Loss of PLA2G6 leads to elevated mitochondrial lipid peroxidation and mitochondrial dysfunction.
Bottom Line: Furthermore, we demonstrate that loss of iPLA2-VIA function leads to a number of mitochondrial abnormalities, including mitochondrial respiratory chain dysfunction, reduced ATP synthesis and abnormal mitochondrial morphology.Moreover, we show that loss of iPLA2-VIA is strongly associated with increased lipid peroxidation levels.Similar abnormalities were seen including elevated mitochondrial lipid peroxidation and mitochondrial membrane defects, as well as raised levels of cytoplasmic and mitochondrial reactive oxygen species.
Affiliation: 1 Institute of Healthy Ageing and Department of Genetics, Evolution and Environment, University College London, London WC1E 6BT, UK 2 Institute of Neurology, University College London, Queen Square, London WC1N 3BG, UK email@example.com.Show MeSH
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Mentions: Given the abnormal mitochondrial membrane morphology in older iPLA2-VIA−/− flies, we investigated whether loss of iPLA2-VIA affected mitochondrial membrane potential (ΔΨm), an indicator of mitochondrial health and function. Mitochondrial membrane potential was assessed in the brains of 25-day-old flies using TMRM fluorescence and imaging with confocal microscopy. This revealed a significant reduction in mitochondrial membrane potential in iPLA2-VIA−/− fly brains compared to controls (Fig. 3A). We next isolated mitochondria from iPLA2-VIA−/− flies and assessed respiratory chain activity by measuring oxygen consumption using a Clark-type oxygen electrode. The steps in respiration were compared in mitochondria isolated from iPLA2-VIA−/− flies and control w1118 flies with substrates and inhibitors specific to individual respiratory complexes. We found that knockout of iPLA2-VIA activity conferred a decrease in complex I and II-dependent respiration (Fig. 3B), manifested as significantly decreased oxygen consumption in state 3 respiration, whether using substrates for complex I (5 mM glutamate and 5 mM malate) or for complex II (5 mM succinate in the presence of rotenone) (Fig. 3B). The respiratory control ratio is the ratio of state 3 respiration (ADP stimulated) to state 4 respiration (no ADP present), and is considered an indication of the degree of coupling of mitochondrial respiratory chain activity to oxidative phosphorylation (Chance and Williams, 1955). The respiratory control ratio was significantly lower in iPLA2-VIA−/− mitochondria compared with age-matched control w1118 flies (Fig. 3C), because of the inhibition of state 3 respiration. Furthermore, the respiratory control ratio of iPLA2-VIA−/− flies in the presence of glutamate/malate was dependent on the age of the flies: older flies having lower respiratory control ratio values (Fig. 3C). Thus, loss of iPLA2-VIA activity reduces mitochondrial membrane potential as a result of mitochondrial uncoupling, which increases with age in the iPLA2-VIA−/− flies.Figure 3
Affiliation: 1 Institute of Healthy Ageing and Department of Genetics, Evolution and Environment, University College London, London WC1E 6BT, UK 2 Institute of Neurology, University College London, Queen Square, London WC1N 3BG, UK firstname.lastname@example.org.