Lemieux 2019 bioRxiv
|Lemieux H, Subarsky P, Doblander C, Wurm M, Troppmair J, Gnaiger E (2019) Mitochondrial respiratory function as an early biomarker of apoptosis induced by growth factor removal. bioRxiv doi: https://doi.org/10.1101/151480 .|
Abstract: Remodeling of mitochondrial metabolism is implicated in progression of cancer. Conversely, however, mitochondrial dysfunction and signaling play key roles in the induction of cell death. Apoptosis is induced following interleukin 3 (IL-3) depletion in mouse pro-myeloid 32D cells. Molecular signals of cell death are absent in 32D cells after short-term IL-3 deprivation (8 h). We addressed the question if changes in mitochondrial function can be detected by high-resolution respirometry as an early event in the induction of apoptosis. Respiration of living 32D cells was suppressed by 10 to 55% following 8 h removal of IL-3, but remained more stable in 32D cells expressing the v-RAF oncogene related to CRAF. In 32D cells deprived of IL3, succinate-supported respiration did not decline significantly, but respiratory capacities of the NADH-pathway and the combined NADH- and succinate-linked (NS) pathway were decreased compared to cells grown in the presence of IL-3. This was consistent with respiratory control exerted by impaired Complex IV activity, since there was not even the slightest excess Complex IV capacity above NS-pathway capacity. In contrast, electron flow reached only 60% when supported by succinate alone through Complexes II, III and IV, and was therefore relatively insensitive to Complex IV injuries up to a threshold of 40% inhibition. After IL-3 depletion respiration increased by 15% following addition of cytochrome c, which provides a marker of mitochondrial outer membrane leakage, thus indicating mitochondrial fragility. Our results highlight a novel link between the key mitogenic and survival kinase CRAF and mitochondrial energy homeostasis.
• Keywords: Mitochondrial respiration, OXPHOS, cytochrome c oxidase, apoptosis, CRAF, interleukin 3 • Bioblast editor: Gnaiger E • O2k-Network Lab: CA_Edmonton_Lemieux H, AT_Innsbruck_Oroboros, AT_Innsbruck_Gnaiger E
Labels: MiParea: Respiration Pathology: Cancer Stress:Cell death Organism: Mouse Tissue;cell: Blood cells Preparation: Intact cells, Permeabilized cells Enzyme: Complex IV;cytochrome c oxidase, Marker enzyme Regulation: Coupling efficiency;uncoupling, Cyt c, Flux control, Inhibitor, Threshold;excess capacity, Uncoupler, Q-junction effect Coupling state: LEAK, ROUTINE, OXPHOS, ET Pathway: N, S, CIV, NS, ROX HRR: Oxygraph-2k
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