1,721,044 research outputs found
In vivo measurements of respiration control by cytochrome c oxidase and in situ analysis of oxidative phosphorylation
No full textIn vivo control of respiration by cytochrome c oxidase in wild-type and mitochondrial DNA mutation-carrying human cells
No full textThe metabolic control of respiration is still poorly understood, due mainly to the lack of suitable approaches for studying it in vivo. Experiments on isolated mammalian mitochondria have indicated that a relatively small fraction of each of several components of the electron transport chain is sufficient to sustain a normal O2 consumption rate. These experiments, however, may not reflect accurately the in vivo situation, due to the lack in the mitochondrial fraction of essential cytosolic components and to the use of excess of substrates in the in vitro assays. An approach is described here whereby the control of respiration by cytochrome c oxidase (COX; EC 1.9.3.1) was analyzed in intact cultured human osteosarcoma 143B.TK- cells and other wild-type cells and in mitochondrial DNA mutation- carrying human cell lines. Surprisingly, in wild-type cells, only a slightly higher COX capacity was detected than required to support the endogenous respiration rate, pointing to a tighter in vivo control of respiration by COX than generally assumed. Cell lines carrying the MERRF mitochondrial tRNA(Lys) gene mutation, which causes a pronounced decrease in mitochondrial protein synthesis and respiration rates, revealed, in comparison, a significantly greater COX capacity relative to the residual endogenous respiration rate, and, correspondingly, a higher COX inhibition threshold above which the overall respiratory flux was affected. The observed relationship between COX respiratory threshold and relative COX capacity and the potential extension of the present analysis to other respiratory complexes have significant general implications for understanding the pathogenetic role of mutations in mtDNA-linked diseases and the tissue specificity of the mutation-associated phenotype
Identification of discrete electrophoretic components among the products of mitochondrial protein synthesis
No full textIn vivo control of respiration by cytochrome c oxidase in human cells
No full textThe metabolic control of oxidative phosphorylation (OXPHOS) has attracted increasing attention in recent years, especially due to its importance for understanding the role of mitochondrial DNA mutations in human diseases and aging. Experiments on isolated mitochondria have indicated that a relatively small fraction of each of several components of the electron transport chain is sufficient to sustain a normal respiration rate. These experiments, however, may have not reflected the in vivo situation, due to the possible loss of essential metabolites during organelle isolation and the disruption of the normal interactions of mitochondria with the cytoskeleton, which may be important for the channeling of respiratory substrate to the organelles. To obtain direct evidence on this question, in particular, as concerns the in vivo control of respiration by cytochrome c oxidase (COX), we have developed an approach for measuring COX activity in intact cells, by means of cyanide titration, either as an isolated step or as a respiratory chain-integrated step. The method has been applied to a variety of human cell types, including wild-type and mtDNA mutation-carrying cells, several tumor- derived semidifferentiated cell lines, as well as specialized cells removed from the organism. The results obtained strongly support the following conclusions: (i) the in vivo control of respiration by COX is much tighter than has been generally assumed on the basis of experiments carried out on isolated mitochondria; (ii) COX thresholds depend on the respiratory fluxes under which they are measured; and (iii) measurements of relative enzyme capacities are needed for understanding the role of mitochondrial respiratory complexes in human physiopathology
Rate-limiting step preceding cytochrome c release in cells primed for Fas-mediated apoptosis revealed by analysis of cellular mosaicism of respiratory changes
No full textIn the present work, Jurkat cells undergoing anti-Fas antibody (anti-Fas)-triggered apoptosis exhibited in increasing proportion a massive release of cytochrome c from mitochondria, as revealed by double-labeling confocal immunofluorescence microscopy. The cytochrome c release was followed by a progressive reduction in the respiratory activity of the last respiratory enzyme, cytochrome c oxidase (COX), and with a little delay, by a decrease in overall endogenous respiration rate, as measured in vivo in the whole cell population. Furthermore, in vivo titration experiments showed that an ∼30% excess of COX capacity over that required to support endogenous respiration, found in naive cells, was maintained in anti-Fas-treated cells having lost ∼40% of their COX respiratory activity. This observation strongly suggested that only a subpopulation of anti-Fas-treated cells, which maintained the excess of COX capacity, respired. Fractionation of cells on annexin V-coated paramagnetic beads did indeed separate a subpopulation of annexin V-binding apoptotic cells with fully released cytochrome c and completely lacking respiration, and a nonbound cell subpopulation exhibiting nearly intact respiration and in their great majority preserving the mitochondrial cytochrome c localization. The above findings showed a cellular mosaicism in cytochrome c release and respiration loss, and revealed the occurrence of a rate-limiting step preceding cytochrome c release in the apoptotic cascade. Furthermore, the striking observation that controlled digitonin treatment caused a massive and very rapid release of cytochrome c and complete loss of respiration in the still respiring anti-Fas-treated cells, but not in naive cells, indicated that the cells responding to digitonin had already been primed for apoptosis, and that this treatment bypassed or accelerated the rate-limiting step most probably at the level of the mitochondrial outer membrane
ANTIBODIES AGAINST THE COOH-TERMINAL UNDECAPEPTIDE OF SUBUNIT-II, BUT NOT THOSE AGAINST THE NH2-TERMINAL DECAPEPTIDE, IMMUNOPRECIPITATE THE WHOLE HUMAN CYTOCHROME-C-OXIDASE COMPLEX
No full text7 UNIDENTIFIED READING FRAMES OF HUMAN MITOCHONDRIAL-DNA ENCODE SUBUNITS OF THE RESPIRATORY-CHAIN NADH DEHYDROGENASE
No full textFontana-Unipi @ HaSpeeDe2: Ensemble of Transformers for the Hate Speech Task at Evalita
Get PDFWe describe our approach and experiments to tackle Task A of the second edition of HaSpeeDe, within the Evalita 2020 evaluation campaign. The proposed model consists in an ensemble of classifiers built from three variants of a common neural architecture. Each classifier uses contextual representations from transformers trained on Italian texts, fine tuned on the training set of the challenge. We tested the proposed model on the two official test sets, the in-domain test set containing just tweets and the out-of-domain one including also news headlines. Our submissions ranked 4th on the tweets test set and 17th on the second test set
DEVELOPMENTAL REGULATION OF EXPRESSION OF MITOCHONDRIAL-DNA ENCODED ND5-SUBUNIT OF NADH DEHYDROGENASE
No full text- …
