membrane.


Assuntos
Proteínas de Bactérias , Helicobacter pylori , Toxinas Biológicas , Canais de Ânion Dependentes de Voltagem , Humanos , Proteínas de Bactérias/química , Microscopia Crioeletrônica/métodos , Detergentes , Helicobacter pylori/química , Lipossomos/química , Toxinas Biológicas/química , Canais de Ânion Dependentes de Voltagem/química , Multimerização Proteica
7.
Nat Commun ; 14(1): 8115, 2023 Dec 08.
Artigo em Inglês | MEDLINE | ID: mdl-38065946

RESUMO

Mitochondria are double-membrane-bounded organelles that depend critically on phospholipids supplied by the endoplasmic reticulum. These lipids must cross the outer membrane to support mitochondrial function, but how they do this is unclear. We identify the Voltage Dependent Anion Channel (VDAC), an abundant outer membrane protein, as a scramblase-type lipid transporter that catalyzes lipid entry. On reconstitution into membrane vesicles, dimers of human VDAC1 and VDAC2 catalyze rapid transbilayer translocation of phospholipids by a mechanism that is unrelated to their channel activity. Coarse-grained molecular dynamics simulations of VDAC1 reveal that lipid scrambling occurs at a specific dimer interface where polar residues induce large water defects and bilayer thinning. The rate of phospholipid import into yeast mitochondria is an order of magnitude lower in the absence of VDAC homologs, indicating that VDACs provide the main pathway for lipid entry. Thus, VDAC isoforms, members of a superfamily of beta barrel proteins, moonlight as a class of phospholipid scramblases - distinct from alpha-helical scramblase proteins - that act to import lipids into mitochondria.


Assuntos
Fosfolipídeos , Canal de Ânion 1 Dependente de Voltagem , Humanos , Canal de Ânion 1 Dependente de Voltagem/metabolismo , Fosfolipídeos/metabolismo , Canais de Ânion Dependentes de Voltagem/metabolismo , Mitocôndrias/metabolismo , Saccharomyces cerevisiae/metabolismo
8.
Redox Biol ; 68: 102959, 2023 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-37977042

RESUMO

Colorectal cancer (CRC) is a common and deadly disease of the digestive system, but its targeted therapy is hampered by the lack of reliable and specific biomarkers. Hence, discovering new therapeutic targets and agents for CRC is an urgent and challenging task. Here we report that carnitine palmitoyltransferase 1A (CPT1A), a mitochondrial enzyme that catalyzes fatty acid oxidation (FAO), is a potential target for CRC treatment. We show that CPT1A is overexpressed in CRC cells and that its inhibition by a secolignan-type compound, 2,6-dihydroxypeperomin B (DHP-B), isolated from the plant Peperomia dindygulensis, suppresses tumor cell growth and induces apoptosis. We demonstrate that DHP-B covalently binds to Cys96 of CPT1A, blocks FAO, and disrupts the mitochondrial CPT1A-VDAC1 interaction, leading to increased mitochondrial permeability and reduced oxygen consumption and energy metabolism in CRC cells. We also reveal that CPT1A expression correlates with the survival of tumor-bearing animals and that DHP-B exhibits anti-CRC activity in vitro and in vivo. Our study uncovers the molecular mechanism of DHP-B as a novel CPT1A inhibitor and provides a rationale for its preclinical development as well as a new strategy for CRC targeted therapy.


Assuntos
Carnitina O-Palmitoiltransferase , Neoplasias Colorretais , Animais , Apoptose , Carnitina O-Palmitoiltransferase/antagonistas & inibidores , Carnitina O-Palmitoiltransferase/genética , Carnitina O-Palmitoiltransferase/metabolismo , Neoplasias Colorretais/tratamento farmacológico , Neoplasias Colorretais/genética , Ácidos Graxos/metabolismo , Metabolismo dos Lipídeos , Oxirredução , Canais de Ânion Dependentes de Voltagem/metabolismo
9.
Gene ; 887: 147784, 2023 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-37689223

RESUMO

Acrosome is inextricably related to membranous organelles. The origin of acrosome is still controversial, one reason is that limited articles were reported about the proteomic analysis of the acrosome. Mitochondrial proteins were found exist in the acrosome, nevertheless, only limited attention has been paid to the function of mitochondrial proteins in the acrosome formation. Eriocheir sinensis sperm has a large acrosome, which makes it an ideal model to study acrosome formation. Here, we firstly compared the rate of acrosome reaction induced by the calcium ionophore A23187 and ionomycin. The rate of acrosome reaction induced by ionomycin is higher (95.8%) than A23187 (58.7%). Morphological changes were observed using light, confocal and transmission electron microscopy. Further more, proteins released during the acrosome reaction as induced by ionomycin were collected for LC-MS/MS analysis. A total of 945 proteins, including malate dehydrogenase (MDH) and voltage-dependent anion channel 3 (VDAC3), were identified in the acrosomal released proteome. The number of proteins from mitochondria (17.57%) was higher compared with endoplasmic reituculum (1.59%) and lysosomes (1.8%). To investigate the functions of target mitochondrial proteins during spermatogenesis, poly-antibodies of MDH in E. sinensis were prepared. The characteristics, further analyzed using immunofluorescence, of two mitochondrial proteins during acrosome formation showed that MDH and VDAC3 were independently involved in the formation of acrosomal membrane. These findings illustrate the acrosomal released proteome and provide important data resource for understanding the relationship between mitochondria and the acrosome in Decapoda crustacean.


Assuntos
Malato Desidrogenase , Proteoma , Masculino , Humanos , Acrossomo , Calcimicina , Cromatografia Líquida , Ionomicina , Proteômica , Sêmen , Espectrometria de Massas em Tandem , Espermatozoides , Espermatogênese , Mitocôndrias , Proteínas Mitocondriais , Canais de Ânion Dependentes de Voltagem , Lisossomos
10.
J Steroid Biochem Mol Biol ; 234: 106400, 2023 11.
Artigo em Inglês | MEDLINE | ID: mdl-37722462

RESUMO

Steroidogenesis machinery involves the steroidogenic acute regulatory protein (StAR), which regulates cholesterol transfer within the mitochondria, and the transport of cholesterol via a channel composed of 18-kDa translocator protein (TSPO), the voltage-dependent anion channel (VDAC) plus some accessory proteins. In this study, we investigated the immunolocalizations and expressions of StAR, TSPO, VDAC and cytochrome P450 side chain cleavage enzyme (P450scc, CYP11A1) in the scent glands of muskrats (Ondatra zibethicus) during the breeding and non-breeding periods. StAR, TSPO, VDAC and CYP11A1 were immunolocalized in the scent glandular, interstitial and epithelial cells in both breeding and non-breeding seasons with stronger immunostaining in the breeding season. The mRNA expression levels of StAR, TSPO, VDAC and CYP11A1 were higher in the scent glands of the breeding season than those of the non-breeding season. The circulating follicle stimulating hormone (FSH), luteinizing hormone (LH) and testosterone (T) as well as scent glandular T and dihydrotestosterone (DHT) concentrations were also significantly higher in the breeding season. Additionally, the transcriptomic study in the scent glands identified that differentially expressed genes might be related to the lipid metabolic process, integral component of membrane, and steroid hormone receptor activity and hormone activity using GO analysis. Further in vitro study verified that StAR, TSPO, VDAC and CYP11A1 expression levels increased significantly after human chorionic gonadotropin, hCG/FSH treatment compared with the control group. The KEGG pathway enriched by differentially expressed genes detected to be involved in endocrine system or amino acid metabolism. These findings suggested that the scent glands of the muskrats have ability to synthesize steroids de novo, and that the steroid hormones may have an important regulatory role in the scent glandular function via an autocrine/paracrine pathway.


Assuntos
Arvicolinae , Glândulas Odoríferas , Animais , Humanos , Estações do Ano , Arvicolinae/metabolismo , Glândulas Odoríferas/metabolismo , Enzima de Clivagem da Cadeia Lateral do Colesterol/genética , Enzima de Clivagem da Cadeia Lateral do Colesterol/metabolismo , Testosterona/metabolismo , Fosfoproteínas/genética , Fosfoproteínas/metabolismo , Canais de Ânion Dependentes de Voltagem/metabolismo , Colesterol/metabolismo , Hormônio Foliculoestimulante/metabolismo , Receptores de GABA/genética , Receptores de GABA/metabolismo
11.
Eur J Pharmacol ; 957: 176034, 2023 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-37652292

RESUMO

Voltage dependent anion channels (VDAC) in the outer mitochondrial membrane regulate the influx of metabolites that sustain mitochondrial metabolism and the efflux of ATP to the cytosol. Free tubulin and NADH close VDAC. The VDAC-binding small molecules X1 and SC18 modulate mitochondrial metabolism. X1 antagonizes the inhibitory effect of tubulin on VDAC. SC18 occupies an NADH-binding pocket in the inner wall of all VDAC isoforms. Here, we hypothesized that X1 and SC18 have a synergistic effect with sorafenib, regorafenib or lenvatinib to arrest proliferation and induce death in hepatocarcinoma cells. We used colony formation assays to determine cell proliferation, and a combination of calcein/propidium iodide, and trypan blue exclusion to assess cell death in the well differentiated Huh7 and the poorly differentiated SNU-449 cells. Synergism was assessed using the Chou-Talalay method. The inhibitory effect of X1, SC18, sorafenib, regorafenib and lenvatinib was concentration and time dependent. IC50s calculated from the inhibition of clonogenic capacity were lower than those determined from cell survival. At IC50s that inhibited cell proliferation, SC18 arrested cells in G0/G1. SC18 at 0.25-2 IC50s had a synergistic effect with sorafenib on clonogenic inhibition in Huh7 and SNU-449 cells, and with regorafenib or lenvatinib in SNU-449 cells. X1 or SC18 also had synergistic effects with sorafenib on promoting cell death at 0.5-2 IC50s for SC18 in Huh7 and SNU-449 cells. These results suggest that small molecules targeting VDAC represent a potential new class of drugs to treat liver cancer.


Assuntos
Carcinoma Hepatocelular , NAD , Humanos , Sorafenibe/farmacologia , Tubulina (Proteína) , Carcinoma Hepatocelular/tratamento farmacológico , Proliferação de Células , Canais de Ânion Dependentes de Voltagem
12.
J Gen Physiol ; 155(10)2023 10 02.
Artigo em Inglês | MEDLINE | ID: mdl-37555782

RESUMO

Using optical and electrical methods, we document that diffusion in the cytoplasm of BL6 murine cardiomyocytes becomes restricted >20-fold as molecular weight increases from 30 to 2,000, roughly as expected for pores with porin channel dimensions. Bodipy-FL ATP diffuses >40-fold slower than in free water at 25°C. From several fluorophores analyzed, bound fluorophore fractions range from 0.1 for a 2 kD FITC-labeled polyethylene glycol to 0.93 for sulforhodamine. Unbound fluorophores diffuse at 0.5-8 × 10-7 cm2/s (5-80 µm2/s). Analysis of Na/K pump and veratridine-modified Na channel currents suggests that Na diffusion is nearly unrestricted at 35°C (time constant for equilibration with the pipette tip, ∼20 s). Using multiple strategies, we estimate that at 35°C, ATP diffuses four to eight times slower than in free water. To address whether restrictions are caused more by protein or membrane networks, we verified first that a protein gel, 10 g% gelatin, restricts diffusion with strong dependence on molecular weight. Solute diffusion in membrane-extracted cardiac myofilaments, confined laterally by suction into large-diameter pipette tips, is less restricted than in intact myocytes. Notably, myofilaments extracted similarly from skeletal (diaphragm) myocytes are less restrictive. Solute diffusion in myocytes with sarcolemma permeabilized by ß-escin (80 µM) is similar to diffusion in intact myocytes. Restrictions are strain-dependent, being twofold greater in BL6 myocytes than in CD1/J6/129svJ myocytes. Furthermore, longitudinal diffusion is 2.5-fold more restricted in CD1/J6/129svJ myocytes lacking the mitochondrial porin, VDAC1, than in WT CD1/J6/129svJ myocytes. Thus, mitochondria networks restrict long-range diffusion while presumably optimizing nucleotide transfer between myofilaments and mitochondria. We project that diffusion restrictions imposed by both myofilaments and the outer mitochondrial membrane are important determinants of total free cytoplasmic AMP and ADP (∼10 µM). However, the capacity of diffusion to deliver ATP to myofilaments remains ∼100-fold greater than ATP consumption.


Assuntos
Miócitos Cardíacos , Miofibrilas , Camundongos , Animais , Miócitos Cardíacos/metabolismo , Miofibrilas/metabolismo , Mitocôndrias/metabolismo , Difusão , Canais de Ânion Dependentes de Voltagem/metabolismo , Trifosfato de Adenosina/metabolismo , Água/metabolismo
13.
Biol Res ; 56(1): 33, 2023 Jun 22.
Artigo em Inglês | MEDLINE | ID: mdl-37344914

RESUMO

BACKGROUND: Voltage-dependent anion selective channels (VDACs) are the most abundant mitochondrial outer membrane proteins, encoded in mammals by three genes, VDAC1, 2 and 3, mostly ubiquitously expressed. As 'mitochondrial gatekeepers', VDACs control organelle and cell metabolism and are involved in many diseases. Despite the presence of numerous VDAC pseudogenes in the human genome, their significance and possible role in VDAC protein expression has not yet been considered. RESULTS: We investigated the relevance of processed pseudogenes of human VDAC genes, both in physiological and in pathological contexts. Using high-throughput tools and querying many genomic and transcriptomic databases, we show that some VDAC pseudogenes are transcribed in specific tissues and pathological contexts. The obtained experimental data confirm an association of the VDAC1P8 pseudogene with acute myeloid leukemia (AML). CONCLUSIONS: Our in-silico comparative analysis between the VDAC1 gene and its VDAC1P8 pseudogene, together with experimental data produced in AML cellular models, indicate a specific over-expression of the VDAC1P8 pseudogene in AML, correlated with a downregulation of the parental VDAC1 gene.


Assuntos
Leucemia Mieloide Aguda , Pseudogenes , Canais de Ânion Dependentes de Voltagem , Humanos , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/metabolismo , Mitocôndrias , Pseudogenes/genética , Transcriptoma , Canais de Ânion Dependentes de Voltagem/genética , Canais de Ânion Dependentes de Voltagem/metabolismo
14.
Sci Immunol ; 8(84): eade7652, 2023 06 23.
Artigo em Inglês | MEDLINE | ID: mdl-37327321

RESUMO

NLRP3 inflammasome activation is a highly regulated process for controlling secretion of the potent inflammatory cytokines IL-1ß and IL-18 that are essential during bacterial infection, sterile inflammation, and disease, including colitis, diabetes, Alzheimer's disease, and atherosclerosis. Diverse stimuli activate the NLRP3 inflammasome, and unifying upstream signals has been challenging to identify. Here, we report that a common upstream step in NLRP3 inflammasome activation is the dissociation of the glycolytic enzyme hexokinase 2 from the voltage-dependent anion channel (VDAC) in the outer membrane of mitochondria. Hexokinase 2 dissociation from VDAC triggers activation of inositol triphosphate receptors, leading to release of calcium from the ER, which is taken up by mitochondria. This influx of calcium into mitochondria leads to oligomerization of VDAC, which is known to form a macromolecule-sized pore in the outer membranes of mitochondria that allows proteins and mitochondrial DNA (mtDNA), often associated with apoptosis and inflammation, respectively, to exit the mitochondria. We observe that VDAC oligomers aggregate with NLRP3 during initial assembly of the multiprotein oligomeric NLRP3 inflammasome complex. We also find that mtDNA is necessary for NLRP3 association with VDAC oligomers. These data, together with other recent work, help to paint a more complete picture of the pathway leading to NLRP3 inflammasome activation.


Assuntos
Inflamassomos , Proteína 3 que Contém Domínio de Pirina da Família NLR , Humanos , Inflamassomos/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Hexoquinase/metabolismo , Cálcio/metabolismo , Mitocôndrias/metabolismo , Canais de Ânion Dependentes de Voltagem/metabolismo , DNA Mitocondrial/metabolismo , Inflamação/metabolismo
15.
Rev Med Virol ; 33(4): e2453, 2023 07.
Artigo em Inglês | MEDLINE | ID: mdl-37170417

RESUMO

Viruses control the host cell by exploiting its molecular machinery to facilitate viral replication and propagation. Understanding different viral mechanisms and biochemical pathways is crucial for finding promising therapeutic solutions to viral infections. The mitochondrion is a vital organelle targeted by various types of viruses. More specifically, viruses interact with the voltage-dependent anion channel (VDAC), a porin protein found in the outer mitochondrial membrane. VDAC controls metabolite flux, regulates reactive oxygen species production, and promotes mitochondrial-mediated apoptosis by releasing pro-apoptotic proteins. Hence, a common pathogenic strategy used by many viruses seems to exploit natural pathways that VDAC regulates. This review aims to address the inhibition and enhancement roles of VDAC in viral pathogenesis and outlines multiple links and interactions between VDAC and viral proteins as potential antiviral targets.


Assuntos
Viroses , Canais de Ânion Dependentes de Voltagem , Humanos , Canais de Ânion Dependentes de Voltagem/metabolismo , Mitocôndrias/metabolismo , Apoptose , Proteínas Virais/metabolismo , Viroses/metabolismo
16.
PLoS One ; 18(5): e0273882, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-37130143

RESUMO

Mitochondrial decline is a key feature of ageing. The retina has more mitochondria than any other tissue and ages rapidly. To understand human retinal ageing it is critical to examine old world primates that have similar visual systems to humans, and do so across central and peripheral regions, as there is evidence for early central decline. Hence, we examine mitochondrial metrics in young and ageing Macaca fascicularis retinae. In spite of reduced ATP with age, primate mitochondrial complex activity did not decline. But mitochondrial membrane potentials were reduced significantly, and concomitantly, mitochondrial membrane permeability increased. The mitochondrial marker Tom20 declined significantly, consistent with reduced mitochondria number, while VDAC, a voltage dependent anion channel and diffusion pore associated with apoptosis increased significantly. In spite of these clear age-related changes, there was almost no evidence for regional differences between the centre and the periphery in these mitochondrial metrics. Primate cones do not die with age, but many showed marked structural decline with vacuous spaces in proximal inner segments normally occupied by endoplasmic reticulum (ER), that regulate mitochondrial autophagy. In many peripheral cones, ER was displaced by the nucleus that transposed across the outer limiting membrane and could become embedded in mitochondrial populations. These data are consistent with significant changes in retinal mitochondria in old world primate ageing but provide little if any evidence that aged central mitochondria suffer more than those in the periphery.


Assuntos
Mitocôndrias , Retina , Animais , Humanos , Idoso , Retina/metabolismo , Envelhecimento/fisiologia , Células Fotorreceptoras Retinianas Cones/metabolismo , Primatas , Macaca fascicularis , Canais de Ânion Dependentes de Voltagem/metabolismo
17.
Biochim Biophys Acta Biomembr ; 1865(6): 184175, 2023 08.
Artigo em Inglês | MEDLINE | ID: mdl-37201560

RESUMO

Mitochondria, composed of two membranes, play a key role in energy production in eukaryotic cells. The main function of the inner membrane is oxidative phosphorylation, while the mitochondrial outer membrane (MOM) seems to control the energy flux and exchange of various charged metabolites between mitochondria and the cytosol. Metabolites cross MOM via the various isoforms of voltage-dependent anion channel (VDAC). In turn, VDACs interact with some enzymes, other proteins and molecules, including drugs. This work aimed to analyze various literature experimental data related to targeting mitochondrial VDACs and VDAC-kinase complexes on the basis of the hypothesis of generation of the outer membrane potential (OMP) and OMP-dependent reprogramming of cell energy metabolism. Our previous model of the VDAC-hexokinase-linked generation of OMP was further complemented in this study with an additional regulation of the MOM permeability by the OMP-dependent docking of cytosolic proteins like tubulin to VDACs. Computational analysis of the model suggests that OMP changes might be involved in the mechanisms of apoptosis promotion through the so-called transient hyperpolarization of mitochondria. The high concordance of the performed computational estimations with many published experimental data allows concluding that OMP generation under physiological conditions is highly probable and VDAC might function as an OMP-dependent gatekeeper of mitochondria, controlling cell life and death. The proposed model of OMP generation allows understanding in more detail the mechanisms of cancer death resistance and anticancer action of various drugs and treatments influencing VDAC voltage-gating properties, VDAC content, mitochondrial hexokinase activity and VDAC-kinase interactions in MOM.


Assuntos
Hexoquinase , Mitocôndrias , Hexoquinase/metabolismo , Mitocôndrias/metabolismo , Membranas Mitocondriais/metabolismo , Metabolismo Energético , Canais de Ânion Dependentes de Voltagem/metabolismo
18.
J Phys Chem B ; 127(15): 3372-3381, 2023 04 20.
Artigo em Inglês | MEDLINE | ID: mdl-37040575

RESUMO

Ca2+/calmodulin-dependent protein kinase II (CaMKII) is a key enzyme that plays a significant role in intracellular signaling and the modulation of mitochondrial membrane properties. It is known that the voltage-dependent anion channel (VDAC) is one of the most abundant outer mitochondrial membrane (OMM) proteins acting as a significant passageway and regulatory site for various enzymes, proteins, ions, and metabolites. Considering this, we hypothesize that VDAC could be one of the targets for CaMKII enzymatic activity. Our in vitro experiments indicate that VDAC can be phosphorylated by the CaMKII enzyme. Moreover, the bilayer electrophysiology experimental data indicate that CaMKII significantly reduces VDAC's single-channel conductivity; its open probability remains high at all the applied potentials between +60 and -60 mV, and the voltage dependency was lost, which suggests that CaMKII disrupted the VDAC's single-channel activities. Hence, we can infer that VDAC interacts with CaMKII and thus acts as a vital target for its activity. Furthermore, our findings suggest that CaMKII could play a significant role during the transport of ions and metabolites across the outer mitochondrial membrane (OMM) through VDAC and thus regulate apoptotic events.


Assuntos
Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina , Bicamadas Lipídicas , Bicamadas Lipídicas/metabolismo , Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina/metabolismo , Canais de Ânion Dependentes de Voltagem/metabolismo , Membranas Mitocondriais/metabolismo , Fosforilação
19.
Dev Cell ; 58(7): 597-615.e10, 2023 04 10.
Artigo em Inglês | MEDLINE | ID: mdl-37040696

RESUMO

Loss of fragile X messenger ribonucleoprotein (FMRP) causes fragile X syndrome (FXS), the most prevalent form of inherited intellectual disability. Here, we show that FMRP interacts with the voltage-dependent anion channel (VDAC) to regulate the formation and function of endoplasmic reticulum (ER)-mitochondria contact sites (ERMCSs), structures that are critical for mitochondrial calcium (mito-Ca2+) homeostasis. FMRP-deficient cells feature excessive ERMCS formation and ER-to-mitochondria Ca2+ transfer. Genetic and pharmacological inhibition of VDAC or other ERMCS components restored synaptic structure, function, and plasticity and rescued locomotion and cognitive deficits of the Drosophila dFmr1 mutant. Expressing FMRP C-terminal domain (FMRP-C), which confers FMRP-VDAC interaction, rescued the ERMCS formation and mito-Ca2+ homeostasis defects in FXS patient iPSC-derived neurons and locomotion and cognitive deficits in Fmr1 knockout mice. These results identify altered ERMCS formation and mito-Ca2+ homeostasis as contributors to FXS and offer potential therapeutic targets.


Assuntos
Proteínas de Drosophila , Síndrome do Cromossomo X Frágil , Animais , Camundongos , Síndrome do Cromossomo X Frágil/genética , Síndrome do Cromossomo X Frágil/metabolismo , Cálcio/metabolismo , Proteína do X Frágil de Retardo Mental/genética , Proteína do X Frágil de Retardo Mental/metabolismo , Drosophila/metabolismo , Camundongos Knockout , Homeostase , Mitocôndrias/metabolismo , Retículo Endoplasmático/metabolismo , Canais de Ânion Dependentes de Voltagem/metabolismo , Proteínas de Drosophila/metabolismo
20.
Int J Mol Sci ; 24(7)2023 Apr 02.
Artigo em Inglês | MEDLINE | ID: mdl-37047622

RESUMO

The voltage-dependent anion channel (VDAC) is the primary regulating pathway of water-soluble metabolites and ions across the mitochondrial outer membrane [...].


Assuntos
Membranas Mitocondriais , Canais de Ânion Dependentes de Voltagem , Canais de Ânion Dependentes de Voltagem/metabolismo , Membranas Mitocondriais/metabolismo , Mitocôndrias/metabolismo , Íons/metabolismo
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA
...