Year: 2008
Type: dissertation
Abstract: In the yeast Saccharomyces cerevisiae, acetic acid triggers a mitochondria-mediated death pathway with apoptotic characteristics. In mammalian cells, the mitochondrial outer membrane permeabilization (MOMP), necessary for the release of pro-apoptotic proteins, is a pivotal event for the activation of the apoptotic cascade in numerous cell death pathways. MOMP is thought to be mediated by a complex of proteins that constitute the permeability transition pore (PTP). Since S. cerevisiae possesses orthologues of the components believed to be involved in mammalian PTP composition/regulation, it was used herein to study such proteins both concerning their role in mitochondrial permeabilization and involvement in the course of cell death. The proteins studied were Por1p (yeast voltage-dependent anion channel, VDAC), Cpr3p (mitochondrial cyclophilin) and Aac1/2/3p (ADP/ATP carrier, AAC). We found that during apoptosis triggered by acetic acid deletion of CPR3 has no effect. Absence of Por1p enhances and absence of AAC proteins decreases acetic acidinduced apoptosis indicating an anti- and pro-apoptotic role, respectively, for these proteins. Moreover, the pro-death role of AAC does not require the ADP/ATP translocase activity. Absence of AAC proteins impairs MOMP and release of cytochrome c, which is degraded along with other mitochondrial inner membrane proteins. We observe that, during acetic acid-induced apoptosis, caspase activation is independent of AAC proteins but strongly dependent on the growth phase of the culture. In addition, a strain deleted for the yeast metacaspase YCA1 shows decreased overall caspase activation but still died exhibiting apoptotic features, supporting the existence of an Yca1p-independent apoptotic pathway. Fragmentation and degradation of mitochondria are common events in mammalian apoptosis. Both fragmentation and degradation are able to strongly affect the course of cell death and a relation has been proposed between these events and MOMP/cytochrome c release. Interestingly, por1Δ cells exhibit fragmented mitochondrial reticulum in the absence of any death stimulus. This phenotype however, does not contribute to the apoptosis stimulation observed in por1Δ mutant. We observe that during acetic acidinduced apoptosis the absence of AAC proteins leads to aggregation of fragmented mitochondria and a slower degradation of these organelles. Degradation of mitochondria in response to acetic acid is not due to classical autophagy or mediated by Uth1p-dependent mitophagy. We show that mitochondrial degradation during acetic acid-induced apoptosis is dependent on the protease Pep4p that is released from the vacuole to the cytosol. pep4Δ cells, which are strongly impaired in mitochondria degradation, are sensitized to acetic acid and mainly die by necrosis. This suggests that mitochondria degradation in response to acetic acid helps to sustain the apoptotic process. Taken together, the results show that vacuolar and mitochondrial proteins interfere with mitochondria morphological remodelling and subsequent degradation, suggesting that there is a complex interplay between these organelles in the regulation of yeast cell death. In conclusion, we were able exploring the distinctive ability of yeast to survive without respiration-competent mitochondria to study the involvement of mitochondria and mitochondria-interacting proteins in cell death. Additional studies using this model will undoubtedly further our understanding of the complex cell death processes.
Author: Clara Pereira
Language: English
Subfield: Molecular Biology
Domain: Life Sciences
Open Access status: green
OpenAlex ID: W111256701