Programmed cell death in Saccharomyces cerevisiae is hampered by the deletion of GUP1 gene
© Tulha et al.; licensee BioMed Central Ltd. 2012
Received: 23 December 2011
Accepted: 27 April 2012
Published: 22 May 2012
During the past years, yeast has been successfully established as a model to study mechanisms of programmed cell death regulation. Saccharomyces cerevisiae commits to cell death showing typical hallmarks of metazoan apoptosis, in response to different stimuli. Gup1p, an O-acyltransferase, is required for several cellular processes that are related to apoptosis development, such as rafts integrity and stability, lipid metabolism including GPI anchor correct remodeling, proper mitochondrial and vacuole function, bud site selection and actin dynamics. Therefore, we hypothesize that apoptotic process would be affected by GUP1 deletion.
In the present work we used two known apoptosis inducing conditions, chronological aging and acetic acid, to assess several apoptotic markers in gup1∆ mutant strain. We found that this mutant presents a significantly reduced chronological lifespan as compared to Wt and it is also highly sensitive to acetic acid treatment. In addition, it presents extremely high levels of ROS. There were notorious differences on apoptotic markers between Wt and gup1∆ mutant strains, namely on the maintenance of plasma membrane integrity, on the phosphatidylserine externalization, on the depolarization of mitochondrial membrane and on the chromatin condensation. Those suggested that the mutant, under either condition, probably dies of necrosis and not from apoptosis.
To Gup1p has been assigned an important function on lipid rafts assembly/integrity, lipid metabolism and GPI anchor remodeling. Our results provide, for the first time, the connection of the integrity of yeast lipid rafts and apoptosis induction and/or signaling, giving new insights into the molecular mechanisms underlying this process in yeast.
Apoptosis is the most common process of programmed cell death (PCD) in eukaryotes. It is vital for the fast elimination of useless or injured cells, and for the differential development of tissues and organs. In humans the malfunction of this process leads to severe diseases, namely neurodegenerative disorders, AIDS and cancer. The existence of PCD processes in lower eukaryotes or bacteria was for long disregarded due to the absence of obvious benefits for unicellular organisms. Nonetheless, numerous works contributed to evidence PCD occurring in single cell organisms [1–4], as well as to the establishment of yeast as a good model to study mechanisms of apoptotic regulation [5, 6]. Multicellular aggregates of microbial cells, like colonies or biofilms, are spatially organized and require the specialization of cells differentially localized to ensure supply of nutrients and water to the whole cell ensemble . The growing concept that microbial multicellular aggregates form functional and higher organized structures, as a kind of proto-tissue, supports the notion that PCD may be a much more spread and conserved mechanism of cellular altruistic behaviour.
The characteristic apoptotic markers, as DNA fragmentation, phosphatidylserine externalization, chromatin condensation, release of cytochrome C, and/or caspases activation are also valid to assess apoptotic yeast cells [1, 8]. Furthermore, an increasing list of homologues of apoptotic regulators in metazoans has been identified in yeast, such as Yca1p, the proposed yeast caspase ; Aifp, the apoptosis inducing factor ; EndoG, an endonuclease which regulates not only life but also death in yeast ; Nma111p, a yeast HtrA-like protein ; Bir1p, an inhibitor-of-apoptosis protein  and Ybh3p, a yeast protein that interacts with Bcl-xL and harbours a functional BH3 domain . Additionally, the expression in S. cerevisiae of the mammalian Bcl-2 family and PKC isoforms , led to the same phenotypes observed in mammalian cells, providing evidence that apoptosis is an evolutionarily conserved mechanism. Several agents can induce yeast PCD, like hydrogen peroxide, UV radiation, the absence of nutrients, hyper-osmotic stress, acetic acid  and aging . Aging in yeast can be studied assessing either replicative or chronological lifespan. Replicative lifespan is defined as the number of daughter cells a single yeast mother cell produces before senescence; chronological lifespan is defined by the length of time cells can survive in a non-dividing, quiescence-like state . Chronological aged yeast cells also exhibit typical apoptotic markers. During chronological aging, the old yeasts die and release certain substances (nutrients) into the medium in order to promote survival of other aged cells, yet fitter ones .
On the other hand, it has been demonstrated that apoptotic S. cerevisiae cells display changes in the expression of some genes associated with the sphingolipids metabolism , which is consistent with changes in the proportions of the various sphingolipid types in dying cells . Carmona-Guitierrez and co-authors  observed the apoptosis induction by external addition of C2-ceramide, whereas Barbosa and co- authors reported changes in sphingolipids during chronological aging, namely a decrease of dihydrosphingosine levels and an increase of dihydro-C(26) -ceramide and phyto-C(26) -ceramide levels . Also, a role in apoptosis and aging of Ydc1p ceramidase was described , and a yeast homologue of mammalian neutral sphingomyelinase 2 was associated with apoptosis . Moreover, some intermediates in sphingolipids biosynthesis act as signalling molecules and growth regulators [22, 23]. Nevertheless, modest attention has been paid to the involvement of sphingolipids in yeast PCD.
In S. cerevisiae, sphingolipids are mainly located in the plasma membrane, being more concentrated along the sphingolipid-sterol rich domains , commonly named rafts. These domains play fundamental roles in connecting the plasma membrane to the cytoskeleton, ER and Golgi, and therefore in the correct protein sorting and trafficking through exocytosis/endocytosis . Moreover, rafts harbour signalling molecules besides sphingolipids, like kinases, PI2P (phosphatidylinositol-3,4-diphosphate), and GPI (glycosylphosphatidylinositol)-anchored proteins [25, 26]. The latter, are proteins attached to the plasma membrane via a lipid anchor that contains either a ceramide or diacylglycerol . Gup1p is a membrane-bound O-acyltransferase [28, 29] involved in lipid metabolism, rafts integrity and assembly  and GPI anchor remodelling . This protein was primarily identified associated with phenotypes on glycerol metabolism and transport , but has further been implicated in a vast number of distinct processes, namely cell wall structure, composition and biogenesis , plasma membrane assembly and composition [30, 34], cytoskeleton polarization and bud site selection , and telomere length , all of which directly or indirectly associated with apoptosis. This work presents evidence that cells lacking GUP1 are not able of undergoing apoptosis, as revealed by the analysis of several apoptotic markers (mainly lack of membrane integrity and of phosphatidylserine externalization). Instead the mutant appears to be experiencing a necrotic cell death process, upon both chronological aging and acetic acid induction. This result adds to the growing view that as in higher eukaryotes, lipids are involved in signalling PCD in yeast.
GUP1 is involved in a wide range of cellular processes, some of which are associated directly or indirectly with apoptosis, such as rafts integrity and lipids metabolism [17, 18, 21, 30, 31, 34], cytoskeleton polarization [35, 37], and telomere length [36, 38]. In the present work, we assess apoptotic markers for gup1∆ mutant strain and compare them with Wt, under two different conditions documented to induce apoptosis in yeast: chronological aging and acetic acid [8, 39].
gup1∆ mutant cells exhibit a reduction in chronological lifespan
Yeast chronological lifespan is described as the length of time a population remains viable in the non-dividing/stationary phase [40, 41]. Chronologically aged yeast cells die exhibiting specific markers of apoptosis [6, 40]. We checked the survival of gup1∆ chronologically aged cells in comparison to Wt, continuously for 30 days throughout stationary phase until complete death of the culture.
Chronological aged gup1∆ mutant seems to be incapable of dying by apoptosis but rather by necrosis
In order to investigate whether chronologically aged Wt and gup1∆ mutant cells die by apoptosis, we analyzed several apoptotic markers in exponentially growing and chronologically aged cultures on both strains [6, 42]. We choose 6 hours growth to assess exponential cells, and day 7 or day 12 to test chronologically aged cells of gup1∆ mutant and Wt, respectively.
Phosphatidylserine has an asymmetric distribution in the lipid bilayer of the cytoplasmic membrane . The exposure of phosphatidylserine at the outer surface of the cytoplasmic membrane occurs at the early stages of apoptosis , when membrane integrity is still retained. We checked this through the FITC-coupled Annexin V reaction followed by flow cytometry of co-labeled Annexin V/PI cells. We observed that gup1∆ mutant aging cells presents a significant percentage (53%) of necrotic cells (Ann (−)/PI(+)). In contrast, in Wt cells the exposure of phosphatidylserine (Ann (+)/PI (−)) increased in aged cells (less than 1% to ~12%) (Figure 2B).
In order to evaluate the mitochondrial membrane depolarization, DiOC6 was used. At a concentration of 20 ng/ml this dye accumulates specifically at mitochondrial membranes and can be observed by fluorescence microscopy. Nonetheless, cells that have low mitochondrial membrane potential will fail to accumulate DiOC6. Both gup1∆ mutant and Wt exponential cells stained with DiOC6 revealed intact mitochondrial networks, confirming a normal polarization of mitochondrial membranes (Figure 2C left panels). Aged cells (7 and 12 days in gup1∆ mutant and Wt, respectively), showed a decrease in green fluorescence of approximately 40% in Wt and 50% in gup1∆ mutant, reflecting a reduction in mitochondrial membrane potential (Figure 2C right panels). Moreover, some cells exhibited a strong green fluorescence all over the cell, mainly in gup1∆ mutant strain, suggesting that these cells possibly had the plasma membrane altered, which in turn resulted in the accumulation of DiOC6 on the cytosol (Figure 2C right panels).
Finally, we evaluated chromatin condensation through DAPI staining (Figure 2D). Moderate chromatin condensation upon DAPI staining was observed in 80% of old gup1∆ mutant cells, which can be visualized by the fluorescent semicircles formed by the chromatin fragments (Figure 2D right panels). Regarding Wt aged cells, we observed some cells with chromatin condensation, but we also detected cells without stained nucleus or even with multiples nucleus (Figure 2D right panels). These are probably due to an endomitosis process [45, 46]. In contrast, in exponentially growing cultures, both Wt and gup1∆ mutant cells presented integral chromatin mirrored as single round fluorescent circles in the middle of the cell (Figure 2D left panels).
gup1∆ mutant cells are sensitive to acetic acid
Acetic acid induces cell death by necrosis similar to that triggered by chronological aging in the gup1∆ mutant strain
Yeast mitochondria undergo both structural and functional changes after the incubation with acetic acid , including mitochondrial membrane depolarization. In order to evaluate this phenomenon, DiOC6 staining was used to visualize mitochondrial membranes (Figure 4C). Just before apoptosis induction with acetic acid, most of Wt and gup1∆ mutant cells presented intact mitochondrial networks (Figure 4C left panels). After the treatment, it was possible to visualize depolarization of mitochondrial membranes in approximately 40% and 30% of gup1∆ mutant and Wt cells, respectively, mirrored by the absence of fluorescence (Figure 4C right panels). Furthermore, we observed a considerable number of gup1∆ mutant cells displayed an increase in DiOC6 green fluorescence, similarly to the results obtained when the apoptotic inductor was chronological aging (Figure 4C right panels).
Additionally, we checked for chromatin condensation during acetic acid treatment by staining cells with DAPI (Figure 4D). Nearly no chromatic condensation was observed in both gup1∆ mutant and Wt untreated cells, as reflected by the single round fluorescent circles in the center of the cells (Figure 4D left panels). Yet, after the treatment with acetic acid, we observed a significant increase in gup1∆ mutant cells exhibiting moderate chromatin condensation along the nuclear envelope (~90%). In Wt, ~25% of cells presented chromatin condensation (Figure 4D right panels).
gup1∆ mutant cells accumulate large amounts of ROS during chronological aging and acetic acid treatment
The finding of an endogenous PCD process with an apoptotic phenotype has turned yeast into a powerful model for apoptosis research [39, 51, 52]. In fact, S. cerevisiae commits to cell death showing typical features of mammalian apoptosis, in response to different stimuli. However, how cell compounds participate in the processes leading to cell death in yeast remains to be established. Gup1p, an O-acyltransferase, is required for several cellular processes that are related to apoptosis development, namely, rafts integrity and stability, lipid metabolism including GPI anchor correct remodeling, proper mitochondrial and vacuole function, and actin dynamics [30, 31, 33, 35, 37, 42, 53–56].
In this work we used two known apoptosis-inducing conditions, chronological aging  and acetic acid , to assess several apoptotic markers in gup1∆ mutant strain. We found that, when compared with Wt, gup1∆ mutant presents a significant reduced chronological lifespan, showing almost no viability after 11 days incubation. Chronologically aged yeast cultures were shown to die exhibiting typical apoptotic markers . Accordingly, we showed that chronologically aged Wt cells predominantly commit to an apoptotic death, as revealed by a) PI negative cells; b) phosphatidylserine externalization; c) depolarization of mitochondrial membrane; and d) chromatin condensation. Yet, gup1∆ mutant aged cells seem to be incapable of undergoing apoptosis. Instead, these cells appeared to be experiencing a necrotic cell death process. The gup1∆ mutant aged culture exhibited a higher number of cells with loss of membrane integrity, and did not reveal an increase of phosphatidylserine exposure on the surface of the plasma membrane. Such observations discredit the possibility that these cells are dying through an apoptotic process, being more likely that the reduction in lifespan is due to a necrotic death. Furthermore, both loss of mitochondrial membrane potential and moderate chromatin condensation that we observed in this mutant have already been described in necrotic phenotypes [57, 58]. Lately, several points of evidence suggest that necrotic cell death also occurs in yeast. Moreover, that can occur under normal physiological conditions or in the presence of cell death inducing substances, and not necessarily resulting from brutal chemical or physical damage, as previously thought .
We also used acetic acid as an apoptotic inducer of cell death in both Wt and gup1∆ mutant strains. Our results revealed that acetic acid induces a cell death process similar to that observed in aging cultures. These results are in accordance with the hypothesis proposed in a previous work, in which the toxicity of acetic acid produced during aging was suggested as the major cause of chronological aging in yeast . Reinforcing such idea are the acidified cultures that we observed during aging, probably resulting from acetic acid production and release to the medium (data not shown). Moreover, it was also reported that the signaling of acetic acid-induced apoptosis is linked to amino-acid metabolism as well as to the TOR pathway , as it happens in the aging process . A necrotic death induced by acetic acid was already observed in other yeast mutants, namely in mutants in class C VPS genes that code for proteins essential for vacuolar and endossomal vesicle function .
Accumulation of ROS has predominantly been associated to yeast apoptosis under numerous conditions [62–64]. Some studies have addressed a fundamental role of ROS on the execution apoptotic death, after treatment with low doses of hydrogen peroxide  and on the superoxide-mediated altruistic program of aging . Interestingly, however, many studies have suggested a crucial involvement of ROS during necrosis of mammalian cells  as well as in yeast necrosis [42, 64]. This evidence is in accordance with our results. We observed a significant difference in ROS accumulation between Wt and gup1∆ mutant strain in both chronological aging and acetic acid treatment. gup1∆ mutant cells, which present a necrotic phenotype, have an extremely higher accumulation of ROS. If ROS can contribute, apart from its role on apoptosis, to the necrotic cell death in yeast as well, or if it is rather a byproduct that accumulates as a result of cellular demise, remains to be elucidated.
Gup1p has been described to have an important function on lipid rafts assembly/integrity . In the literature, rafts have been increasingly implicated on regulation of apoptotic signaling in mammalian cells [54, 67]. In response to intra or extracellular stimuli, lipid rafts can include or exclude proteins to variable extents. This favors specific protein-protein interactions and modulates the activity of signalling apoptotic cascades. Moreover, in mammalian cells a number of proteins involved in apoptotic signals have been found to locate in lipid rafts, namely Fas/CD95 receptor  and the pro-apoptotic protein of Bcl-2 family, Bad . Our results showed that the PCD processes in S. cerevisiae is altered by GUP1 deletion and reinforce the importance of lipid rafts on the regulation of apoptotic signaling in yeast. Moreover, our findings point to that these membrane domains seem to be indispensable for a proper development of PCD, under aging and acetic acid conditions, namely in the switch from a necrotic to an apoptotic death phenotype.
We demonstrate that gup1∆ mutant strain present a significantly reduced chronological lifespan comparing to Wt. Moreover, this mutant showed to be highly sensitive to acetic acid. Yet, while chronologically aged and acetic acid treated Wt cells die exhibiting apoptotic markers, gup1∆ mutant cells under the same conditions seems to be incapable of undergoing apoptosis. Instead, these cells appeared to be experiencing a necrotic cell death process. In addition, those cells also present extremely high levels of ROS. Being gup1∆ mutant affected in lipid rafts integrity/assembly, lipid metabolism and GPI anchor remodeling we propose that the integrity of rafts may be essential for apoptosis induction and/or signaling. This provides for the first time the possible participation of lipid rafts in yeast apoptosis, giving new insights into the molecular mechanisms underlying this particular process of PCD, and highlighting the complex network of cellular structures that interact, cooperate and compete to regulate cell death.
Strains and growth conditions
The Saccharomyces cerevisiae strains used in this study were W303-1A  and BHY54 . Yeast batch cultures were grown aerobically in minimal medium (0.67% (wt/v) YNB (Difco)) with 2% (wt/v) glucose and adequate quantities of auxotrophic requirements . Incubation was performed at 30°C, 200 rpm, orbital shaking and air/liquid ratio 3/1. Yeast strains maintenance was done on rich medium (YPD (Difco) with 2% agar), grown at 30°C for 48 h and kept at 4°C up to 5 days.
For chronological lifespan experiments, pre-inoculum cultures grown overnight on YNB were used to start batch cultures at 0.05 (OD600nm) in fresh YNB medium. At the stipulated time points, culture aliquots were taken to assess growth through OD600 and colony forming units (c.f.u.), and for apoptotic assays. c.f.u. were determined plating 1,000 cells, counted on a Neubauer chamber, on YPD agar, as previously described . Colonies were counted after 48 h incubation at 30°C. No further colonies appeared after that incubation period.
Sensitivity to acetic acid
Dropout tests were performed from mid-exponential YNB cultures containing approximately 1 × 106 cells/ml. Ten-fold serial dilutions were made, and 5 μl of each suspension was applied on YNB medium supplemented with different acetic acid concentrations (50, 80 and 100 mM). Results were scored after 48 h incubation at 30°C.
Acetic acid treatment
Yeast strains were grown until exponential phase (OD600 = 0.5–0.6) on YNB medium. Then the cultures were collected and resuspended to a final concentration of 107 cells ml-1 in fresh YNB adjusted to pH 3.0 with HCl and containing 160 mM acetic acid. Incubation took place for 180 min at 30°C as previously described [4, 72]. At determined time points, 40 μl from a 10−4 cell suspension were inoculated onto YPD agar plates and c.f.u. were counted after 48 h incubation at 30°C. The percentage of viable cells was estimated considering 100% survival the number of c.f.u. obtained in time 0.
Membrane integrity was assessed by PI (Propidium Iodide) staining. Cells were harvested, washed and resuspended in PBS (137 mM NaCl; 2.7 mM KCl; 100 mM Na2HPO4; 2 mM KH2PO4; pH 7.4) containing PI (4 μg/ml) (Sigma). The samples were incubated for 10 min at room temperature in the dark and analyzed in an Epics® XL™ (Beckman Coulter) flow cytometer. At least 20,000 cells from each sample were analyzed.
Phosphatidylserine exposure was detected by an FITC-coupled Annexin V reaction with the ApoAlertAnnexin V Apoptosis Kit (CLONTECH Laboratories). For that, cells were primarily harvested and washed in digesting buffer (1.2 M sorbitol; 0.5 mM MgCl2; 35 mM K2HPO4; pH 6.8). To promote the drug course through cell wall, an incubation step with Zymolyase (20 T) at 30°C was performed. Phase-contrast microscopy was used to monitor that step, preventing this way damage to the unfixed spheroplasts. Cells were subsequently centrifuged (10 min at 1500 rpm) and resuspended in 200 μl of binding buffer (1.2 M sorbitol; 10 mM HEPES/NaOH, pH 7.4; 140 mM NaCl; 2.5 mM Cacl2). To 40 μl of this cell suspension, 2 μl Annexin V (1 μg/ml) and 1 μl PI (4 μg/ml) were added, and the mixture incubated for 20 min at room temperature in the dark. Finally, extra 400 μl of binding buffer were added to the mixture just prior to analysis in an Epics® XL™ (Beckman Coulter) flow cytometer. At least 20,000 cells from each sample were analyzed.
For evaluation of mitochondrial potential the probe DiOC6 (3,3′dihexyloxacarbocyanine iodide) (Invitrogen) was used. Cells were harvested, washed, and resuspended in DiOC6 buffer (10 mM MES; 0.1 mM MgCl2; 2% (wt/v) glucose, adjusted to pH 6 set with Ca(OH)2) containing DiOC6 (20 ng/ml). Cells were visualized by light microscopy (LM) after 30 min at room temperature in the dark. At least, 300 cells selected randomly were counted per sample. The number of cells counted with mitochondrial depolarization (cells without fluorescence) was indexed to our 100% (300 cells).
Chromatin condensation was assessed by DAPI (4,6-diamino-2-phenylindole dihydrochloride) (Sigma) staining. Cells were harvested, washed, fixed for 45 min with 3.7% formaldehyde, permeabilized with a solution of 70% (v/v) ethanol for 30 min, sonicated for 5 sec and afterwards stained with DAPI (1 μg/ml). Cells were visualized by LM after 5 min at room temperature in the dark. At least 300 cells selected randomly were counted per sample. The number of cells counted with chromatin condensation was indexed to our 100% (300 cells).
Stained cells were visualized in a Leica Microsystems DM-5000B epifluorescence microscope with appropriate filter settings using a 100× oil-immersion objective. Images were acquired with a Leica DCF350FX digital camera and processed with LAS AF Leica Microsystems software.
Assessment of ROS
To visualize accumulation of ROS cells were harvested by centrifugation, resuspended in PBS in the presence of DHE (Dihydroethidium) (4 μg/ml), and further incubated in the dark for 30 min at room temperature. To quantify the number of cells displaying high ROS levels, at least 20,000 cells were counted in an Epics® XL™ (Beckman Coulter) flow cytometer.
This work was supported by FEDER funds through the COMPETE (Programa Operacional Factores de Competitividade) and national funds through FCT (Fundação para a Ciência e a Tecnologia) through FCOMP-01-0124-FEDER-07047. Fábio Faria-Oliveira is a PhD grantee from FCT (SFRH/BD/45368/2008). Authors would like to acknowledge Manuela Côrte-Real for profitable discussions of the results and to Rui Silva for assistance on cytometry experiments. We also thank Hugh S. Johnson for the critical reading of the manuscript regarding English usage.
- Madeo F, Frohlich E, Frohlich KU: A yeast mutant showing diagnostic markers of early and late apoptosis. J Cell Biol. 1997, 139 (3): 729-734. 10.1083/jcb.139.3.729.PubMedPubMed CentralView ArticleGoogle Scholar
- Ligr M, Madeo F, Frohlich E, Hilt W, Frohlich KU, Wolf DH: Mammalian Bax triggers apoptotic changes in yeast. FEBS Lett. 1998, 438 (1–2): 61-65.PubMedView ArticleGoogle Scholar
- Madeo F, Frohlich E, Ligr M, Grey M, Sigrist SJ, Wolf DH, Frohlich KU: Oxygen stress: a regulator of apoptosis in yeast. J Cell Biol. 1999, 145 (4): 757-767. 10.1083/jcb.145.4.757.PubMedPubMed CentralView ArticleGoogle Scholar
- Ludovico P, Sousa MJ, Silva MT, Leao C, Corte-Real M: Saccharomyces cerevisiae commits to a programmed cell death process in response to acetic acid. Microbiology. 2001, 147 (Pt 9): 2409-2415.PubMedView ArticleGoogle Scholar
- Frohlich KU, Fussi H, Ruckenstuhl C: Yeast apoptosis–from genes to pathways. Semin Cancer Biol. 2007, 17 (2): 112-121. 10.1016/j.semcancer.2006.11.006.PubMedView ArticleGoogle Scholar
- Herker E, Jungwirth H, Lehmann KA, Maldener C, Frohlich KU, Wissing S, Buttner S, Fehr M, Sigrist S, Madeo F: Chronological aging leads to apoptosis in yeast. J Cell Biol. 2004, 164 (4): 501-507. 10.1083/jcb.200310014.PubMedPubMed CentralView ArticleGoogle Scholar
- Vachova L, Palkova Z: Physiological regulation of yeast cell death in multicellular colonies is triggered by ammonia. J Cell Biol. 2005, 169 (5): 711-717. 10.1083/jcb.200410064.PubMedPubMed CentralView ArticleGoogle Scholar
- Madeo F, Herker E, Wissing S, Jungwirth H, Eisenberg T, Frohlich KU: Apoptosis in yeast. Curr Opin Microbiol. 2004, 7 (6): 655-660. 10.1016/j.mib.2004.10.012.PubMedView ArticleGoogle Scholar
- Madeo F, Herker E, Maldener C, Wissing S, Lachelt S, Herlan M, Fehr M, Lauber K, Sigrist SJ, Wesselborg S, et al.: A caspase-related protease regulates apoptosis in yeast. Mol Cell. 2002, 9 (4): 911-917. 10.1016/S1097-2765(02)00501-4.PubMedView ArticleGoogle Scholar
- Wissing S, Ludovico P, Herker E, Buttner S, Engelhardt SM, Decker T, Link A, Proksch A, Rodrigues F, Corte-Real M, et al.: An AIF orthologue regulates apoptosis in yeast. J Cell Biol. 2004, 166 (7): 969-974. 10.1083/jcb.200404138.PubMedPubMed CentralView ArticleGoogle Scholar
- Buttner S, Eisenberg T, Carmona-Gutierrez D, Ruli D, Knauer H, Ruckenstuhl C, Sigrist C, Wissing S, Kollroser M, Frohlich KU, et al.: Endonuclease G regulates budding yeast life and death. Mol Cell. 2007, 25 (2): 233-246. 10.1016/j.molcel.2006.12.021.PubMedView ArticleGoogle Scholar
- Fahrenkrog B, Sauder U, Aebi U: The S. cerevisiae HtrA-like protein Nma111p is a nuclear serine protease that mediates yeast apoptosis. J Cell Sci. 2004, 117 (Pt 1): 115-126.PubMedView ArticleGoogle Scholar
- Walter D, Wissing S, Madeo F, Fahrenkrog B: The inhibitor-of-apoptosis protein Bir1p protects against apoptosis in S. cerevisiae and is a substrate for the yeast homologue of Omi/HtrA2. J Cell Sci. 2006, 119 (Pt 9): 1843-1851.PubMedView ArticleGoogle Scholar
- Buttner S, Ruli D, Vogtle FN, Galluzzi L, Moitzi B, Eisenberg T, Kepp O, Habernig L, Carmona-Gutierrez D, Rockenfeller P, et al.: A yeast BH3-only protein mediates the mitochondrial pathway of apoptosis. EMBO J. 2011, 30 (14): 2779-2792. 10.1038/emboj.2011.197.PubMedPubMed CentralView ArticleGoogle Scholar
- Saraiva L, Silva RD, Pereira G, Goncalves J, Corte-Real M: Specific modulation of apoptosis and Bcl-xL phosphorylation in yeast by distinct mammalian protein kinase C isoforms. J Cell Sci. 2006, 119 (Pt 15): 3171-3181.PubMedView ArticleGoogle Scholar
- Kaeberlein M: Longevity and aging in the budding yeast. In: Handbook of models for human aging. Edited by: Conn PM. 2006, Academic, San DiegoGoogle Scholar
- Laun P, Ramachandran L, Jarolim S, Herker E, Liang P, Wang J, Weinberger M, Burhans DT, Suter B, Madeo F, et al.: A comparison of the aging and apoptotic transcriptome of Saccharomyces cerevisiae. FEMS Yeast Res. 2005, 5 (12): 1261-1272. 10.1016/j.femsyr.2005.07.006.PubMedView ArticleGoogle Scholar
- Aerts AM, Zabrocki P, Francois IE, Carmona-Gutierrez D, Govaert G, Mao C, Smets B, Madeo F, Winderickx J, Cammue BP, et al.: Ydc1p ceramidase triggers organelle fragmentation, apoptosis and accelerated ageing in yeast. Cell Mol Life Sci. 2008, 65 (12): 1933-1942. 10.1007/s00018-008-8129-8.PubMedView ArticleGoogle Scholar
- Carmona-Gutierrez D, Reisenbichler A, Heimbucher P, Bauer MA, Braun RJ, Ruckenstuhl C, Buttner S, Eisenberg T, Rockenfeller P, Frohlich KU, et al.: Ceramide triggers metacaspase-independent mitochondrial cell death in yeast. Cell Cycle. 2011, 10 (22): 3973-3978. 10.4161/cc.10.22.18212.PubMedView ArticleGoogle Scholar
- Barbosa AD, Osorio H, Sims KJ, Almeida T, Alves M, Bielawski J, Amorim MA, Moradas-Ferreira P, Hannun YA, Costa V: Role for Sit4p-dependent mitochondrial dysfunction in mediating the shortened chronological lifespan and oxidative stress sensitivity of Isc1p-deficient cells. Mol Microbiol. 2011, 81 (2): 515-527. 10.1111/j.1365-2958.2011.07714.x.PubMedPubMed CentralView ArticleGoogle Scholar
- Almeida T, Marques M, Mojzita D, Amorim MA, Silva RD, Almeida B, Rodrigues P, Ludovico P, Hohmann S, Moradas-Ferreira P, et al.: Isc1p plays a key role in hydrogen peroxide resistance and chronological lifespan through modulation of iron levels and apoptosis. Mol Biol Cell. 2008, 19 (3): 865-876.PubMedPubMed CentralView ArticleGoogle Scholar
- Ogretmen B, Hannun YA: Biologically active sphingolipids in cancer pathogenesis and treatment. Nat Rev Cancer. 2004, 4 (8): 604-616. 10.1038/nrc1411.PubMedView ArticleGoogle Scholar
- Thevissen K, Francois IE, Winderickx J, Pannecouque C, Cammue BP: Ceramide involvement in apoptosis and apoptotic diseases. Mini Rev Med Chem. 2006, 6 (6): 699-709. 10.2174/138955706777435643.PubMedView ArticleGoogle Scholar
- Jacobson K, Mouritsen OG, Anderson RG: Lipid rafts: at a crossroad between cell biology and physics. Nat Cell Biol. 2007, 9 (1): 7-14. 10.1038/ncb0107-7.PubMedView ArticleGoogle Scholar
- Bagnat M, Keranen S, Shevchenko A, Simons K: Lipid rafts function in biosynthetic delivery of proteins to the cell surface in yeast. Proc Natl Acad Sci U S A. 2000, 97 (7): 3254-3259. 10.1073/pnas.97.7.3254.PubMedPubMed CentralView ArticleGoogle Scholar
- Alvarez-Vasquez F, Sims KJ, Voit EO, Hannun YA: Coordination of the dynamics of yeast sphingolipid metabolism during the diauxic shift. Theor Biol Med Model. 2007, 4: 42-10.1186/1742-4682-4-42.PubMedPubMed CentralView ArticleGoogle Scholar
- Pittet M, Conzelmann A: Biosynthesis and function of GPI proteins in the yeast Saccharomyces cerevisiae. Biochim Biophys Acta. 2007, 1771 (3): 405-420. 10.1016/j.bbalip.2006.05.015.PubMedView ArticleGoogle Scholar
- Hofmann K: A superfamily of membrane-bound O-acyltransferases with implications for wnt signaling. Trends Biochem Sci. 2000, 25 (3): 111-112. 10.1016/S0968-0004(99)01539-X.PubMedView ArticleGoogle Scholar
- Neves L, Oliveira R, Lucas C: Yeast orthologues associated with glycerol transport and metabolism. FEMS Yeast Res. 2004, 5 (1): 51-62. 10.1016/j.femsyr.2004.06.012.PubMedView ArticleGoogle Scholar
- Ferreira C, Lucas C: The yeast O-acyltransferase Gup1p interferes in lipid metabolism with direct consequences on the sphingolipid-sterol-ordered domains integrity/assembly. Biochim Biophys Acta. 2008, 1778 (11): 2648-2653. 10.1016/j.bbamem.2008.08.011.PubMedView ArticleGoogle Scholar
- Bosson R, Jaquenoud M, Conzelmann A: GUP1 of Saccharomyces cerevisiae encodes an O-acyltransferase involved in remodeling of the GPI anchor. Mol Biol Cell. 2006, 17 (6): 2636-2645. 10.1091/mbc.E06-02-0104.PubMedPubMed CentralView ArticleGoogle Scholar
- Holst B, Lunde C, Lages F, Oliveira R, Lucas C, Kielland-Brandt MC: GUP1 and its close homologue GUP2, encoding multimembrane-spanning proteins involved in active glycerol uptake in Saccharomyces cerevisiae. Mol Microbiol. 2000, 37 (1): 108-124. 10.1046/j.1365-2958.2000.01968.x.PubMedView ArticleGoogle Scholar
- Ferreira C, Silva S, van Voorst F, Aguiar C, Kielland-Brandt MC, Brandt A, Lucas C: Absence of Gup1p in Saccharomyces cerevisiae results in defective cell wall composition, assembly, stability and morphology. FEMS Yeast Res. 2006, 6 (7): 1027-1038. 10.1111/j.1567-1364.2006.00110.x.PubMedView ArticleGoogle Scholar
- Oelkers P, Tinkelenberg A, Erdeniz N, Cromley D, Billheimer JT, Sturley SL: A lecithin cholesterol acyltransferase-like gene mediates diacylglycerol esterification in yeast. J Biol Chem. 2000, 275 (21): 15609-15612. 10.1074/jbc.C000144200.PubMedView ArticleGoogle Scholar
- Ni L, Snyder M: A genomic study of the bipolar bud site selection pattern in Saccharomyces cerevisiae. Mol Biol Cell. 2001, 12 (7): 2147-2170.PubMedPubMed CentralView ArticleGoogle Scholar
- Askree SH, Yehuda T, Smolikov S, Gurevich R, Hawk J, Coker C, Krauskopf A, Kupiec M, McEachern MJ: A genome-wide screen for Saccharomyces cerevisiae deletion mutants that affect telomere length. Proc Natl Acad Sci U S A. 2004, 101 (23): 8658-8663. 10.1073/pnas.0401263101.PubMedPubMed CentralView ArticleGoogle Scholar
- Gourlay CW, Ayscough KR: Identification of an upstream regulatory pathway controlling actin-mediated apoptosis in yeast. J Cell Sci. 2005, 118 (Pt 10): 2119-2132.PubMedView ArticleGoogle Scholar
- Qi H, Li TK, Kuo D, Nur EKA, Liu LF: Inactivation of Cdc13p triggers MEC1-dependent apoptotic signals in yeast. J Biol Chem. 2003, 278 (17): 15136-15141. 10.1074/jbc.M212808200.PubMedView ArticleGoogle Scholar
- Carmona-Gutierrez D, Jungwirth H, Eisenberg T, Madeo F: Cell cycle control of cell death in yeast. Cell Cycle. 2010, 9 (20): 4046-View ArticleGoogle Scholar
- Fabrizio P, Longo VD: The chronological life span of Saccharomyces cerevisiae. Aging Cell. 2003, 2 (2): 73-81. 10.1046/j.1474-9728.2003.00033.x.PubMedView ArticleGoogle Scholar
- Rockenfeller P, Madeo F: Apoptotic death of ageing yeast. Exp Gerontol. 2008, 43 (10): 876-881. 10.1016/j.exger.2008.08.044.PubMedView ArticleGoogle Scholar
- Schauer A, Knauer H, Ruckenstuhl C, Fussi H, Durchschlag M, Potocnik U, Frohlich KU: Vacuolar functions determine the mode of cell death. Biochim Biophys Acta. 2009, 1793 (3): 540-545. 10.1016/j.bbamcr.2008.11.006.PubMedView ArticleGoogle Scholar
- Cerbon J, Calderon V: Changes of the compositional asymmetry of phospholipids associated to the increment in the membrane surface potential. Biochim Biophys Acta. 1991, 1067 (2): 139-144. 10.1016/0005-2736(91)90035-7.PubMedView ArticleGoogle Scholar
- Martin SJ, Reutelingsperger CP, McGahon AJ, Rader JA, van Schie RC, LaFace DM, Green DR: Early redistribution of plasma membrane phosphatidylserine is a general feature of apoptosis regardless of the initiating stimulus: inhibition by overexpression of Bcl-2 and Abl. J Exp Med. 1995, 182 (5): 1545-1556. 10.1084/jem.182.5.1545.PubMedView ArticleGoogle Scholar
- Pichova A, Vondrakova D, Breitenbach M: Mutants in the Saccharomyces cerevisiae RAS2 gene influence life span, cytoskeleton, and regulation of mitosis. Can J Microbiol. 1997, 43 (8): 774-781. 10.1139/m97-111.PubMedView ArticleGoogle Scholar
- Laun P, Pichova A, Madeo F, Fuchs J, Ellinger A, Kohlwein S, Dawes I, Frohlich KU, Breitenbach M: Aged mother cells of Saccharomyces cerevisiae show markers of oxidative stress and apoptosis. Mol Microbiol. 2001, 39 (5): 1166-1173. 10.1111/j.1365-2958.2001.02317.x.PubMedView ArticleGoogle Scholar
- Ludovico P, Rodrigues F, Almeida A, Silva MT, Barrientos A, Corte-Real M: Cytochrome c release and mitochondria involvement in programmed cell death induced by acetic acid in Saccharomyces cerevisiae. Mol Biol Cell. 2002, 13 (8): 2598-2606. 10.1091/mbc.E01-12-0161.PubMedPubMed CentralView ArticleGoogle Scholar
- Longo VD, Gralla EB, Valentine JS: Superoxide dismutase activity is essential for stationary phase survival in Saccharomyces cerevisiae. Mitochondrial production of toxic oxygen species in vivo. J Biol Chem. 1996, 271 (21): 12275-12280. 10.1074/jbc.271.21.12275.PubMedView ArticleGoogle Scholar
- Nestelbacher R, Laun P, Vondrakova D, Pichova A, Schuller C, Breitenbach M: The influence of oxygen toxicity on yeast mother cell-specific aging. Exp Gerontol. 2000, 35 (1): 63-70. 10.1016/S0531-5565(99)00087-X.PubMedView ArticleGoogle Scholar
- Buttner S, Eisenberg T, Herker E, Carmona-Gutierrez D, Kroemer G, Madeo F: Why yeast cells can undergo apoptosis: death in times of peace, love, and war. J Cell Biol. 2006, 175 (4): 521-525. 10.1083/jcb.200608098.PubMedPubMed CentralView ArticleGoogle Scholar
- Fleury C, Pampin M, Tarze A, Mignotte B: Yeast as a model to study apoptosis?. Biosci Rep. 2002, 22 (1): 59-79. 10.1023/A:1016013123094.PubMedView ArticleGoogle Scholar
- Madeo F, Engelhardt S, Herker E, Lehmann N, Maldener C, Proksch A, Wissing S, Frohlich KU: Apoptosis in yeast: a new model system with applications in cell biology and medicine. Curr Genet. 2002, 41 (4): 208-216. 10.1007/s00294-002-0310-2.PubMedView ArticleGoogle Scholar
- Dickson RC, Lester RL: Sphingolipid functions in Saccharomyces cerevisiae. Biochim Biophys Acta. 2002, 1583 (1): 13-25. 10.1016/S1388-1981(02)00210-X.PubMedView ArticleGoogle Scholar
- Garcia A, Cayla X, Fleischer A, Guergnon J, Alvarez-Franco Canas F, Rebollo MP, Roncal F, Rebollo A: Rafts: a simple way to control apoptosis by subcellular redistribution. Biochimie. 2003, 85 (8): 727-731. 10.1016/j.biochi.2003.09.005.PubMedView ArticleGoogle Scholar
- Pereira C, Silva RD, Saraiva L, Johansson B, Sousa MJ, Corte-Real M: Mitochondria-dependent apoptosis in yeast. Biochim Biophys Acta. 2008, 1783 (7): 1286-1302. 10.1016/j.bbamcr.2008.03.010.PubMedView ArticleGoogle Scholar
- dos Santos SC, Sa-Correia I: Genome-wide identification of genes required for yeast growth under imatinib stress: vacuolar H + −ATPase function is an important target of this anticancer drug. OMICS. 2009, 13 (3): 185-198. 10.1089/omi.2008.0086.PubMedView ArticleGoogle Scholar
- Galluzzi L, Maiuri MC, Vitale I, Zischka H, Castedo M, Zitvogel L, Kroemer G: Cell death modalities: classification and pathophysiological implications. Cell Death Differ. 2007, 14 (7): 1237-1243. 10.1038/sj.cdd.4402148.PubMedView ArticleGoogle Scholar
- Sripriya P, Vedantam LV, Podile AR: Involvement of mitochondria and metacaspase elevation in harpin Pss-induced cell death of Saccharomyces cerevisiae. J Cell Biochem. 2009, 107 (6): 1150-1159. 10.1002/jcb.22217.PubMedView ArticleGoogle Scholar
- Burtner CR, Murakami CJ, Kennedy BK, Kaeberlein M: A molecular mechanism of chronological aging in yeast. Cell Cycle. 2009, 8 (8): 1256-1270. 10.4161/cc.8.8.8287.PubMedPubMed CentralView ArticleGoogle Scholar
- Almeida B, Ohlmeier S, Almeida AJ, Madeo F, Leao C, Rodrigues F, Ludovico P: Yeast protein expression profile during acetic acid-induced apoptosis indicates causal involvement of the TOR pathway. Proteomics. 2009, 9 (3): 720-732. 10.1002/pmic.200700816.PubMedView ArticleGoogle Scholar
- Powers RW, Kaeberlein M, Caldwell SD, Kennedy BK, Fields S: Extension of chronological life span in yeast by decreased TOR pathway signaling. Genes Dev. 2006, 20 (2): 174-184. 10.1101/gad.1381406.PubMedPubMed CentralView ArticleGoogle Scholar
- Pozniakovsky AI, Knorre DA, Markova OV, Hyman AA, Skulachev VP, Severin FF: Role of mitochondria in the pheromone- and amiodarone- induced programmed death of yeast. J Cell Biol. 2005, 168 (2): 257-269. 10.1083/jcb.200408145.PubMedPubMed CentralView ArticleGoogle Scholar
- Braun RJ, Zischka H, Madeo F, Eisenberg T, Wissing S, Buttner S, Engelhardt SM, Buringer D, Ueffing M: Crucial mitochondrial impairment upon CDC48 mutation in apoptotic yeast. J Biol Chem. 2006, 281 (35): 25757-25767. 10.1074/jbc.M513699200.PubMedView ArticleGoogle Scholar
- Eisenberg T, Buttner S, Kroemer G, Madeo F: The mitochondrial pathway in yeast apoptosis. Apoptosis. 2007, 12 (5): 1011-1023. 10.1007/s10495-007-0758-0.PubMedView ArticleGoogle Scholar
- Fabrizio P, Battistella L, Vardavas R, Gattazzo C, Liou LL, Diaspro A, Dossen JW, Gralla EB, Longo VD: Superoxide is a mediator of an altruistic aging program in Saccharomyces cerevisiae. J Cell Biol. 2004, 166 (7): 1055-1067. 10.1083/jcb.200404002.PubMedPubMed CentralView ArticleGoogle Scholar
- Festjens N, Vanden Berghe T, Vandenabeele P: Necrosis, a well-orchestrated form of cell demise: signalling cascades, important mediators and concomitant immune response. Biochim Biophys Acta. 2006, 1757 (9–10): 1371-1387.PubMedView ArticleGoogle Scholar
- Mollinedo F, Gajate C: Lipid rafts and clusters of apoptotic signaling molecule-enriched rafts in cancer therapy. Future Oncol. 2010, 6 (5): 811-821. 10.2217/fon.10.34.PubMedView ArticleGoogle Scholar
- Gajate C, Mollinedo F: The antitumor ether lipid ET-18-OCH(3) induces apoptosis through translocation and capping of Fas/CD95 into membrane rafts in human leukemic cells. Blood. 2001, 98 (13): 3860-3863. 10.1182/blood.V98.13.3860.PubMedView ArticleGoogle Scholar
- Ayllon V, Fleischer A, Cayla X, Garcia A, Rebollo A: Segregation of Bad from lipid rafts is implicated in the induction of apoptosis. J Immunol. 2002, 168 (7): 3387-3393.PubMedView ArticleGoogle Scholar
- Thomas BJ, Rothstein R: Elevated recombination rates in transcriptionally active DNA. Cell. 1989, 56 (4): 619-630. 10.1016/0092-8674(89)90584-9.PubMedView ArticleGoogle Scholar
- Sherman F: Getting started with yeast. Methods Enzymol. 2002, 350: 3-41.Google Scholar
- Guaragnella N, Pereira C, Sousa MJ, Antonacci L, Passarella S, Corte-Real M, Marra E, Giannattasio S: YCA1 participates in the acetic acid induced yeast programmed cell death also in a manner unrelated to its caspase-like activity. FEBS Lett. 2006, 580 (30): 6880-6884. 10.1016/j.febslet.2006.11.050.PubMedView ArticleGoogle Scholar
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