Type

Journal Article

Authors

D B Papkovsky
P J Pollard
R O'Connor
J Adam
L O'Flaherty
C Favre
A V Zhdanov

Subjects

Biochemistry

Topics
rna interference deficiency oxidative phosphorylation physiology oligomycins fumarate hydratase pharmacology embryo mammalian animals oxygen consumption glycolysis extracellular space gene deletion citric acid cycle fibroblasts energy metabolism cytology gene knockout techniques mice drug effects humans pnc1 protein human mitochondrial proteins hydrogen ion concentration nucleotide transport proteins hela cells lactic acid cell respiration rna small interfering carbon dioxide carbonyl cyanide p trifluoromethoxyphenylhydrazone genetics metabolism adenosine triphosphate

Comparative bioenergetic assessment of transformed cells using a cell energy budget platform. (2011)

Abstract The aberrant expression and functional activity of proteins involved in ATP production pathways may cause a crisis in energy generation for cells and compromise their survival under stressful conditions such as excitation, starvation, pharmacological treatment or disease states. Under resting conditions such defects are often compensated for, and therefore masked by, alternative pathways which have significant spare capacity. Here we present a multiplexed 'cell energy budget' platform which facilitates metabolic assessment and cross-comparison of different cells and the identification of genes directly or indirectly involved in ATP production. Long-decay emitting O(2) and pH sensitive probes and time-resolved fluorometry are used to measure changes in cellular O(2) consumption, glycolytic and total extracellular acidification (ECA), along with the measurement of total ATP and protein content in multiple samples. To assess the extent of spare capacity in the main energy pathways, the cells are also analysed following double-treatment with carbonyl cyanide p-(trifluoromethoxy)phenylhydrazone and oligomycin. The four-parametric platform operating in a high throughput format has been validated with two panels of transformed cells: mouse embryonic fibroblasts (MEFs) lacking the Krebs cycle enzyme fumarate hydratase (Fh1) and HeLa cells with reduced expression of pyrimidine nucleotide carrier 1. In both cases, a marked reduction in both respiration and spare respiratory capacity was observed, accompanied by a compensatory activation of glycolysis and consequent maintenance of total ATP levels. At the same time, in Fh1-deficient MEFs the contribution of non-glycolytic pathways to the ECA did not change.
Collections Ireland -> University College Cork -> PubMed

Full list of authors on original publication

D B Papkovsky, P J Pollard, R O'Connor, J Adam, L O'Flaherty, C Favre, A V Zhdanov

Experts in our system

1
Dmitri B Papkovsky
University College Cork
Total Publications: 89
 
2
Robert O'Connor
Dublin City University
Total Publications: 74
 
3
C Favre
University College Cork
Total Publications: 4
 
4
Alexander V Zhdanov
University College Cork
Total Publications: 37