Conformational Changes of Mitochondria associated with Uncoupling of Oxidative Phosphorylation in vivo and in vitro

Nature ◽  
1970 ◽  
Vol 226 (5242) ◽  
pp. 272-274 ◽  
Author(s):  
P. BUFFA ◽  
V. GUARRIERA-BOBYLEVA ◽  
U. MUSCATELLO ◽  
I. PASQUALI-RONCHETTI
Keyword(s):  
Oxidative Phosphorylation ◽  
Conformational Changes ◽  
Uncoupling Of Oxidative Phosphorylation

THE UNCOUPLING OF OXIDATIVE PHOSPHORYLATION BY IONIZING RADIATION

1962 ◽  
Vol 40 (8) ◽  
pp. 1025-1042 ◽  
Author(s):  
J. F. Scaife ◽  
B. Hill
Keyword(s):  
Oxidative Phosphorylation ◽  
Liver Mitochondria ◽  
Cell Suspensions ◽  
Whole Body ◽  
X Rays ◽  
Succinate Oxidation ◽  
Mouse Ascites ◽  
Uncoupling Of Oxidative Phosphorylation

Whole body irradiation of rabbits or rats with X-rays or Co60γ-rays causes uncoupling of oxidative phosphorylation in thymus mitochondria, which is not prevented by the prior administration of AET. Whole body irradiation was not found to affect oxidative phosphorylation in liver or mouse ascites cell mitochondria. The radiation lesion can be repaired in vitro by the addition of cytochrome c, bovine serum albumin, or vitamin K1to mitochondria. Vitamin E and coenzyme Q10 were without effect. Both phosphorylating steps in the electron transport chain associated with succinate oxidation are affected by irradiation. The diphosphopyridine nucleotide dependent steps in the oxidation of α-ketoglutarate by thymus mitochondria are damaged by in vivo irradiation. Diphosphopyridine nucleotide levels of thymus and spleen but not liver or ascites cells are reduced by in vivo irradiation. No effect of in vitro irradiation on oxidative phosphorylation could be found for thymocyte cell suspensions, isolated thymus or liver mitochondria, or ascites or HeLa cell suspensions. Respiration of ascites or thymocyte cells was unaffected by in vitro irradiation.

THE UNCOUPLING OF OXIDATIVE PHOSPHORYLATION BY IONIZING RADIATION

1962 ◽  
Vol 40 (1) ◽  
pp. 1025-1042 ◽  
Author(s):  
J. F. Scaife ◽  
B. Hill
Keyword(s):  
Oxidative Phosphorylation ◽  
Liver Mitochondria ◽  
Cell Suspensions ◽  
Whole Body ◽  
X Rays ◽  
Succinate Oxidation ◽  
Mouse Ascites ◽  
Uncoupling Of Oxidative Phosphorylation

Whole body irradiation of rabbits or rats with X-rays or Co60γ-rays causes uncoupling of oxidative phosphorylation in thymus mitochondria, which is not prevented by the prior administration of AET. Whole body irradiation was not found to affect oxidative phosphorylation in liver or mouse ascites cell mitochondria. The radiation lesion can be repaired in vitro by the addition of cytochrome c, bovine serum albumin, or vitamin K1to mitochondria. Vitamin E and coenzyme Q10 were without effect. Both phosphorylating steps in the electron transport chain associated with succinate oxidation are affected by irradiation. The diphosphopyridine nucleotide dependent steps in the oxidation of α-ketoglutarate by thymus mitochondria are damaged by in vivo irradiation. Diphosphopyridine nucleotide levels of thymus and spleen but not liver or ascites cells are reduced by in vivo irradiation. No effect of in vitro irradiation on oxidative phosphorylation could be found for thymocyte cell suspensions, isolated thymus or liver mitochondria, or ascites or HeLa cell suspensions. Respiration of ascites or thymocyte cells was unaffected by in vitro irradiation.

UNCOUPLING OF OXIDATIVE PHOSPHORYLATION BY VANADATE

1966 ◽  
Vol 44 (7) ◽  
pp. 983-988 ◽  
Author(s):  
John N. Hathcock ◽  
C. H. Hill ◽  
S. B. Tove
Keyword(s):  
Electron Transport ◽  
Oxidative Phosphorylation ◽  
Adenosine Triphosphate ◽  
Adenosine Diphosphate ◽  
Liver Mitochondria ◽  
In Vitro Studies ◽  
Exchange Reactions ◽  
Uncoupling Of Oxidative Phosphorylation

The addition of ammonium metavanadate to the diet of chicks at a level to supply 25 parts per million vanadium uncoupled oxidative phosphorylation in mitochondria isolated from the livers. In vitro studies revealed that 1 mM vanadate uncoupled oxidative phosphorylation in liver mitochondria. This uncoupling was manifest whether succinate or β-hydroxybutyrate was used as the substrate, suggesting that all three phosphorylating sites associated with electron transport were uncoupled.At a concentration of 0.1 mM, vanadate increased the destruction of adenosine triphosphate by mitochondria. As the concentration of vanadate was increased the destruction of adenosine triphosphate became progressively less. The exchange reactions of adenosine triphosphate with orthophosphate and with adenosine diphosphate, catalyzed by liver mitochondria, were inhibited by 0.1 mM vanadate. These results suggest the possibility that the known toxic effects of vanadium in vivo are related to the uncoupling of oxidative phosphorylation.

CarR, a MarR-family regulator from Corynebacterium glutamicum, modulated antibiotic and aromatic compound resistance

2019 ◽  
Vol 476 (21) ◽  
pp. 3141-3159 ◽  
Author(s):  
Meiru Si ◽  
Can Chen ◽  
Zengfan Wei ◽  
Zhijin Gong ◽  
GuiZhi Li ◽  
...  
Keyword(s):  
Stress Response ◽  
Ligand Binding ◽  
Corynebacterium Glutamicum ◽  
Conformational Changes ◽  
Cysteine Oxidation ◽  
Transcriptional Regulatory ◽  
Ligand Free ◽  
Redox Sensing

Abstract MarR (multiple antibiotic resistance regulator) proteins are a family of transcriptional regulators that is prevalent in Corynebacterium glutamicum. Understanding the physiological and biochemical function of MarR homologs in C. glutamicum has focused on cysteine oxidation-based redox-sensing and substrate metabolism-involving regulators. In this study, we characterized the stress-related ligand-binding functions of the C. glutamicum MarR-type regulator CarR (C. glutamicum antibiotic-responding regulator). We demonstrate that CarR negatively regulates the expression of the carR (ncgl2886)–uspA (ncgl2887) operon and the adjacent, oppositely oriented gene ncgl2885, encoding the hypothetical deacylase DecE. We also show that CarR directly activates transcription of the ncgl2882–ncgl2884 operon, encoding the peptidoglycan synthesis operon (PSO) located upstream of carR in the opposite orientation. The addition of stress-associated ligands such as penicillin and streptomycin induced carR, uspA, decE, and PSO expression in vivo, as well as attenuated binding of CarR to operator DNA in vitro. Importantly, stress response-induced up-regulation of carR, uspA, and PSO gene expression correlated with cell resistance to β-lactam antibiotics and aromatic compounds. Six highly conserved residues in CarR were found to strongly influence its ligand binding and transcriptional regulatory properties. Collectively, the results indicate that the ligand binding of CarR induces its dissociation from the carR–uspA promoter to derepress carR and uspA transcription. Ligand-free CarR also activates PSO expression, which in turn contributes to C. glutamicum stress resistance. The outcomes indicate that the stress response mechanism of CarR in C. glutamicum occurs via ligand-induced conformational changes to the protein, not via cysteine oxidation-based thiol modifications.

scholarly journals E3 Ubiquitin Ligase SPL2 Is a Lanthanide-Binding Protein

2021 ◽  
Vol 22 (11) ◽  
pp. 5712
Author(s):  
Michał Tracz ◽  
Ireneusz Górniak ◽  
Andrzej Szczepaniak ◽  
Wojciech Białek
Keyword(s):  
Ubiquitin Ligase ◽  
Conformational Changes ◽  
Protein Interactions ◽  
E3 Ubiquitin Ligase ◽  
Lanthanide Ions ◽  
E3 Ligases ◽  
Fluorescence Measurements ◽  
Partial Unfolding

The SPL2 protein is an E3 ubiquitin ligase of unknown function. It is one of only three types of E3 ligases found in the outer membrane of plant chloroplasts. In this study, we show that the cytosolic fragment of SPL2 binds lanthanide ions, as evidenced by fluorescence measurements and circular dichroism spectroscopy. We also report that SPL2 undergoes conformational changes upon binding of both Ca2+ and La3+, as evidenced by its partial unfolding. However, these structural rearrangements do not interfere with SPL2 enzymatic activity, as the protein retains its ability to auto-ubiquitinate in vitro. The possible applications of lanthanide-based probes to identify protein interactions in vivo are also discussed. Taken together, the results of this study reveal that the SPL2 protein contains a lanthanide-binding site, showing for the first time that at least some E3 ubiquitin ligases are also capable of binding lanthanide ions.

scholarly journals Furin Processing and Proteolytic Activation of Semliki Forest Virus

2003 ◽  
Vol 77 (5) ◽  
pp. 2981-2989 ◽  
Author(s):  
Xinyong Zhang ◽  
Martin Fugère ◽  
Robert Day ◽  
Margaret Kielian
Keyword(s):  
Conformational Changes ◽  
Secretory Pathway ◽  
Semliki Forest Virus ◽  
Fusion Reaction ◽  
Low Ph ◽  
Protein Fusion ◽  
Proteolytic Activation ◽  
Control Virus

ABSTRACT The alphavirus Semliki Forest virus (SFV) infects cells via a low-pH-dependent membrane fusion reaction mediated by the E1 envelope protein. Fusion is regulated by the interaction of E1 with the receptor-binding protein E2. E2 is synthesized as a precursor termed “p62,” which forms a stable heterodimer with E1 and is processed late in the secretory pathway by a cellular furin-like protease. Once processing to E2 occurs, the E1/E2 heterodimer is destabilized so that it is more readily dissociated by exposure to low pH, allowing fusion and infection. We have used FD11 cells, a furin-deficient CHO cell line, to characterize the processing of p62 and its role in the control of virus fusion and infection. p62 was not cleaved in FD11 cells and cleavage was restored in FD11 cell transfectants expressing human furin. Studies of unprocessed virus produced in FD11 cells (wt/p62) demonstrated that the p62 protein was efficiently cleaved by purified furin in vitro, without requiring prior exposure to low pH. wt/p62 virus particles were also processed during their endocytic uptake in furin-containing cells, resulting in more efficient virus infection. wt/p62 virus was compared with mutant L, in which p62 cleavage was blocked by mutation of the furin-recognition motif. wt/p62 and mutant L had similar fusion properties, requiring a much lower pH than control virus to trigger fusion and fusogenic E1 conformational changes. However, the in vivo infectivity of mutant L was more strongly inhibited than that of wt/p62, due to additional effects of the mutation on virus-cell binding.

scholarly journals Let-7a regulates EV secretion and mitochondrial oxidative phosphorylation by targeting SNAP23 in colorectal cancer

2021 ◽  
Vol 40 (1) ◽  
Author(s):  
You Dong Liu ◽  
Xiao Peng Zhuang ◽  
Dong Lan Cai ◽  
Can Cao ◽  
Qi Sheng Gu ◽  
...  
Keyword(s):  
Oxidative Phosphorylation ◽  
Metabolic Reprogramming ◽  
Luciferase Reporter ◽  
Mitochondrial Oxidative Phosphorylation ◽  
In Vivo Assays ◽  
Reporter Assays ◽  
Extracellular Mirna

Abstract Background MicroRNAs (miRNAs) are abundant in tumor-derived extracellular vesicles (EVs) and the functions of extracellular miRNA to recipient cells have been extensively studied with tumorigenesis. However, the role of miRNA in EV secretion from cancer cells remains unknown. Methods qPCR and bioinformatics analysis were applied for determining extracellular let-7a expression from CRC patient serum and cells. Nanosight particle tracking analysis was performed for investigating the effect of let-7a on EV secretion. Luciferase reporter assays was used for identifying targeted genes synaptosome-associated protein 23 (SNAP23). In vitro and in vivo assays were used for exploring the function of let-7a/SNAP23 axis in CRC progression. Bioenergetic assays were performed for investigating the role of let-7a/SNAP23 in cellular metabolic reprogramming. Results let-7a miRNA was elevated in serum EVs from CRC patients and was enriched in CRC cell-derived EVs. We determined that let-7a could suppress EV secretion directly targeting SNAP23. In turn, SNAP23 promotes EV secretion of let-7a to downregulate the intracellular let-7a expression. In addition, we found a novel mechanism of let-7a/SNAP23 axis by regulating mitochondrial oxidative phosphorylation (OXPHOS) through Lin28a/SDHA signaling pathway. Conclusions Let-7a plays an essential role in not only inhibiting EV secretion, but also suppressing OXPHOS through SNAP23, resulting in the suppression of CRC progression, suggesting that let-7a/SNAP23 axis could provide not only effective tumor biomarkers but also novel targets for tumor therapeutic strategies.

scholarly journals IL-32 is a metabolic regulator promoting survival and proliferation of malignant plasma cells

2021 ◽  
Author(s):  
Kristin Roseth Aass ◽  
Robin Mjelle ◽  
Martin H. Kastnes ◽  
Synne S. Tryggestad ◽  
Luca M. van den Brink ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Oxidative Phosphorylation ◽  
Plasma Cells ◽  
Inflammatory Diseases ◽  
Mitochondrial Respiratory Chain ◽  
Gene Signature ◽  
Growth And Survival ◽  
Novel Concept

AbstractIL-32 is a non-classical cytokine expressed in cancers, inflammatory diseases and infections. IL-32 can have both extracellular and intracellular functions, and its receptor is not identified. We here demonstrate that endogenously expressed, intracellular IL-32 binds to components of the mitochondrial respiratory chain and promotes oxidative phosphorylation. Knocking out IL-32 in malignant plasma cells significantly reduced survival and proliferation in vitro and in vivo. High throughput transcriptomic and MS-metabolomic profiling of IL-32 KO cells revealed that loss of IL-32 leads to profound perturbations in metabolic pathways, with accumulation of lipids, pyruvate precursors and citrate, indicative of reduced mitochondrial function. IL-32 is expressed in a subgroup of multiple myeloma patients with an inferior prognosis. Primary myeloma cells expressing IL-32 were characterized by a plasma cell gene signature associated with immune activation, proliferation and oxidative phosphorylation. We propose a novel concept for regulation of metabolism by an intracellular cytokine and identify IL-32 as an endogenous growth and survival factor for malignant plasma cells. IL-32 is a potential prognostic biomarker and a treatment target in multiple myeloma.

In Vivo Retrieval of a Trapped Merci Embolectomy Device: Technical Case Report

2010 ◽  
Vol 67 (3) ◽  
pp. onsE304-onsE304 ◽  
Author(s):  
Ajeet Gordhan ◽  
John Soliman
Keyword(s):  
Vertebral Artery ◽  
Conformational Changes ◽  
Basilar Artery ◽  
Technical Note ◽  
Vessel Injury ◽  
Artery Segment ◽  
In Vitro Assessment ◽  
Artery Flow

Abstract BACKGROUND AND IMPORTANCE: This technical note describes a complication related to the use of the Merci embolectomy device not previously reported. The device can induce critical flow limitation within an accessed vessel because of a combination of vasospasm and anatomic conformational changes. Furthermore, this can limit the safe removal of the device from intracranial vasculature. We present a novel rescue technique that can be used to safely retrieve the entrapped Merci device without inciting localized vessel injury. CLINICAL PRESENTATION: A 51-year-old male with embolic occlusion of the distal basilar artery and dissection-related occlusion of the left cervical vertebral underwent mechanical thrombolysis. Flow-limiting vasospasm and/or anatomic conformational changes/ telescoping of the intracranial right vertebral artery segment was induced during deployment with subsequent entrapment of the device. Reclamation of the entrapped device was performed by initially removing the Merci microcatheter. The entrapped and fixated device was then resheathed into a 4F slip catheter within the intracranial vertebral artery. The Merci device and the slip catheter were then removed. Right vertebral and proximal basilar artery flow was reestablished after removal of the Merci device. Successful clot extraction was thereafter performed using a microsnare. CONCLUSION: In vitro assessment of the device has demonstrated its propensity to induce vasospasm. In vivo entrapment of the device has not been previously reported. Successful retrieval can be achieved if the Merci device becomes entrapped and fixated. This may be an important consideration as increased utilization of the device occurs.

A dual-action peptide-containing hydrogel targets wound infection and inflammation

2020 ◽  
Vol 12 (524) ◽  
pp. eaax6601 ◽  
Author(s):  
Manoj Puthia ◽  
Marta Butrym ◽  
Jitka Petrlova ◽  
Ann-Charlotte Strömdahl ◽  
Madelene Å. Andersson ◽  
...  
Keyword(s):  
Conformational Changes ◽  
Inflammatory Responses ◽  
Therapeutic Potential ◽  
Nuclear Factor Κb ◽  
Reporter Mice ◽  
Dual Action ◽  
Infection And Inflammation ◽  
Molecular Patterns

There is a clinical need for improved wound treatments that prevent both infection and excessive inflammation. TCP-25, a thrombin-derived peptide, is antibacterial and scavenges pathogen-associated molecular patterns (PAMPs), such as lipopolysaccharide, thereby preventing CD14 interaction and Toll-like receptor dimerization, leading to reduced downstream immune activation. Here, we describe the development of a hydrogel formulation that was functionalized with TCP-25 to target bacteria and associated PAMP-induced inflammation. In vitro studies determined the polymer prerequisites for such TCP-25–mediated dual action, favoring the use of noncharged hydrophilic hydrogels, which enabled peptide conformational changes and LPS binding. The TCP-25–functionalized hydrogels killed Gram-positive Staphylococcus aureus and Gram-negative Pseudomonas aeruginosa bacteria in vitro, as well as in experimental mouse models of subcutaneous infection. The TCP-25 hydrogel also mediated reduction of LPS-induced local inflammatory responses, as demonstrated by analysis of local cytokine production and in vivo bioimaging using nuclear factor κB (NF-κB) reporter mice. In porcine partial thickness wound models, TCP-25 prevented infection with S. aureus and reduced concentrations of proinflammatory cytokines. Proteolytic fragmentation of TCP-25 in vitro yielded a series of bioactive TCP fragments that were identical or similar to those present in wounds in vivo. Together, the results demonstrate the therapeutic potential of TCP-25 hydrogel, a wound treatment based on the body’s peptide defense, for prevention of both bacterial infection and the accompanying inflammation.

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