Semiautonomous, self-reproducing organelles that occur in the cytoplasm of all cells of most, but not all, eukaryotes. Each mitochondrion is surrounded by a double limiting membrane. The inner membrane is highly invaginated, and its projections are called cristae. Mitochondria are the sites of the reactions of oxidative phosphorylation, which result in the formation of ATP. They contain distinctive RIBOSOMES, transfer RNAs (RNA, TRANSFER); AMINO ACYL T RNA SYNTHETASES; and elongation and termination factors. Mitochondria depend upon genes within the nucleus of the cells in which they reside for many essential messenger RNAs (RNA, MESSENGER). Mitochondria are believed to have arisen from aerobic bacteria that established a symbiotic relationship with primitive protoeukaryotes. (King & Stansfield, A Dictionary of Genetics, 4th ed)
Mitochondria in hepatocytes. As in all mitochondria, there are an outer membrane and an inner membrane, together creating two separate mitochondrial compartments: the internal matrix space and a much narrower intermembrane space. In the liver mitochondrion, an estimated 67% of the total mitochondrial proteins is located in the matrix. (From Alberts et al., Molecular Biology of the Cell, 2d ed, p343-4)
Glycosides from plants of the genus DIGITALIS. Some of these are useful as cardiotonic and anti-arrhythmia agents. Included also are semi-synthetic derivatives of the naturally occurring glycosides. The term has sometimes been used more broadly to include all CARDIAC GLYCOSIDES, but here is restricted to those related to Digitalis.
Cell Membrane Permeability
The two lipoprotein layers in the MITOCHONDRION. The outer membrane encloses the entire mitochondrion and contains channels with TRANSPORT PROTEINS to move molecules and ions in and out of the organelle. The inner membrane folds into cristae and contains many ENZYMES important to cell METABOLISM and energy production (MITOCHONDRIAL ATP SYNTHASE).
Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone
Components of a cell produced by various separation techniques which, though they disrupt the delicate anatomy of a cell, preserve the structure and physiology of its functioning constituents for biochemical and ultrastructural analysis. (From Alberts et al., Molecular Biology of the Cell, 2d ed, p163)
Microscopy using an electron beam, instead of light, to visualize the sample, thereby allowing much greater magnification. The interactions of ELECTRONS with specimens are used to provide information about the fine structure of that specimen. In TRANSMISSION ELECTRON MICROSCOPY the reactions of the electrons that are transmitted through the specimen are imaged. In SCANNING ELECTRON MICROSCOPY an electron beam falls at a non-normal angle on the specimen and the image is derived from the reactions occurring above the plane of the specimen.
A basic element found in nearly all organized tissues. It is a member of the alkaline earth family of metals with the atomic symbol Ca, atomic number 20, and atomic weight 40. Calcium is the most abundant mineral in the body and combines with phosphorus to form calcium phosphate in the bones and teeth. It is essential for the normal functioning of nerves and muscles and plays a role in blood coagulation (as factor IV) and in many enzymatic processes.
Membrane Potential, Mitochondrial
Mitochondrial Membrane Transport Proteins
Electron Transport Complex IV
A multisubunit enzyme complex containing CYTOCHROME A GROUP; CYTOCHROME A3; two copper atoms; and 13 different protein subunits. It is the terminal oxidase complex of the RESPIRATORY CHAIN and collects electrons that are transferred from the reduced CYTOCHROME C GROUP and donates them to molecular OXYGEN, which is then reduced to water. The redox reaction is simultaneously coupled to the transport of PROTONS across the inner mitochondrial membrane.
A closely related group of toxic substances elaborated by various strains of Streptomyces. They are 26-membered macrolides with lactone moieties and double bonds and inhibit various ATPases, causing uncoupling of phosphorylation from mitochondrial respiration. Used as tools in cytochemistry. Some specific oligomycins are RUTAMYCIN, peliomycin, and botrycidin (formerly venturicidin X).
A chemical reaction in which an electron is transferred from one molecule to another. The electron-donating molecule is the reducing agent or reductant; the electron-accepting molecule is the oxidizing agent or oxidant. Reducing and oxidizing agents function as conjugate reductant-oxidant pairs or redox pairs (Lehninger, Principles of Biochemistry, 1982, p471).
Rats, Inbred Strains
Centrifugation, Density Gradient
Reactive Oxygen Species
Molecules or ions formed by the incomplete one-electron reduction of oxygen. These reactive oxygen intermediates include SINGLET OXYGEN; SUPEROXIDES; PEROXIDES; HYDROXYL RADICAL; and HYPOCHLOROUS ACID. They contribute to the microbicidal activity of PHAGOCYTES, regulation of signal transduction and gene expression, and the oxidative damage to NUCLEIC ACIDS; PROTEINS; and LIPIDS.
Cytochrome c Group
One of the mechanisms by which CELL DEATH occurs (compare with NECROSIS and AUTOPHAGOCYTOSIS). Apoptosis is the mechanism responsible for the physiological deletion of cells and appears to be intrinsically programmed. It is characterized by distinctive morphologic changes in the nucleus and cytoplasm, chromatin cleavage at regularly spaced sites, and the endonucleolytic cleavage of genomic DNA; (DNA FRAGMENTATION); at internucleosomal sites. This mode of cell death serves as a balance to mitosis in regulating the size of animal tissues and in mediating pathologic processes associated with tumor growth.
Molecular Sequence Data
Descriptions of specific amino acid, carbohydrate, or nucleotide sequences which have appeared in the published literature and/or are deposited in and maintained by databanks such as GENBANK, European Molecular Biology Laboratory (EMBL), National Biomedical Research Foundation (NBRF), or other sequence repositories.
The second enzyme in the committed pathway for CHOLESTEROL biosynthesis, this enzyme catalyzes the first oxygenation step in the biosynthesis of STEROLS and is thought to be a rate limiting enzyme in this pathway. Specifically, this enzyme catalyzes the conversion of SQUALENE to (S)-squalene-2,3-epoxide.
Mitochondrial ADP, ATP Translocases
A class of nucleotide translocases found abundantly in mitochondria that function as integral components of the inner mitochondrial membrane. They facilitate the exchange of ADP and ATP between the cytosol and the mitochondria, thereby linking the subcellular compartments of ATP production to those of ATP utilization.
The class of all enzymes catalyzing oxidoreduction reactions. The substrate that is oxidized is regarded as a hydrogen donor. The systematic name is based on donor:acceptor oxidoreductase. The recommended name will be dehydrogenase, wherever this is possible; as an alternative, reductase can be used. Oxidase is only used in cases where O2 is the acceptor. (Enzyme Nomenclature, 1992, p9)
Carbonyl Cyanide m-Chlorophenyl Hydrazone
Amino Acid Sequence
A system of cisternae in the CYTOPLASM of many cells. In places the endoplasmic reticulum is continuous with the plasma membrane (CELL MEMBRANE) or outer membrane of the nuclear envelope. If the outer surfaces of the endoplasmic reticulum membranes are coated with ribosomes, the endoplasmic reticulum is said to be rough-surfaced (ENDOPLASMIC RETICULUM, ROUGH); otherwise it is said to be smooth-surfaced (ENDOPLASMIC RETICULUM, SMOOTH). (King & Stansfield, A Dictionary of Genetics, 4th ed)
A coenzyme composed of ribosylnicotinamide 5'-diphosphate coupled to adenosine 5'-phosphate by pyrophosphate linkage. It is found widely in nature and is involved in numerous enzymatic reactions in which it serves as an electron carrier by being alternately oxidized (NAD+) and reduced (NADH). (Dorland, 27th ed)
bcl-2-Associated X Protein
Electron Transport Complex I
A flavoprotein and iron sulfur-containing oxidoreductase complex that catalyzes the conversion of UBIQUINONE to ubiquinol. In MITOCHONDRIA the complex also couples its reaction to the transport of PROTONS across the internal mitochondrial membrane. The NADH DEHYDROGENASE component of the complex can be isolated and is listed as EC 22.214.171.124.
Electrophoresis, Polyacrylamide Gel
A water-soluble, colorless crystal with an acid taste that is used as a chemical intermediate, in medicine, the manufacture of lacquers, and to make perfume esters. It is also used in foods as a sequestrant, buffer, and a neutralizing agent. (Hawley's Condensed Chemical Dictionary, 12th ed, p1099; McGraw-Hill Dictionary of Scientific and Technical Terms, 4th ed, p1851)
The voltage differences across a membrane. For cellular membranes they are computed by subtracting the voltage measured outside the membrane from the voltage measured inside the membrane. They result from differences of inside versus outside concentration of potassium, sodium, chloride, and other ions across cells' or ORGANELLES membranes. For excitable cells, the resting membrane potentials range between -30 and -100 millivolts. Physical, chemical, or electrical stimuli can make a membrane potential more negative (hyperpolarization), or less negative (depolarization).
Mitochondrial Proton-Translocating ATPases
Proto-Oncogene Proteins c-bcl-2
Membrane proteins encoded by the BCL-2 GENES and serving as potent inhibitors of cell death by APOPTOSIS. The proteins are found on mitochondrial, microsomal, and NUCLEAR MEMBRANE sites within many cell types. Overexpression of bcl-2 proteins, due to a translocation of the gene, is associated with follicular lymphoma.
A family of intracellular CYSTEINE ENDOPEPTIDASES that play a role in regulating INFLAMMATION and APOPTOSIS. They specifically cleave peptides at a CYSTEINE amino acid that follows an ASPARTIC ACID residue. Caspases are activated by proteolytic cleavage of a precursor form to yield large and small subunits that form the enzyme. Since the cleavage site within precursors matches the specificity of caspases, sequential activation of precursors by activated caspases can occur.
Indicators and Reagents
Substances used for the detection, identification, analysis, etc. of chemical, biological, or pathologic processes or conditions. Indicators are substances that change in physical appearance, e.g., color, at or approaching the endpoint of a chemical titration, e.g., on the passage between acidity and alkalinity. Reagents are substances used for the detection or determination of another substance by chemical or microscopical means, especially analysis. Types of reagents are precipitants, solvents, oxidizers, reducers, fluxes, and colorimetric reagents. (From Grant & Hackh's Chemical Dictionary, 5th ed, p301, p499)
Voltage-Dependent Anion Channels
Diseases caused by abnormal function of the MITOCHONDRIA. They may be caused by mutations, acquired or inherited, in mitochondrial DNA or in nuclear genes that code for mitochondrial components. They may also be the result of acquired mitochondria dysfunction due to adverse effects of drugs, infections, or other environmental causes.
Acidic phospholipids composed of two molecules of phosphatidic acid covalently linked to a molecule of glycerol. They occur primarily in mitochondrial inner membranes and in bacterial plasma membranes. They are the main antigenic components of the Wassermann-type antigen that is used in nontreponemal SYPHILIS SERODIAGNOSIS.
Electron Transport Complex III
A multisubunit enzyme complex that contains CYTOCHROME B GROUP; CYTOCHROME C1; and iron-sulfur centers. It catalyzes the oxidation of ubiquinol to UBIQUINONE, and transfers the electrons to CYTOCHROME C. In MITOCHONDRIA the redox reaction is coupled to the transport of PROTONS across the inner mitochondrial membrane.
The first committed enzyme of the biosynthesis pathway that leads to the production of STEROLS. it catalyzes the synthesis of SQUALENE from farnesyl pyrophosphate via the intermediate PRESQUALENE PYROPHOSPHATE. This enzyme is also a critical branch point enzyme in the biosynthesis of ISOPRENOIDS that is thought to regulate the flux of isoprene intermediates through the sterol pathway.
Digitonin as a Biological Detergent | AG Scientific Blog
Digitonin (C56H92O29) is a steroidal, monodesmosidic saponin (a toxic chemical with a plant-based origin) derived from the ... Mitochondria isolation requires selective detergents (i.e., digitonin) that target lipids not located within the cell, such as ... Test results of digitonin application showed improved yield and respiratory control ratio of isolated mitochondria compared to ... Researchers commonly apply this detergent in studies that involve membrane proteins and isolation of mitochondria. Digitonin ...
An N-terminal addressing sequence targets NLRX1 to the mitochondrial matrix | Journal of Cell Science
Digitonin treatment of isolated mitochondria. Isolated mitochondria were incubated for 15 minutes on ice with an increasing ... Moreover, digitonin treatment of purified mitochondria demonstrated a similar profile for endogenous NLRX1 (Fig. 2D) as the one ... B) Purified mitochondria from HEK293T cells overexpressing NLRX1-FLAG were treated with increasing concentrations of digitonin ... Mitochondria sub-fractionation. Isolated mitochondria were resuspended at a protein concentration of 4-8 mg/ml in a hypotonic ...
Increased Vulnerability of Brain Mitochondria in Diabetic (Goto-Kakizaki) Rats With Aging and Amyloid-β Exposure | Diabetes
24), with slight modifications, using 0.02% digitonin to free mitochondria from the synaptosomal fraction. In brief, a rat was ... A: Mitochondria isolated from 1.5-month-old GK rats. B: Mitochondria isolated from 12-month-old GK rats. C: Mitochondria ... Isolation of brain mitochondria.. Brain mitochondria were isolated from male Wistar and GK rats by the method of Rosenthal et ... Effect of Aβ25-35 on ΔΨm of mitochondria isolated from 24-month-old GK rats. Freshly isolated brain mitochondria (0.8 mg) in 1 ...
Distinct stages in stress granule assembly and disassembly. - PubMed - NCBI
Yeast were grown in presence of 1,6-Hexanediol and 10 μg/mL Digitonin for 10 min or just 10 μg/mL Digitonin prior to lysis. ... Mitochondria. Get1, Guided Entry of Tail-anchored proteins (YGL020C). Atp1, ATP synthase (YBL099W). Scale bar is 2 μm. (B) Time ... Yeast were grown in the presence of 10% 1,6-Hexanediol and 10 μg/mL Digitonin for 10 min or just 10 μg/mL Digitonin ± ... 6-Hexanediol and 10 μg/mL Digitonin for 2 min or 30 min; or just 10 μg/mL Digitonin for 2 min or 30 min. For Nsp1-GFP and Atp1- ...
Mitochondria Regulate the Ca2+-Exocytosis Relationship of Bovine Adrenal Chromaffin Cells | Journal of Neuroscience
1986) Guanine nucleotide effects on catecholamine secretion from digitonin-permeabilized adrenal chromaffin cells. J Biol Chem ... Mitochondria can sequester large amounts of calcium and function as a cytosolic Ca2+ buffer of low affinity and high capacity ( ... 1994) Mitochondria buffer physiological calcium loads in cultured rat dorsal root ganglion neurons. J Neurosci 14:348-356. ... Mitochondria Regulate the Ca2+-Exocytosis Relationship of Bovine Adrenal Chromaffin Cells. David R. Giovannucci, Michael D. ...
Accumulation of Amyloid Precursor Protein in the Mitochondrial Import Channels of Human Alzheimer's Disease Brain Is Associated...
Accumulation of APP in mitochondria of AD brains. Next, we investigated whether APP is targeted to mitochondria in human AD and ... The pellet was further treated with digitonin, as described previously (Brown et al., 2004), to remove microsomal and ... 4D) were markedly reduced in mitochondria from AD brains compared with that of NC mitochondria. Inhibition of cytochrome c ... It may be possible that Aβ is generated outside the mitochondria and is imported into the mitochondria. Furthermore, our ...
Motor recruitment to the TIM23 channel's lateral gate restricts polypeptide release into the inner membrane | Nature...
e WT mitochondria or mitochondria containing Pam18His or HisS*Tim23 were solubilized in digitonin buffer and subjected to Ni- ... f After import of [35S]-labeled b2(220)-DHFR without proteinase K treatment, mitochondria were solubilized in digitonin buffer ... d Western blot analysis of steady-state protein using isolated mitochondria (5, 10, and 20 µg). e Digitonin-solubilized ... Isolation of mitochondria. Mitochondria were isolated as previously described45. Yeast were grown in YPG medium at 30 °C and ...
Mitochondria pelleted together with nuclei from 293 Cells using Sucrose gradient - Molecular Biology
I found digitonin a very reliable detergent. Last time I even tried Tween 20 but I couldnt fractionate well. However, it ... Mitochondria pelleted together with nuclei from 293 Cells using Sucrose gradient - (Mar/27/2012 ). Hi there,. I am doing an ... I would use digitonin instead of Triton or NP40. Look for Afshin Samalis method for rapid isolation of cytoplasm with ... which was suspected to be mitochondria. Could this indicate that the sucrose gradient separation was carried out improperly?. I ...
Enriching Mitochondria - SENS Research Foundation
Rapid enrichment of mitochondria from mammalian cell cultures using digitonin ... Mitochondria purification involves cell lysis followed by separation of the organelles from the rest of the cellular components ... Rapid enrichment of mitochondria from mammalian cells using commonly available reagents.. *Multiple samples can be processed ... The MitoSENS team has recently published a new protocol which enables researchers to isolate mitochondria from mammalian cells ...
Search Results | Biochemical Journal | Portland Press
CY(-) and CY(+) mitochondria are sensitive and resistant respectively to rotenone. Growth of cybrids in rutamycin for 2 weeks ... treatment with digitonin and differential centrifugation. Similar results were obtained with each technique. δ-Aminolaevulate ... There was no evidence for an increase in the relative amount of cristae protein in mitochondria during this period to account ... The ATPase of TL mitochondria is similarly resistant to rutamycin whether grown in the presence or absence of antibiotic. The ...
Digitonin Used as non-ionic detergent | Digitin | Sigma-Aldrich
Digitonin; Mild nonionic detergent used to solubilize receptors and permeabilize cellular and nuclear membranes. Bulk and ... Digitonin has been used in a study to assess a procedure for isolating mitochondria from ascites tumor cells. It has also been ... What is the solubility of Product D141, Digitonin? The solubility of D141 is tested by heating a 5% solution in water to 95-98° ... How pure is Product D141, Digitonin? The purity of this product is evaluated by Thin Layer Chromatography. The actual results ...
NDUFA10 mutations cause complex I deficiency in a patient with Leigh disease | European Journal of Human Genetics
Blue native-gradient gels (5-15%) were cast and run with 40 μg of digitonin-isolated mitochondria as described previously.22, ... Janssen AJ, Trijbels FJ, Sengers RC et al: Measurement of the energy-generating capacity of human muscle mitochondria: ... Analysis of steady-state protein phosphorylation in mitochondria using a novel fluorescent phosphosensor dye. J Biol Chem 2003 ... and in-gel complex I activity measurements with mitochondria-enriched fractions of the cultured patient fibroblasts. Figure 2a ...
Frontiers | STARD1 Functions in Mitochondrial Cholesterol Metabolism and Nascent HDL Formation. Gene Expression and Molecular...
STARD1 expression also exhibits a novel, slow splicing that slows delivery of mRNA to mitochondria. Stimulation of ... 1 complex of 3.5 kb mRNA with peri-nuclear mitochondria. Adrenal cells are similar but distinguished from testis cells by ... STARD1 and STARD7 target their respective activities to mitochondria, via N-terminal domains (NTD) of over 50 amino acids. The ... STARD1 and STARD7 target their respective activities to mitochondria, via N-terminal domains (NTD) of over 50 amino acids. The ...
Structural and Functional Association of Trypanosoma brucei MIX Protein with Cytochrome c Oxidase Complex | Eukaryotic Cell
ATP production was measured in digitonin-extracted mitochondria. The three ATP-producing pathways were triggered by the ... Digitonin fractionation.The crude mt fraction isolated by hypotonic lysis (10) was treated with 320 μM digitonin followed by ... ATP production in mitochondria isolated from noninduced cells and tested without any additions (None) was set to 100%. All ... An RNA-binding respiratory component mediates import of type II tRNAs into Leishmania mitochondria. J. Biol. Chem.281:25270- ...
Distinct Forms of Mitochondrial TOM-TIM Supercomplexes Define Signal-Dependent States of Preprotein Sorting | Molecular and...
After reisolation, mitochondria were washed and lysed in ice-cold digitonin buffer (1% [wt/vol] digitonin, 20 mM Tris-HCl [pH ... and the mitochondria were lysed with digitonin and subjected to affinity purification. In wild-type and tim17-4 mitochondria, ... The resulting mitochondria show a reduced efficiency for import of PAM-dependent proteins (4, 33). We subjected the digitonin- ... Tim21ProtA mitochondria were solubilized in digitonin buffer, and Tim21-containing TIM23 complexes were purified by IgG ...
Unraveling the Intricate Organization of Mammalian Mitochondrial Presequence Translocases: Existence of Multiple Translocases...
A) HEK293T mitochondria depleted for Magmas (Mag), JC19, or JC15 were digitonin lysed and subjected to CoIP using the indicated ... Mitochondria were isolated using a cell mitochondrion isolation kit (Sigma). Equivalent amounts of purified mitochondria were ... A) HEK293T mitochondria were solubilized in 1% digitonin buffer and analyzed by glycerol gradient centrifugation (20 to 50%). ... Glycerol gradient centrifugation.A total of 2 mg of yeast or human mitochondria was lysed in 1% digitonin buffer (25 mM Tris [ ...
Cells (6 × 105) grown in 35-mm dishes were treated with Dox and/or PFT-α for 16 h. Mitochondria were isolated by the digitonin ... H9c2 cells were treated with 5 μM of Dox and/or 20 μM PFT-α for 16 h. Digitonin was used to permeabilize plasma membrane. ... Digitonin is a weak nonionic detergent. At low concentrations, it can permeabilize plasma membrane and release cytosolic ... Robertson JD, Enoksson M, Suomela M, Zhivotovsky B, and Orrenius S. Caspase-2 acts upstream of mitochondria to promote ...
Independent evolution of functionally exchangeable mitochondrial outer membrane import complexes | eLife
Mitochondria or mitochondria-enriched fractions were solubilised with buffer (1% digitonin or 0.2% TritonX-100, 20 mM Tris, 0.1 ... For mitochondria enriched fractions by digitonin extraction of T. brucei, the cells were incubated for 10 min on ice in 20 mM ... D) The mitochondria described in (A) were solubilised in a buffer containing 1% digitonin and then analysed by BN-PAGE followed ... B) Top panels: immunoblot analysis of whole cells (Tot), soluble (Cyt) and digitonin-extracted mitochondria-enriched pellet ( ...
Differential effects of l -buthionine sulfoximine and ethacrynic acid on glutathione levels and mitochondrial function in PC12...
The conserved GTPase Gem1 regulates endoplasmic reticulum-mitochondria connections | PNAS
We purified the ERMES complex from digitonin-solubilized whole-cell extracts of strains bearing functional, C-terminally tandem ... Altered ER-mitochondria contact impacts mitochondria calcium homeostasis and contributes to neurodegeneration in vivo in ... Mitochondria were visualized by MitoTracker staining (Fig. 4A, Upper Left) or an α-Tomm-20 antibody (Fig. S4F), and the ER ... 4A, Lower Left). Miro-1 staining was observed in a few foci per mitochondria, strongly reminiscent of ERMES foci (Fig. 4A, ...
Ups1p and Ups2p antagonistically regulate cardiolipin metabolism in mitochondria | JCB
500 µg of mitochondria was solubilized at 2 mg protein/ml in digitonin buffer (1% digitonin, 20 mM Tris-HCl, pH 7.5, 50 mM NaCl ... 150 µg of mitochondria was solubilized in 40 µl 1.0% digitonin buffer (1.0% digitonin, 20 mM Tris-HCl, pH 7.5, 50 mM NaCl, and ... Mitochondria were solubilized at 2 mg protein/ml in digitonin buffer for 20 min on ice and then centrifuged at 16,100 g for 15 ... The mitochondria were lysed with digitonin at the indicated time points and analyzed by BN-PAGE. As a negative control, 10 µg/ ...
ER-mitochondria cross-talk is regulated by the Ca2+ sensor NCS1 and is impaired in Wolfram syndrome | Science Signaling
After a 2-min washing (to remove digitonin), [Ca2+]m uptake was estimated by mt-AEQ after cell perfusion with the same ... Bar graph shows the percentage of mitochondria in contact with ER (means ± SEM, n = 403 mitochondria in control cells and n = ... Bar graph shows the relative percentage of mitochondria in contact with ER (n = 257 mitochondria for siScr and n = 239 ... Mitochondria interact physically and functionally with endoplasmic reticulum (ER) through mitochondria-associated membranes ( ...
Assessment of mitochondrial functions and cell viability in renal cells overexpressing protein kinase C isozymes. - PubMed -...
Respiration is assessed in digitonin-permeabilized RPTC as state 3 (maximum respiration in the presence of excess substrates ... a fluorescent dye that specifically accumulates in mitochondria, and live monolayers are examined under a fluorescent ... respiratory chain is assessed by measuring activities of all four complexes of the respiratory chain in isolated mitochondria. ...
F, effect of IB05204 in cytochrome c release from mitochondria. Cells were permeabilized with 10 μg digitonin, and cytosolic ... 5F, cytochrome c was found to be effectively released from the mitochondria into the cytosol after treating BAEC with 5 μmol/L ... A precursor to caspase activation is the release of cytochrome c, as the mitochondria become leakier after the activation of ... Apoptosis can be induced by two pathways: the extrinsic death receptor cascade or the intrinsic mitochondria cascade. In both ...
Acetylcholine Has Dose-Dependent Bimodal Effects on Coronary Blood Flow in Man | Clinical Science | Portland Press
Measurement of mitochondrial respiration in permeabilized murine neuroblastoma (N-2alpha) cells, a simple and rapid in situ...
Most mitochondria-based methods used to investigate toxins require the use of relatively large amounts of material and hence ... Most mitochondria-based methods used to investigate toxins require the use of relatively large amounts of material and hence ... Eukaryotic cell membranes are selectively permeabilized with digitonin to render a system in which mitochondrial respiration ... Most mitochondria-based methods used to investigate toxins require the use of relatively large amounts of material and hence ...
Somatic mtDNA mutations cause aging phenotypes without affecting reactive oxygen species production | PNAS
... before and after digitonin permeabilization. Cells were permeabilized by using 1.1 μl of 5 mg/ml digitonin per 1 × 106 cells. ... Aconitase Activity. Isolated mitochondria were diluted in a solution consisting of 0.1% Triton X-100, 50 mM KH2PO4, and 1 mM ... Robinson J. B., Sumegi, B. L. & Srere P. A. (1987) in Mitochondria: A Practical Approach, eds. Darley-Usmar, V. M., Rickwood, D ... We found normal aconitase activities in heart mitochondria from 12-, 25-, and 40-week-old mtDNA mutator mice (Fig. 3B ). Hence ...
NCX3 regulates mitochondrial Ca2+ handling through the AKAP121-anchored signaling complex and prevents hypoxia-induced neuronal...
1B). Similar results were obtained by treating mitochondria with digitonin (500 µM) (Fig. 1B). Interestingly, confocal ... Extraction of brain mitochondria by the Percoll gradient method. The Percoll gradient method was used to purify mitochondria ... 2002). Heterogeneous distribution of pyruvate dehydrogenase in the matrix of mitochondria. Mitochondrion 1, 327-338. doi: ... or treated with digitonin (500 µM, 10 or 20 minutes). (C) Distribution of NCX3 in mitochondria (arrows), PM and ER and ...
Arabidopsis PPR40 Connects Abiotic Stress Responses to Mitochondrial Electron Transport | Plant Physiology
A, Purified mitochondria were solubilized by digitonin, and protein complexes were separated by Suc gradient centrifugation. ... Preparation of Mitochondria and Protein Isolation and Detection. Intact mitochondria were isolated from either 1-week-old cell ... Giegé P, Brennicke A (1999) RNA editing in Arabidopsis mitochondria effects 441 C to U changes ORFs. Proc Natl Acad Sci USA 96: ... In wild-type mitochondria, electrons flow through either Complexes I-III-IV or Complexes II-III-IV consuming oxygen and ...
MembranesIntact mitochondriaOuterProteinsFunction of mitochondriaImport into mitochondriaApoptosisCellularCytoplasmBrain mitochondria isolatedOxidative phosphorylationIsolated rat liver mitochondriaProtein translocationPermeabilizationChloroplastsInner membrane of mitochondriaCristaeLocalizationEndoplasmicSelectivelyAccumulationOrganellesCytochrome c releaseLiverDetergentPresent in mitochondriaCytosol and mitochondriaIntegrity of mitochondriaTumorSubcellularBiogenesisIsolate mitochondriaReleased in the cytosolCellsComplexesMembrane potentialEnter mitochondriaPlant mitochondriaYeast mitochondriaIsolationPathwaysFractionsEukaryotic organismsCalciumElectronAssayRespirationNucleiPermeabilityRespiratory chainRegulateLipidDownstreamUltrastructure
- The experiment involved digitonin application with various natural and artificial membranes, including large unilamellar vesicles (LUVs) and giant unilamellar vesicles (GUVs). (agscientific.com)
- The permeabilization of cell membranes usually includes two critical reagents: organic solvents and a biological detergent such as digitonin. (agscientific.com)
- The process involves a two-step approach that expresses fluorescent-protein fusion proteins and permeabilization of plasma membranes, which require digitonin preparations. (agscientific.com)
- Protein transport machineries of the endoplasmic reticulum, chloroplasts, and mitochondria facilitate anterograde protein translocation across membranes as well as lateral protein transport of precursors with hydrophobic transmembrane regions into the lipid phase. (nature.com)
- Eukaryotic cell membranes are selectively permeabilized with digitonin to render a system in which mitochondrial respiration can be measured rapidly and with considerable sensitivity. (worldcat.org)
- Mitochondria are made up by two membranes: the outer membrane (OM) and the inner membrane (IM). (rupress.org)
- Estimates of the degree of damage to the outer membrane reveal 3-5% breakage in the M1 mitochondria and 11-17% in PM mitochondria, 1 Optimum procedures for subfractionation of mitochondria into inner and outer membranes, and a soluble fraction are developed employing sonication and sucrose density gradient analysis. (gla.ac.uk)
- Digitonin based selective permeabilization of cell membranes further illustrated that DHFR-TS is not cytosolic but localized and released alongside known mitochondrial protein. (washington.edu)
- Evidence for the presence of the dimeric ATP synthase complexes came initially from molecular sizing analysis performed on complexes, which were solubilized from mitochondrial membranes under mild detergent lysis conditions (e.g., by use of digitonin) ( 3 , 4 ). (asm.org)
- The distribution of the newly inserted PC between the outer and the inner membrane was assessed by separation of the two membranes using digitonin treatment. (epfl.ch)
- Digitonin permeabilizes plasma membranes but leaves lysosomal membranes intact, allowing for a quantifiable measurement of lysosomal hydrolases and comparison to normal cellular enzyme activity. (goldbio.com)
- Digitonin at lower concentrations (approximately 25 μg/ml), allows the plasma membrane to be solubilized with very little effect on intracellular membranes because of its lower cholesterol content. (goldbio.com)
- Using Digitonin at this concentration for the extraction of cytosolic proteins does not dramatically impact these organelle membranes. (goldbio.com)
- The genome of Saccharomyces cerevisiae contains 35 members of a family of transport proteins that, with a single exception, are found in the inner membranes of mitochondria. (embopress.org)
- The rationale for using digitonin to improve mitochondria recovery is that it selectively disrupts the lipid membranes enriched in sterols. (oxphos.org)
- As the molecular weight of digitonin is about three times that of cholesterol, the intercalation of digitonin into cell membranes significantly disrupts the packing of lipids in the bilayer and facilitates its rupture. (oxphos.org)
- Therefore, digitonin greatly facilitates the disruption of cholesterol-rich cellular membranes and synaptosomal membranes and dissolves fragments of endoplasmic reticulum and cellular plasmalemma. (oxphos.org)
- 23) How many membranes enclose a mitochondrion? (blogspot.com)
- Mitochondria are surrounded by two distinct membranes: the outer and the inner membrane. (chemweb.com)
- Imported mTim44 was protected from protease digestion in which outer membranes were selectively permeabilized with digitonin. (northwestern.edu)
- Digitonin solubilizes lipids, especially in membranes and is used as a tool in cellular biochemistry, and reagent for precipitating cholesterol. (uchicago.edu)
- Scatchard analysis and drug displacement studies of the binding of [3H] PK11195 to intact mitochondria and to the outer membrane-enriched digitonin extract further confirm the localization of the peripheral-type benzodiazepine receptor to the mitochondrial outer membrane. (nih.gov)
- In intact mitochondria, there were approximately the same number of binding sites for [3H]PK11195 as for [3H]Ro5-4864, and their apparent Kd values were identical. (aspetjournals.org)
- When a study requires functional structurally intact mitochondria, digitonin based mitochondria isolation protocols such as described in [1, 7, 8] should better be avoided. (oxphos.org)
- We demonstrate that intact mitochondria are important to prevent cisplatin-induced apoptosis in MDCK-C7 cells and that acidic conditions can aggravate the toxic effects of cisplatin. (labome.org)
- Mitochondria are composed of an outer and an inner membrane (MOM and MIM, respectively), and the large majority of mitochondrial proteins are encoded in the nucleus, synthesized in the cytosol and transported to specific mitochondrial compartments via mechanisms that have been extensively studied ( Neupert and Herrmann, 2007 ). (biologists.org)
- Furthermore, in AD brains, mitochondrially associated APP formed stable ∼480 kDa complexes with the translocase of the outer mitochondrial membrane 40 (TOM40) import channel and a super complex of ∼620 kDa with both mitochondrial TOM40 and the translocase of the inner mitochondrial membrane 23 (TIM23) import channel TIM23 in an "N in mitochondria -C out cytoplasm " orientation. (jneurosci.org)
- The mechanism of membrane insertion and assembly of b-barrel proteins is a central question of outer membrane biogenesis of mitochondria, chloroplasts, and Gram-negative bacteria. (sciencemag.org)
- Integrity of the outer membrane in isolated mitochondria is monitored by an assay using exogeneous cytochrome c as substrate for cytochrome c oxidase, an inner membrane protein complex. (gla.ac.uk)
- Titration of isolated mitochondria with digitonin results in the simultaneous release of the peripheral-type benzodiazepine receptor and of monoamine oxidase, but not cytochrome oxidase, indicating association of the peripheral-type benzodiazepine receptor with the mitochondrial outer membrane. (nih.gov)
- Specifically, permeabilization of the outer mitochondrial membrane (OMM) and the release of certain proteins (including cytochrome c ) from the intermembrane space of mitochondria are regarded as key events in apoptosis induction. (rsc.org)
- In order to study the individual steps involved in the import of phosphatidylcholine (PC) into rat liver mitochondria, a number of PC analogues were introduced into the outer membrane of isolated mitochondria. (epfl.ch)
- Rat kidney mitochondria were fractionated by decompression and digitonin-based methods into outer and inner membrane-containing fractions before and after incorporation of the MBR-specific photoaffinity ligand [3H]PK14105. (aspetjournals.org)
- Confocal microscopy of mitochondria devoid of the outer membrane and Western blotting on fractionated mitochondria showed that Cx43 is located at the inner mitochondrial membrane, and coimmunoprecipitation of Cx43 with Tom20 ( T ranslocase of the o uter m embrane 20 ) and with heat shock protein 90 (Hsp90) indicated that it interacts with the regular mitochondrial protein import machinery. (ahajournals.org)
- When fixed cells are permeabilized with 0.005% digitonin, the antibody to cyctochrome c (red), a mitochondrial matrix marker, is unable to reach and bind cytochrome c, but the antibody to Tom20 (white), a mitochondrial outer membrane marker, is now able to bind Tom20. (cellimagelibrary.org)
- A partial rupture of the outer mitochondrial membrane induced by digitonin during the isolation of mitochondria is difficult to detect. (oxphos.org)
- Many critically important proteins such as cytochrome c, creatine kinase, sulfite oxidase, proteases and several pro-apoptotic proteins are located in the intermembrane space and could become depleted during the isolation of mitochondria if their outer membrane is ruptured. (oxphos.org)
- 3]. Dorbani L, Jancsik V, Linden M, Leterrier JF, Nelson BD, Rendon A. Subfractionation of the outer membrane of rat brain mitochondria: evidence for the existence of a domain containing the porin-hexokinase complex. (oxphos.org)
- TXNI is a cytosolic protein while TXNII due to a C-terminal hydrophobic tail is anchored in the outer membrane of the mitochondrion, endoplasmic reticulum and glycosomes. (bvsalud.org)
- Digitonin functions as a nonionic detergent with an uncharged hydrophilic head group and is often used to break down chemical interactions involving proteins and lipids. (agscientific.com)
- Researchers commonly apply this detergent in studies that involve membrane proteins and isolation of mitochondria. (agscientific.com)
- Thus, mitochondria would play a critical role in the control of secretory activity in chromaffin cells that undergo tonic or repetitive depolarizing activity, likely by limiting the Ca 2+ -dependent activation of specific proteins that recruit or prime secretory granules for exocytosis. (jneurosci.org)
- In the current study, the hypothesis that mitochondria regulate secretory activity by limiting rises in [Ca 2+ ] c and the subsequent activation of specific proteins that recruit or prime secretory granules for exocytosis was tested by monitoring stimulus-evoked changes in [Ca 2+ ] c and the secretory activity of single bovine chromaffin cells after selective pharmacological inhibition of mitochondrial Ca 2+ transport. (jneurosci.org)
- ii ) ROS production induces damage to lipids, proteins, and nucleic acids in mitochondria. (pnas.org)
- We show here that M11L is antiapoptotic when expressed independently of other viral proteins, and is directed specifically to mitochondria by a short COOH-terminal region that is necessary and sufficient for targeting. (rupress.org)
- Recent experimental findings have indicated a pivotal role for mitochondria in the "decision to die" checkpoint regulated by Bcl-2 proteins ( 10 )( 11 )( 12 )( 13 ). (rupress.org)
- In this context, Bcl-2 proteins are believed to regulate the mitochondrial permeability transition (PT) pore and thereby control the release of cytochrome c and other proteins from within mitochondria into the cytoplasm. (rupress.org)
- Other proteins released from the intermembrane space of mitochondria are the Apoptosis Inducing Factor (AIF), Smac/Diablo and Omi. (rsc.org)
- Targeting of these proteins to the mitochondrion was verified by a green fluorescence protein (GFP) reporter system, and by subcellular pre-fractionation using digitonin followed by western blotting. (gla.ac.uk)
- The β‐barrel proteins of mitochondria are synthesized on cytosolic ribosomes. (embopress.org)
- So far, three proteins have been identified as being responsible for the transport of cofactors into mitochondria. (embopress.org)
- 46) Where are electron transport proteins located in mitochondria? (blogspot.com)
- Furthermore, a great part of the matrix adenine nucleotides is bound to proteins , a notion supported by the fact that rat liver mitochondria retain more than 50% of their total adenine nucleotide content after permeabilization by toluene . (oxphos.org)
- While the majority of mitochondrial proteins are encoded in nuclear DNA (nuDNA), mitochondria contain their own genome, the circular dsDNA in the matrix mitochondrial DNA (mtDNA). (embopress.org)
Function of mitochondria3
- This assay is derived from a classic experiment to assess mitochondria and serves as a framework with which to build more complex experiments aimed at understanding both physiologic and pathophysiologic function of mitochondria and to predict the ability of cells to respond to stress and/or insults. (jove.com)
- 1. A specific inhibitor of mitochondrial protein translocation, which inhibitor modulates the assembly or function of mitochondria, wherein the inhibitor specifically targets the protein translocation pathway thereby modulating the assembly and function of the mitochondrion with respect to protein translocation and import. (freepatentsonline.com)
- 9. A method, comprising modulate the assembly or function of mitochondria by applying to a body of mitochondria or a cell a specific inhibitor of mitochondria protein translocation, wherein the inhibitor specifically targets the protein translocation pathway thereby modulating the assembly and function of the mitochondrion with respect to protein translocation and import. (freepatentsonline.com)
Import into mitochondria3
- In adrenal cells, STARD1 activates translation-coupled NTD-directed cholesterol import into mitochondria for metabolism by CYP11A1. (frontiersin.org)
- A decrease in calcium import into mitochondria increases cytosolic calcium concentration, which leads to transcriptional upregulation of over 100 genes including those involved in glucose metabolism, apoptosis and tumorigenesis (Kotiadis et al. (springer.com)
- Paschen S, Neupert W. Protein import into mitochondria. (labome.org)
- M11L blocks staurosporine-induced apoptosis by preventing mitochondria from undergoing a permeability transition, and the mitochondrial localization of this protein is essential for this function. (rupress.org)
- Investigation of mitochondrial function represents an important parameter of cardiac physiology as mitochondria are involved in energy metabolism, oxidative stress, apoptosis, aging, mitochondrial encephalomyopathies and drug toxicity. (jove.com)
- Selective targeting of survivin to mitochondria enhances colony formation in soft agar, accelerates tumor growth in immunocompromised animals, and abolishes tumor cell apoptosis in vivo. (jci.org)
- Schwerdt G, Freudinger R, Schuster C, Weber F, Thews O, Gekle M. Cisplatin-induced apoptosis is enhanced by hypoxia and by inhibition of mitochondria in renal collecting duct cells. (labome.org)
- The present study expands the contemporary view of mitochondria as important participants in cellular Ca 2+ dynamics and provides evidence that mitochondria regulate the supply of release-competent secretory granules. (jneurosci.org)
- Disruption of cellular Ca 2+ homeostasis by poisoning mitochondria enhanced the secretory responsiveness of chromaffin cells by increasing the amplitude of the transient rise and the time course of recovery to baseline of the evoked Δ[Ca 2+ ] c . (jneurosci.org)
- Mitochondria purification involves cell lysis followed by separation of the organelles from the rest of the cellular components. (sens.org)
- Mitochondria are indispensable organelles implicated in multiple aspects of cellular processes, including tumorigenesis. (asm.org)
- Normal cellular function requires homeostatic counterbalance of various metabolic pathways, with mitochondria playing a central role in the complex processes. (asm.org)
- Mitochondria are in the center of regulation of cellular energy homeostasis and redox balance, and integrate numerous metabolic pathways that are important in adaptive responses to extreme environmental conditions. (plantphysiol.org)
- Mitochondria are the critical cellular organelles responsible for energy generation and cellular homeostasis. (nih.gov)
- Mitochondria are vital in providing cellular energy via their oxidative phosphorylation system, which requires the coordinated expression of genes encoded by both the nuclear and mitochondrial genomes (mtDNA). (sciencemag.org)
- How mitochondria regulate a cellular process through retrograde signaling specifically remains relatively unexplored. (springer.com)
- Mitochondria are double-membrane enclosed eukaryotic organelles with a central role in numerous cellular functions. (chemweb.com)
- A second pathway "intrinsic" apoptotic pathway is initiated at the mitochondria (8) by cellular stresses, including antineoplastic drugs and radiation. (aacrjournals.org)
- Read this term paper to learn about the role of mitochondria in cellular oxidation. (biologydiscussion.com)
- Mitochondria produce ATP through oxidative phosphorylation (OXPHOS) and are involved in various biosynthesis and cellular signaling processes [ 1 ]. (embopress.org)
- Look for Afshin Samali's method for rapid isolation of cytoplasm with digitonin on google. (protocol-online.org)
- Subcellular fractionation was performed to evaluate expression of Bax and cytochrome c in the cytosol and mitochondria, and also translocation of cytochrome c from the cytoplasm to the nucleus. (biomedcentral.com)
- The various enzymes, reactants, and products of glycolysis are in solution in the cytoplasm outside mitochondria and apparently interact through random collisions. (biologydiscussion.com)
Brain mitochondria isolated2
- This study evaluated the respiratory indexes (respiratory control ratio [RCR] and ADP/O ratio), mitochondrial transmembrane potential (ΔΨm), repolarization lag phase, repolarization level, ATP/ADP ratio, and induction of the permeability transition pore of brain mitochondria isolated from normal Wistar and GK diabetic rats of different ages (1.5, 12, and 24 months of age). (diabetesjournals.org)
- It had been reported that digitonin-isolated guinea pig brain mitochondria exhibited non-standard patterns of response to the respiratory chain inhibitors  and that rat brain mitochondria isolated with digitonin exhibited altered cytochrome c release and Ca2 + -handling properties . (oxphos.org)
- 11) Who discovered mitochondria as the sites of oxidative phosphorylation in eukaryotes? (blogspot.com)
Isolated rat liver mitochondria1
- In isolated rat liver mitochondria, reports indicate a wide range of intramatrix free ATP to ADP ratios in State 3, ranging from 0.01 to 4.5 [2-4;8;9;12;22;24;27-30] or even to 8-12 [31;32]. (oxphos.org)
- The present invention provides compounds that modulate protein translocation in mitochondria, compositions thereof, and methods of identifying, making and using these. (freepatentsonline.com)
- The study corroborated the crucial role of digitonin concentration in human neutrophils permeabilization, alongside other factors such as permeabilizing period and temperature. (agscientific.com)
- In another experiment, scientists examined digitonin in the permeabilization of the plasma membrane in Trypanosoma cruzi , a parasite that grows in the guts of insects. (agscientific.com)
- The permeabilization of Trypanosoma brucei procyclic and bloodstream trypomastigotes with digitonin permitted the quantitative estimation of a mitochondrial membrane potential of the order of 130-140 mV, in both forms, using safranine O. Dependence on substrate oxidation and response of the procyclic mitochondrial membrane potential to phosphate, FCCP, valinomycin, and Ca2+ indicate that these mitochondria behave similarly to vertebrate mitochondria regarding the properties of their electrochemical proton gradient. (nih.gov)
- Following the removal of contaminating mitochondria from isolated bean etioplasts and greening chloroplasts, all photochemical activities were vanishingly low. (royalsocietypublishing.org)
- Our next strategy involved the non-ionic detergent, digitonin, to fragment spinach chloroplasts derived from hydroponically grown plants from which functional chloroplasts had been obtained overseas. (royalsocietypublishing.org)
Inner membrane of mitochondria3
- In many human mitochondrial diseases, the inner membrane of mitochondria folds incorrectly, in similar ways to the misfolding seen in the yeast cells that did not contain the MICOS complex. (elifesciences.org)
- Recent experiments, however, identify large assemblies of two homologous prohibitin subunits, PHB1 and PHB2, in the inner membrane of mitochondria as the physiologically active structure. (chemweb.com)
- The ring-like assembly of prohibitins and their sequence similarity with lipid raft-associated SPFH-family members suggests a scaffolding function of prohibitins, which may lead to functional compartmentalization in the inner membrane of mitochondria. (chemweb.com)
- In contrast, in the procyclic form (PF) vector stage of the parasite, the mitochondrion has many cristae, a complete cytochrome-mediated respiratory chain, and Krebs cycle enzymes ( 3 ). (asm.org)
- Mitochondria are larger and fewer in older individuals, and mitochondrial abnormalities such as vacuoles, abnormal cristae, and paracrystalline inclusions are more often found ( 4 ). (pnas.org)
- Despite the apparent importance of the structural organization of mitochondria, the components responsible for the morphology and biogenesis of cristae, and in particular of CJs, are largely unknown. (rupress.org)
- Mitochondrial prohibitin complexes control cell proliferation, cristae morphogenesis and the functional integrity of mitochondria. (chemweb.com)
- By immunoelectron microscopy, mTim44 was seen in the paracrystalline structures within the mitochondria, as well as in their cristae. (northwestern.edu)
- The mitochondrial Na + /Ca 2+ exchanger, which was first postulated in 1974, has been primarily investigated only from a functional point of view, and its identity and localization in the mitochondria have been a matter of debate over the past three decades. (biologists.org)
- Comparisons between DHFR-TS and mitochondria-marking dyes, an engineered mitochondrial protein (HProtein-GFP), and native DHODH enzyme all indicate mitochondrial localization of this important drug target, DHFR-TS. (washington.edu)
- Here we investigated the intramitochondrial localization of Cx43, the mechanism of Cx43 translocation to mitochondria and the effect of inhibiting translocation on the protection of preconditioning. (ahajournals.org)
- By using anti-FLAG epitope M2 monoclonal antibody, mTim44 was found to have its subcellular localization associated with mitochondria. (northwestern.edu)
- Cx43 expression and localization, analyzed by Western blot and confirmed by immunofluorescence analysis, showed that DZX co-treatement increases Cx43 amount both on sarcoplasmic membrane and on mitochondria. (mdpi.com)
- Mitochondria are connected to the endoplasmic reticulum (ER) through specialized protein complexes. (pnas.org)
- The endoplasmic reticulum (ER) and mitochondria have recently emerged as a paradigm for interorganelle communication ( 1 - 3 ). (pnas.org)
- It has also been shown not to significantly affect the structure of Ca 2+ -sequestering organelles, like the endoplasmic reticulum and mitochondria. (goldbio.com)
- As evident from the experiment, digitonin functioned as a cholesterol-binding agent to selectively permeabilize plasma through the high cholesterol level in its lipid bilayer. (agscientific.com)
- Lipid accumulation in skeletal muscle ( 7 ) or metabolic overload in mitochondria ( 8 ) have also been shown to occur with IR, and it has been shown that incomplete β-oxidation contributes to skeletal muscle IR ( 9 ). (diabetesjournals.org)
- The main site for ROS production is the respiratory chain inside the mitochondria and accumulation of mtDNA mutations, and impaired respiratory chain function have been associated with degenerative diseases and aging. (pnas.org)
- Mitochondria are a direct site of A beta accumulation in Alzheimer's disease neurons: implications for free radical generation and oxidative damage in disease progression. (semanticscholar.org)
- The accumulation of mitochondria-targeted delocalized lipophilic cations such as Mito-CP ( Fig. 1 ) in tumor cells is controlled by several factors including the lipophilicity and the chain length of mitochondria-targeted drugs (MTD), the negative mitochondrial membrane potential, and the levels of p-glycoprotein-mediated MDR protein that actively pumps cationic drugs out of the tumor cells ( 24 ). (aacrjournals.org)
- H. von Grafenstein and E. Neumann, "ATP-stimulated accumulation of calcium by chromaffin granules and mitochondria from the adrenal medulla", Biochemical and Biophysical Research Communications , vol. 117, 1983, pp. 245-251. (uni-bielefeld.de)
- We recently identified the ER-mitochondria encounter structure (ERMES) tethering complex, which plays a role in phospholipid exchange between the two organelles. (pnas.org)
- Mitochondria are ubiquitous organelles and fulfill a multitude of crucial functions in eukaryotic organisms. (rupress.org)
- The organelles of these Digitonin-treated cells can be extracted via centrifuge. (goldbio.com)
- The respiratory organelles in the eukaryotic cells are called mitochondria. (blogspot.com)
- Mitochondria are essential organelles of the eukaryotic cells that are made by expansion and division of pre-existing mitochondria. (chemweb.com)
Cytochrome c release3
- Calcium-induced Cytochrome c release from rat brain mitochondria is altered by digitonin. (oxphos.org)
- The resultant truncated Bid translocates to mitochondria where it can trigger cytochrome c release. (aacrjournals.org)
- HNHA treatment caused cytochrome c release from mitochondria, which was mediated by increased Bax expression and caspase activation. (biomedcentral.com)
- The effect of intact diphtheria toxin and of its fragment A on protein synthesis in mouse liver mitoplasts (digitonin-treated mitochondria) was studied. (ias.ac.in)
- Treatment of rat liver mitochondria with low levels of digitonin (0.01mg/mg protein) releases approx. (gla.ac.uk)
- 2]. Cr mel G, Filliol D, Jancsik V, Rendon A. Cholesterol distribution in rat liver and brain mitochondria as determined by stopped-flow kinetics with filipin. (oxphos.org)
Present in mitochondria2
- We have previously shown that connexin 43 (Cx43) is present in mitochondria, that its genetic depletion abolishes the protection of ischemia- and diazoxide-induced preconditioning, and that it is involved in reactive oxygen species (ROS) formation in response to diazoxide. (ahajournals.org)
- 20) What kind of ribosomes are present in mitochondria? (blogspot.com)
Cytosol and mitochondria1
- The Par14 protein was initially detected within the nucleus, cytosol and mitochondria of human HEK 293 cells [ 11 ], but how this distribution was accomplished remained to be determined. (biomedcentral.com)
Integrity of mitochondria1
- Cell viability can be calculated from respirometric indices, based on the assumption that respiratory integrity of mitochondria is maintained in dce suspended in mitochondrial respiration medium MiR05 . (oroboros.at)
- Digitonin has been used in a study to assess a procedure for isolating mitochondria from ascites tumor cells. (sigmaaldrich.com)
- We studied mitochondria of digitonin-permeabilized Hela cells, since digitonin (60 ug/10 6 cells) increases plasma membrane Ca2+ permeability, in order to avoid problems such as low mitochondrial yield and the possibility of obtaining damaged or uncoupled mitochondria from tumor cells. (bireme.br)
- 7]. Moreadith RW, Fiskum G. Isolation of mitochondria from ascites tumor cells permeabilized with digitonin. (oxphos.org)
- The subcellular distribution of enzymes involved in thiamine metabolism shows that ThPP is synthesized in the cytosol and then imported into mitochondria. (embopress.org)
- The results of digitonin titration experiments showed that in control fibroblasts at least 70% of the catalase activity is present in subcellular particles distinct from mitochondria or lysosomes. (uu.nl)
- The presequence translocase or the TIM23 complex accounts for import of approximately 60% of the total mitochondrial proteome and hence is critical for mitochondria biogenesis ( 4 ). (asm.org)
- The MitoSENS team has recently published a new protocol which enables researchers to isolate mitochondria from mammalian cells more efficiently and scalably than any prior technique, using only readily available and economical reagents and equipment. (sens.org)
- In earlier studies, digitonin treatment had frequently been used to isolate mitochondria from cells and tissue. (oxphos.org)
Released in the cytosol1
- We identified a novel pool of survivin that localizes to mitochondria and is rapidly released in the cytosol in response to apoptotic stimuli. (jci.org)
- Digitonin also exists in a wide range of processes, such as hemolysis (the destruction of red blood cells commonly caused by Gram-positive bacteria). (agscientific.com)
- Researchers applied digitonin in permeabilizing human neutrophils , a component that makes up 40 to 70 percent of white blood cells. (agscientific.com)
- Using pharmacological probes to inhibit mitochondrial Ca 2+ import, the ability of mitochondria to modulate secretory activity in single, patch-clamped bovine chromaffin cells was examined by simultaneously monitoring rapid changes in membrane surface area (Δ C m ) and cytosolic Ca 2+ levels ([Ca 2+ ] c ). (jneurosci.org)
- Rapid enrichment of mitochondria from mammalian cells using commonly available reagents. (sens.org)
- Trypanosomatid cells contain only a single large mitochondrion per cell that contains uniquely structured mt DNA that is called kinetoplast DNA ( 15 ). (asm.org)
- and Mdm10, an integral β-barrel OMM protein ( Fig. S1 B ). In cells, these protein complexes are arranged in larger assemblies, visible microscopically as one to five discrete puncta per cell at the ER-mitochondria interface. (pnas.org)
- Mitochondrial morphology in live cells is visualized using MitoTracker Red 580, a fluorescent dye that specifically accumulates in mitochondria, and live monolayers are examined under a fluorescent microscope. (nih.gov)
- Structures called mitochondria provide energy that cells need to live and grow. (elifesciences.org)
- To do this, mitochondria convert energy stored within sugars and other carbon-rich compounds into the energy currency of cells, a molecule called adenosine triphosphate (called ATP for short). (elifesciences.org)
- Defective mitochondria can cause cells to starve and also cause severe human diseases. (elifesciences.org)
- The intracellular transport mechanisms involved in maintaining Ca2+ homeostasis in T. cruzi have been characterized by measuring Ca2+ transport in digitonin-permeabilized cells. (bireme.br)
- Addition of Ca2+ to digitonin-permeabilized Hela cells gave rise to a cycle of respiratory stimulation. (bireme.br)
- Ca2+ release occurred upon addition of carbonyl cyanide p-trifluoromethoxy-phenylhydrazone (FCCP) to digitonin-permeabilized Hela cells, under state conditions. (bireme.br)
- Digitonin has been shown to induce hemolysis in red blood cells. (goldbio.com)
- Biological responses of cells to visible and near IR (laser) radiation occur due to physical and/or chemical changes in photoacceptor molecules, components of respiratory chains (cyt a/a 3 in mitochondria). (spie.org)
- We have found that the transient overexpression of the Htt fragments increases Ca2+ transients in the mitochondria of cells stimulated with Ca2+-mobilizing agonists. (plos.org)
- Mitochondria in HeLa cells are identified by mito-YFP (green). (cellimagelibrary.org)
- Mito-YFP transfected HeLa cells were treated with DMSO for 3 hrs, fixed with 4% paraformaldehyde and permeabilized with 0.005% digitonin/PBS. (cellimagelibrary.org)
- Mitochondria in HeLa cells are identified by mito-YFP (green) following treatment with the mitochondrial depolarizing agent CCCP (carbonyl cyanide m-chlorophenyl hydrazone). (ucsd.edu)
- Mito-YFP transfected HeLa cells were treated with CCCP (10µM) for 3 hrs, fixed with 4% paraformaldehyde and permeabilized with 0.005% digitonin/PBS. (ucsd.edu)
- In confirmation of its identity, cells lacking the gene for this carrier had reduced levels of ThPP in their mitochondria, and decreased activity of acetolactate synthase, a ThPP‐requiring enzyme found in the organellar matrix. (embopress.org)
- The following protocol uses a calcium-sensitive, membrane-impermeable fluorophore to monitor extra-mitochondrial calcium levels in the presence of permeabilized mammalian cells harboring activated mitochondria. (bio-protocol.org)
- HEK-293 cells with activated mitochondria are suspended in a recording buffer along with a membrane-impermeable calcium-sensitive fluorophore. (bio-protocol.org)
- 4) Who discovered mitochondria in plant cells? (blogspot.com)
- The ultrastructure of mitochondria varies considerably between tissues, organisms, and the physiological state of cells. (chemweb.com)
- Calcium homeostasis in digitonin-permeabilized bovine chromaffin cells. (uni-bielefeld.de)
- Respiratory control ratios (i.e., uncoupled respiration at optimum carbonyl cyanide p-trifluoromethoxyphenylhydrazone concentration divided by oligomycin inhibited respiration measured in unpermeabilized cells) remained as high as 11, indicating well-coupled mitochondria and functional integrity of the inner mitochondrial membrane. (oroboros.at)
- if mitochondria of dce are intact, respiration of dce is identical to digitonin-permeabilized cells ( pce ). (oroboros.at)
- Cardiolipin is required for the integrity of several protein complexes in the IM, including the TIM23 translocase, a dynamic complex which mediates protein import into the mitochondria through interactions with the import motor presequence translocase-associated motor (PAM). (rupress.org)
- Integrity of the respiratory chain is assessed by measuring activities of all four complexes of the respiratory chain in isolated mitochondria. (nih.gov)
- In mitochondria, the F 1 F o -ATP synthase complex does not exist as a separate entity in the membrane but rather forms dimeric complexes. (asm.org)
- It is useful for creating Digitonin-cholesterol complexes which form when Digitonin disrupts the cell membrane and binds with cholesterol in a nearly 1:1 stoichiometric ratio. (goldbio.com)
- Digitonin complexes with most unesterified 3-b-hydroxysterols, including cholesterol, the principal free sterol present in vertebrate tissues. (oxphos.org)
- Digitonin was also shown to alter the mobility of the mitochondrial respiratory complexes and the rates of electron traansfer . (oxphos.org)
- Par17 was imported into mitochondria in a time and membrane potential-dependent manner, where it reached the mitochondrial matrix. (biomedcentral.com)
- In contrast, in bloodstream mitochondria, development of a membrane potential was independent of substrate oxidation and dependent on hydrolysis of ATP by the mitochondrial oligomycin-sensitive ATPase, as demonstrated by collapse of the membrane potential by oligomycin and its insensitivity to the respiratory chain-inhibitor antimycin A. Mitochondria of T. brucei bloodstream forms were also able to take up Ca2+ by an electrophoretic mechanism. (nih.gov)
- 5]. Tretter L, Mayer-Takacs D, Adam-Vizi V. The effect of bovine serum albumin on the membrane potential and reactive oxygen species generation in succinate-supported isolated brain mitochondria. (oxphos.org)
- Our model  predicts that the concentrations of ADP and ATP in the matrix of mitochondria at State III (mitochondrial membrane potential is about -145 mV) are equal to 8.7 mM and 3.3 mM, respectively. (oxphos.org)
- These membrane potential values are for isolated mitochondria. (oxphos.org)
- For in situ mitochondria, membrane potential is lower at about -135 mV (Note 1). (oxphos.org)
- Glycerol enters the glycolytic pathway in the first stage, and the fatty acids enter mitochondria for oxidation in the second stage. (biologydiscussion.com)
- 5) In which plant mitochondria were first discovered? (blogspot.com)
- Par14 and Par17 associated with isolated human, rat and yeast mitochondria at low salt concentrations, but only the Par17 mitochondrial association was resistant to higher salt concentrations. (biomedcentral.com)
- Import of transcription factor MTF1 into the yeast mitochondria takes place through an unusual pathway. (uchicago.edu)
- But from running the miniprep isolation on a gel, it seems that I am getting huge DNA band at about 12-16kbp length, which was suspected to be mitochondria. (protocol-online.org)
- Use of Digitonin in mitochondria isolation. (oxphos.org)
- One of the most efficient protocols for mitochondria isolation from small amounts of brain tissue (~50 mg of adult mouse brain) combines Percoll gradient and digitonin treatment of tissue homogenates . (oxphos.org)
- Although it appears that distinct pathways leading to cell death are triggered by different signals, they often merge at a common "regulator" of this multistep process - mitochondria. (rsc.org)
- They are targeted to the mitochondria and inserted into their target membrane via various pathways. (chemweb.com)
- We have investigated these questions in our study, which we regard as the first rigorous system study of single‐cell bioenergetics subsequent to cyt‐ c release and one that bridges single‐cell microscopy, in vitro analysis with ordinary differential equations (ODE) based modelling of bioenergetics pathways in the mitochondria and the cytosol. (embopress.org)
- In the absence of rotenone, S-OXPHOS capacity may be underestimated in some types of mitochondria, whereas it is overestimated if anaplerotic pathways from malate are active, e.g. mt-malic enzyme would additionally activate the N-pathway. (oroboros.at)
- With the arrival of a Cary model 14R spectrophotometer and a Plant Industry workshop copy of the Bonner type cuvette assembly, I determined the cytochrome difference spectra at 20°C and 77 K in the digitonin fractions. (royalsocietypublishing.org)
- 22) Name the eukaryotic organisms without mitochondria? (blogspot.com)
- Mitochondrial Calcium Uniporter (MCU)-mediated calcium flux is the primary way in which calcium enters the mitochondria. (bio-protocol.org)
- One, calcium in the mitochondria activates key dehydrogenases in the Krebs cycle which leads to increased ATP production. (bio-protocol.org)
- Following this, calcium is added to the cell-suspension and MCU-mediated calcium flux can be followed by observing the changing fluorescence of the fluorophore, which cannot follow calcium into the mitochondria. (bio-protocol.org)
- The PPR40 protein is localized in the mitochondria and found in association with Complex III of the electron transport system. (plantphysiol.org)
- Additionally, troubleshooting advice as well as specific substrates, inhibitors and uncouplers that may be used to determine mitochondria function at specific sites of the electron transport chain are provided. (jove.com)
- Most mitochondria-based methods used to investigate toxins require the use of relatively large amounts of material and hence compromised sensitivity in assay. (worldcat.org)
- Respiration is assessed in digitonin-permeabilized RPTC as state 3 (maximum respiration in the presence of excess substrates and ADP) and uncoupled respirations. (nih.gov)
- Mitochondria remain intact, uninjured, and in their natural environment where mitochondrial respiration can be measured in situ under physiologically relevant conditions. (worldcat.org)
- 7) Who first related mitochondria with cell respiration? (blogspot.com)
- 34) What kind of respiration is performed by mitochondria? (blogspot.com)
- Mutations in anc-1 (nuclear anchorage defective) disrupt the positioning of nuclei and mitochondria in Caenorhabditis elegans . (sciencemag.org)
- As such, the concentrations of digitonin and cholesterol determines membrane permeability. (agscientific.com)
- Brain mitochondria from older diabetic rats were more prone to the induction of the permeability transition pore, i.e., mitochondria from 24-month-old diabetic rats accumulated much less Ca 2+ (20 μmol/l) than those isolated from 12-month-old rats (50 μmol/l) or 1.5-month-old rats (100 μmol/l). (diabetesjournals.org)
- In the mammalian host, the bloodstream form (BF) of the T. brucei mitochondrion lacks a complete respiratory chain, and energy production occurs via glycolysis ( 7 ). (asm.org)
- Furthermore, it has also been shown that SIL A can regulate the Ca 2+ influx into mitochondria [ 9 ]. (hindawi.com)
- Digitonin targets lipid rafts in the plasma membrane layer, resulting in disrupted bilayers. (agscientific.com)
- Cytosolic TERC-53 levels respond to mitochondrial functions, but have no direct effect on these functions, suggesting that cytosolic TERC-53 functions downstream of mitochondria as a signal of mitochondrial functions. (springer.com)
- Bcl-2 overexpression attenuated TRAIL-induced cleavage of caspase-8, indicating its activation upstream and downstream of mitochondria, as well as cleavage of Bid and caspase-3. (aacrjournals.org)
- 13) Which apparatus is used to study the ultrastructure of mitochondria? (blogspot.com)