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Genetic reduction of mitochondrial phosphatidylethanolamine in mammalian cells impairs mitochondrial function
S6
Abstracts / Chemistry and Physics of Lipids 163S (2010) S1–S18
work of fatty acid flux and its metabolic and regulatory involvement in lipid and membrane homeostasis.
Acknowledgements Supported by FWF projects 18857 and DK Molecular Enzymology W901-305. doi:10.1016/j.chemphyslip.2010.05.018 SO7 The targeting of plasmalemmal ceramide to mitochondria during apoptosis E.B. Babiychuk ∗ , K. Monastyrskaya, A. Draeger Department of Cell Biology, Institute of Anatomy, University of Bern, 3012 Bern, Switzerland Ceramide is a key lipid mediator in cellular processes such as differentiation, proliferation, growth arrest and apoptosis. Upon its genesis during apoptosis, ceramide self-associates into platforms thereby promoting extensive structural changes within the plasma membrane. In most cells, apoptosis is initiated by the release of mitochondrial proteins into the cytoplasm and there is abundant evidence to suggest that an increase in permeability is caused by the appearance of pores in the outer mitochondrial membrane. Recent data suggest that generation of intracellular ceramide increases mitochondrial permeability. Whereas mitochondria are central organelles in the ceramide-dependent induction of apoptosis, they are not held to participate in ceramide production themselves. Since intracellular ceramide transport is hampered by the insoluble nature of this molecule, it is an open question how ceramide transfer into mitochondria is effected. We have previously established annexin A1, a member of a family of Ca2+ and membrane-binding proteins, as a marker for ceramide platforms. Using fluorescently tagged annexin A1 we have shown that, upon its generation within the plasma membrane, ceramide selfassociated into platforms, which subsequently invaginated and fused with mitochondria. Electron microscopic tomography confirmed that upon ceramide platform formation, the invaginated regions of the plasma membrane extend deeply into the cytoplasm forming physical contacts with mitochondrial outer membranes. Thus, ceramide is transferred from the plasma membrane to the mitochondrial outer membrane. It is conceivable that this “Kissof-Death” increases the permeability of the outer mitochondrial membrane and triggers apoptosis.
ity and profound abnormalities in mitochondrial morphology. We have now tested the hypothesis that reduction in mitochondrial PE production via the decarboxylation of PS impairs mitochondrial function. We used three models of genetically modified Chinese hamster ovary (CHO) cells in which: (i) PS synthetic capacity was only 5% of normal, (ii) PS import into mitochondria was blocked, or (iii) PS decarboxylase mRNA was reduced by >85% by RNA silencing. In each of these models, cell viability was not impaired but the PE content of mitochondria was reduced by ∼40%, mitochondrial morphology and motility were markedly altered, cellular ATP levels were reduced, and mitochondrial membrane potential was greatly increased. These data demonstrate for the first time that the mitochondrial content of PE profoundly influences mitochondrial function. doi:10.1016/j.chemphyslip.2010.05.020 Session 3: Bioactive lipids and lipidomics PL10 Control of free arachidonic acid levels within immunoinflammatory cells by phospholipases A2 and acyltransferases Jesús Balsinde Institute of Molecular Biology and Genetics, Spanish National Research Council (CSIC), 47003 Valladolid, Spain Arachidonic acid (AA) and its oxygenated derivatives, the eicosanoids, mediate a variety of physiological and pathophysiological states. The distribution of AA among cellular glycerophospholipids is finely regulated by the CoA-dependent acylation of lysophospholipids followed by transacylation reactions. Under the appropriate stimulatory conditions, AA is liberated from its phospholipid storage sites by the action of one or various phospholipases A2 . Thus, cellular availability of AA depends on an exquisite balance between phospholipid reacylation and hydrolysis reactions (Pérez-Chacón et al., 2009; Balboa and Balsinde, 2006). Phagocytic cells mobilize large amounts of free AA during stimulation with stimuli of the innate immune response (Balsinde et al., 2000; Pérez-Chacón et al., 2010). Utilizing HPLC coupled to electrospray ionization mass spectrometry, we have characterized changes in AA-containing phospholipid species in human monocytes stimulated with zymosan (Balgoma et al., 2010). We have identified three AA-containing phospholipids, namely PI(20:4/20:4), PC(20:4/20:4) and PE(16:1/20:4) in stimulated cells which are not present in resting cells or are found at very low levels. Thus these species can be regarded as lipid markers of human monocyte activation.
doi:10.1016/j.chemphyslip.2010.05.019 SO8
References
Daniel H. Bai, Jean E. Vance ∗
Balboa, M.A., Balsinde, J., 2006. Biochim. Biophys. Acta 1761, 385–391. Balgoma, et al., 2010. J. Immunol. 184, 3857–3865. Balsinde, J., Balboa, M.A., Dennis, E.A., 2000. J. Biol. Chem. 275, 22544–22549. Pérez-Chacón, et al., 2009. Biochim. Biophys. Acta 1791, 1103–1113. Pérez-Chacón, et al., 2010. J. Immunol. 184, 1071–1078.
Group on Molecular and Cell Biology of Lipids and Department of Medicine, University of Alberta, Edmonton, AB, Canada
doi:10.1016/j.chemphyslip.2010.05.021
Genetic reduction of mitochondrial phosphatidylethanolamine in mammalian cells impairs mitochondrial function
In mammalian cells, phosphatidylethanolamine (PE) is made by two spatially distinct pathways: in the endoplasmic reticulum (ER) [from the CDP-ethanolamine pathway] and in mitochondria [from the decarboxylation of phosphatidylserine (PS)]. The majority of mitochondrial PE is made in mitochondria by the decarboxylation of PS which requires the import of PS into mitochondria from the ER. We have previously demonstrated that targeted disruption of the PS decarboxylase gene in mice results in embryonic lethal-
Abstracts / Chemistry and Physics of Lipids 163S (2010) S1–S18
work of fatty acid flux and its metabolic and regulatory involvement in lipid and membrane homeostasis.
Acknowledgements Supported by FWF projects 18857 and DK Molecular Enzymology W901-305. doi:10.1016/j.chemphyslip.2010.05.018 SO7 The targeting of plasmalemmal ceramide to mitochondria during apoptosis E.B. Babiychuk ∗ , K. Monastyrskaya, A. Draeger Department of Cell Biology, Institute of Anatomy, University of Bern, 3012 Bern, Switzerland Ceramide is a key lipid mediator in cellular processes such as differentiation, proliferation, growth arrest and apoptosis. Upon its genesis during apoptosis, ceramide self-associates into platforms thereby promoting extensive structural changes within the plasma membrane. In most cells, apoptosis is initiated by the release of mitochondrial proteins into the cytoplasm and there is abundant evidence to suggest that an increase in permeability is caused by the appearance of pores in the outer mitochondrial membrane. Recent data suggest that generation of intracellular ceramide increases mitochondrial permeability. Whereas mitochondria are central organelles in the ceramide-dependent induction of apoptosis, they are not held to participate in ceramide production themselves. Since intracellular ceramide transport is hampered by the insoluble nature of this molecule, it is an open question how ceramide transfer into mitochondria is effected. We have previously established annexin A1, a member of a family of Ca2+ and membrane-binding proteins, as a marker for ceramide platforms. Using fluorescently tagged annexin A1 we have shown that, upon its generation within the plasma membrane, ceramide selfassociated into platforms, which subsequently invaginated and fused with mitochondria. Electron microscopic tomography confirmed that upon ceramide platform formation, the invaginated regions of the plasma membrane extend deeply into the cytoplasm forming physical contacts with mitochondrial outer membranes. Thus, ceramide is transferred from the plasma membrane to the mitochondrial outer membrane. It is conceivable that this “Kissof-Death” increases the permeability of the outer mitochondrial membrane and triggers apoptosis.
ity and profound abnormalities in mitochondrial morphology. We have now tested the hypothesis that reduction in mitochondrial PE production via the decarboxylation of PS impairs mitochondrial function. We used three models of genetically modified Chinese hamster ovary (CHO) cells in which: (i) PS synthetic capacity was only 5% of normal, (ii) PS import into mitochondria was blocked, or (iii) PS decarboxylase mRNA was reduced by >85% by RNA silencing. In each of these models, cell viability was not impaired but the PE content of mitochondria was reduced by ∼40%, mitochondrial morphology and motility were markedly altered, cellular ATP levels were reduced, and mitochondrial membrane potential was greatly increased. These data demonstrate for the first time that the mitochondrial content of PE profoundly influences mitochondrial function. doi:10.1016/j.chemphyslip.2010.05.020 Session 3: Bioactive lipids and lipidomics PL10 Control of free arachidonic acid levels within immunoinflammatory cells by phospholipases A2 and acyltransferases Jesús Balsinde Institute of Molecular Biology and Genetics, Spanish National Research Council (CSIC), 47003 Valladolid, Spain Arachidonic acid (AA) and its oxygenated derivatives, the eicosanoids, mediate a variety of physiological and pathophysiological states. The distribution of AA among cellular glycerophospholipids is finely regulated by the CoA-dependent acylation of lysophospholipids followed by transacylation reactions. Under the appropriate stimulatory conditions, AA is liberated from its phospholipid storage sites by the action of one or various phospholipases A2 . Thus, cellular availability of AA depends on an exquisite balance between phospholipid reacylation and hydrolysis reactions (Pérez-Chacón et al., 2009; Balboa and Balsinde, 2006). Phagocytic cells mobilize large amounts of free AA during stimulation with stimuli of the innate immune response (Balsinde et al., 2000; Pérez-Chacón et al., 2010). Utilizing HPLC coupled to electrospray ionization mass spectrometry, we have characterized changes in AA-containing phospholipid species in human monocytes stimulated with zymosan (Balgoma et al., 2010). We have identified three AA-containing phospholipids, namely PI(20:4/20:4), PC(20:4/20:4) and PE(16:1/20:4) in stimulated cells which are not present in resting cells or are found at very low levels. Thus these species can be regarded as lipid markers of human monocyte activation.
doi:10.1016/j.chemphyslip.2010.05.019 SO8
References
Daniel H. Bai, Jean E. Vance ∗
Balboa, M.A., Balsinde, J., 2006. Biochim. Biophys. Acta 1761, 385–391. Balgoma, et al., 2010. J. Immunol. 184, 3857–3865. Balsinde, J., Balboa, M.A., Dennis, E.A., 2000. J. Biol. Chem. 275, 22544–22549. Pérez-Chacón, et al., 2009. Biochim. Biophys. Acta 1791, 1103–1113. Pérez-Chacón, et al., 2010. J. Immunol. 184, 1071–1078.
Group on Molecular and Cell Biology of Lipids and Department of Medicine, University of Alberta, Edmonton, AB, Canada
doi:10.1016/j.chemphyslip.2010.05.021
Genetic reduction of mitochondrial phosphatidylethanolamine in mammalian cells impairs mitochondrial function
In mammalian cells, phosphatidylethanolamine (PE) is made by two spatially distinct pathways: in the endoplasmic reticulum (ER) [from the CDP-ethanolamine pathway] and in mitochondria [from the decarboxylation of phosphatidylserine (PS)]. The majority of mitochondrial PE is made in mitochondria by the decarboxylation of PS which requires the import of PS into mitochondria from the ER. We have previously demonstrated that targeted disruption of the PS decarboxylase gene in mice results in embryonic lethal-