Cytochrome c, released from cerebellar granule cells undergoing apoptosis or excytotoxic death, can generate protonmotive force and drive ATP synthesis in isolated mitochondria

Abstract

In rat cerebellar granule cells, cytochrome c release takes place during glutamate toxicity and apoptosis due to deprivation of depolarising levels of potassium. We show that, as in necrosis, the released cytochrome c present in the cytosolic fraction obtained from cerebellar granule cells undergoing apoptosis can operate as a reactive oxygen species (ROS) scavenger and as a respiratory substrate. The capability of the cytosolic fraction containing cytochrome c, obtained from cerebellar granule cells undergoing either necrosis or apoptosis, to energise coupled mitochondria isolated by the same cells is also investigated. We show that, in both cases, the cytosolic fraction containing cytochrome c, added to mitochondria, can cause proton ejection, and membrane potential generation and can drive ATP synthesis and export in the extramitochondrial phase, as photometrically measured via the ATP detecting system. Cytochrome c, separated immunologically from the cytosolic fraction of apoptotic cells when added to mitochondria, is found to cause proton ejection to generate membrane potential and to drive ATP synthesis and export in a manner not sensitive to the further addition of the cytosolic fraction depleted of cytochrome c, which failed to do this. In the light of these findings we propose that in apoptosis the released cytochrome c can contribute to provide ATP required for the cell programmed death to occur.

Akt-dependent Activation of the Heart 6-Phosphofructo-2-kinase/Fructose-2,6-bisphosphatase (PFKFB2) Isoenzyme by Amino Acids

“…The pellet was resuspended in somatic cell ATP-releasing reagent (Sigma). ATP was determined in the pellet and supernatant as described in Atlante et al (19) with some modifications. The composition of the assay buffer used was 10 mM phosphate buffer (pH 8), 2.5 mM glucose, hexokinase-glucose-6-phosphate dehydrogenase (0.5 specific units) and 0.2 mM NADP.…”

Does the term ‘trophic’ actually mean anti-amyloidogenic? The case of NGF

“…36 In addition, the pro-apoptotic shifting to a low potassium medium induced a marked failure of mitochondrial oxidative phosphorylation, accompanied by a dramatic intracellular ATP drop and by an increased production of reactive oxidative species. [37][38][39] A functional impairment of proteasomes, causing a progressive accumulation of ubiquitinated-unfolded proteins in the cytoplasm, 40 and an early perturbation of autophagiclysosomal structures were also revealed in apoptotic neurons. 41 Interestingly, co-culture experiments showed that an Ab antibody partially inhibited the 'transfer' of apoptotic process to healthy, but separated neighboring neurons.…”

Cytochrome c Is Released in a Reactive Oxygen Species-Dependent Manner and Is Degraded via Caspase-Like Proteases in Tobacco Bright-Yellow 2 Cells en Route to Heat Shock-Induced Cell Death

“…On the other hand, among a variety of events occurring during PCD, the release of cytochrome c (cyt c) has been shown both in mammalian and plant mitochondria. In the former case, it was shown that the released cyt c (1) can work as a ROS scavenger and as an electron donor to cytochrome oxidase, thus driving ATP synthesis, which is necessary for apoptosis to occur (Atlante et al, 2003a(Atlante et al, , 2003b and (2) binds to apoptotic protease-activating factor (Apaf-1) protein and, in the presence of ATP/dATP, activates caspase-9, thus triggering activation of a caspase cascade that leads to the morphological changes typical of apoptosis (Li et al, 1997;Zou et al, 1999). In plant cells, cyt c release was already shown in a variety of cases, including menadione-induced death in tobacco protoplasts (Sun et al, 1999), D-Mannose-induced Cyt c release from mitochondria was investigated by immunoblot analysis using a monoclonal antibody against cyt c. Both cytosolic and mitochondrial fractions, obtained from TBY-2 cells subjected to HS (cells in these conditions will be referred to as HS cells), were examined.…”