Publicação
Characterization of the role of sphingolipids in the modulation of acetic acid-induced apoptosis
| Resumo: | The yeast Saccharomyces cerevisiae can undergo programmed cell death in response to different stimuli. Exposure of yeast cells to acetic acid has been shown to trigger a mitochondrial pathway displaying, as in mammalian cells, typical apoptotic markers such as externalization of phosphatidylserine, DNA fragmentation, chromatin condensation, mitochondrial dysfunction with cytochrome c release and production of reactive oxygen species (ROS). Sphingolipids are lipid second messengers generated in response to different physiological signals and stress stimuli. They affect multiple aspects of cellular function, including apoptosis. Changes in sphingolipid metabolism have been linked to apoptosis and oxidative stress in both yeast and mammalian cells. The increase of ceramide and sphingosine levels leads to cell growth arrest and apoptosis whereas the increase of sphingosine-1-phosphate levels promotes proliferation and inhibits apoptosis. Moreover, ceramides have been detected in mitochondria and accumulate upon stress treatments, increasing the permeability of the mitochondria to cytochrome c and leading to the generation of ROS. Our working hypothesis was that acetic acid may elicit ceramide production and, therefore, may trigger apoptosis by a signal transduction pathway modulated by ceramide. For that reason, we aimed to characterize the relative contribution of biosynthesis versus catabolism of ceramides to the apoptotic cell death induced by acetic acid in yeast. For our studies, yeast cells lacking Lag1p, Lac1p (unable to generate ceramide by de novo synthesis), Ydc1p and Ypc1p (unable to breakdown ceramide) and Isc1p (unable to generate ceramide by degradation of inositolphosphosphingolipids), were generated by homologous recombination. Our results showed that lag1Δ and isc1Δ mutant cells exhibited a higher resistance to acetic acid that was correlated with lower levels of ROS production and reduced mitochondrial alterations. In comparison with the wild-type strain, lag1Δ and isc1Δ mutant cells display, under acetic acid stress, lower levels of mitochondrial fragmentation and degradation, and reduced alterations of the mitochondrial membrane potential. Associated with these events, there was also less translocation of cytochrome c to the cytosol in response to acetic acid than in the wild-type strain. In conclusion, our results suggest that ceramide production contributes to cell death induced by acetic acid, especially through the hydrolysis of complex lipids catalyzed by Isc1p and through de novo synthesis catalyzed by Lag1p. |
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| Autores principais: | Rego, António Miguel Araújo |
| Ano: | 2011 |
| País: | Portugal |
| Tipo de documento: | dissertação de mestrado |
| Tipo de acesso: | acesso aberto |
| Instituição associada: | Universidade do Minho |
| Idioma: | português |
| Origem: | RepositóriUM - Universidade do Minho |
| Resumo: | The yeast Saccharomyces cerevisiae can undergo programmed cell death in response to different stimuli. Exposure of yeast cells to acetic acid has been shown to trigger a mitochondrial pathway displaying, as in mammalian cells, typical apoptotic markers such as externalization of phosphatidylserine, DNA fragmentation, chromatin condensation, mitochondrial dysfunction with cytochrome c release and production of reactive oxygen species (ROS). Sphingolipids are lipid second messengers generated in response to different physiological signals and stress stimuli. They affect multiple aspects of cellular function, including apoptosis. Changes in sphingolipid metabolism have been linked to apoptosis and oxidative stress in both yeast and mammalian cells. The increase of ceramide and sphingosine levels leads to cell growth arrest and apoptosis whereas the increase of sphingosine-1-phosphate levels promotes proliferation and inhibits apoptosis. Moreover, ceramides have been detected in mitochondria and accumulate upon stress treatments, increasing the permeability of the mitochondria to cytochrome c and leading to the generation of ROS. Our working hypothesis was that acetic acid may elicit ceramide production and, therefore, may trigger apoptosis by a signal transduction pathway modulated by ceramide. For that reason, we aimed to characterize the relative contribution of biosynthesis versus catabolism of ceramides to the apoptotic cell death induced by acetic acid in yeast. For our studies, yeast cells lacking Lag1p, Lac1p (unable to generate ceramide by de novo synthesis), Ydc1p and Ypc1p (unable to breakdown ceramide) and Isc1p (unable to generate ceramide by degradation of inositolphosphosphingolipids), were generated by homologous recombination. Our results showed that lag1Δ and isc1Δ mutant cells exhibited a higher resistance to acetic acid that was correlated with lower levels of ROS production and reduced mitochondrial alterations. In comparison with the wild-type strain, lag1Δ and isc1Δ mutant cells display, under acetic acid stress, lower levels of mitochondrial fragmentation and degradation, and reduced alterations of the mitochondrial membrane potential. Associated with these events, there was also less translocation of cytochrome c to the cytosol in response to acetic acid than in the wild-type strain. In conclusion, our results suggest that ceramide production contributes to cell death induced by acetic acid, especially through the hydrolysis of complex lipids catalyzed by Isc1p and through de novo synthesis catalyzed by Lag1p. |
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