Fig. 6

Dependence on mitochondrial energetic flux renders TCL1-positive leukemic cells more susceptible towards agents interfering with mitochondrial homeostasis. (a) Representative immunoblots on cytosolic and nuclear fractions from 3 experiments on CLL-like JVM3 cells and their TCL1-positive sub-line after 24 h of metal-containing nucleoside treatment. Treatment with these substances reduces the levels of the mitochondrial proteins AIF, smac, and cytochrome c in this subcellular fraction, specifically in the TCL1-overexpressing sub-line. (b, c) Mitochondrial respiration (oxygen consumption rate; OCR) and extracellular acidification rate (ECAR; the result of anaerobic glycolysis) were measured (XFe96 flux analyzer) under basal conditions and in response to the indicated substances in JVM3 cells (control) and those transfected with TCL1. (b) Left: baseline OCR is similar between JVM3 cells and their stable TCL1 transfectants, while a lower degree of ECAR in the presence of overexpressed TCL1 is noted. (c) The effects on OCR are shown as percentages of baseline (set as 100 %) for each treatment (substances added after 3^rd, 6^th, and 9^th time-point). The stronger changes in OCR after oligomycin (ATP synthase inhibitor to differentiate the ATP-linked respiration from the proton leak) in the TCL1-positive sub-line are indicative of higher degrees of respiration (OCR) linked to ATP synthesis. Maximal respiratory capacity as determined by addition of the uncoupling agent FCCP was not different. Antimycin A/rotenone stop the reaction (respiration) and the remaining level of OCR (non-mitochondrial respiration) is lower in the TCL1-carrying sub-line. All experiments of (b)-(c) were performed at least 6 times with *** indicating P < 0.001 and bars for the standard error of the mean