expression of caspase-8 is cytotoxic. In contrast, autoactivation of executioner
caspase-3 or -6 from proforms cannot be detected in yeast. However, coexpression
of procaspase-8 results in procaspase-3 activation and produces a pattern of
morphologic changes similar to mammalian apoptosis.
Expression of CED-4 from C. elegans in S.pombe also leads to an apoptotic
phenotype, whereas coexpression of CED-9, a bcl-2 homolog, prevents chromatin
condensation (James et al., 1997). The possibility of rescuing yeast with antiapoptotic
factors acting downstream in the mammalian apoptotic cascade indicates that Bax,
Bak, CED-4, or caspases do not simply act as cytotoxic substances in yeast, but seem
to activate the same or a similar mechanism as in metazoan organisms.
3.
Mutations leading to apoptosis in yeastAll previously described examples of yeast apoptotic cell death involved heterologous
expression of metazoan apoptotic inducers. In 1997, we found a yeast mutant dying
with a typical apoptotic phenotype: exposure of phosphatidylserine, margination of
chromatin, and formation of cell fragments (Madeo et al., 1997). While the TUNEL
test indicates massive DNA breakage, no DNA ladder is observed. Nucleosome
linkers therefore appear not to be preferred targets of DNA cleavage in yeast, probably
due to their short length (Lowary and Widom, 1989).
The mutation causing the apoptotic phenotype is a point-mutation of CDC48
(cdc48S565G), a S.cerevisiae protein belonging to the AAA family. AAA proteins are
ATPases that extract proteins from protein complexes and are involved in protein
degradation and in vesicle fusion. CDC48p is necessary for homotypic vesicle fusion
in both ER and the Golgi apparatus (Latterich et al., 1995), and participates in the
extraction and degradation of misfolded proteins from the ER (ERAD) (Ye et al.,
2001; Rabinovich et al., 2002).
Recently, Granot and colleagues showed that Bax-triggered apoptosis in yeast can
be blocked by enhancement of vesicle trafficking. Moreover, a downregulation of
vesicular transport enhances the susceptibility of yeast cells to apoptosis, providing
an explanation for the cdc48S565G-mediated apoptosis^ (Levine et al., 2001). Future
work will reveal whether vesicular trafficking also plays a role in metazoan apoptosis,
and whether this process is closely linked to apoptosis, or whether the connection is
rather indirect.
In the case of CDC48, yeast has demonstrated its potential to characterize new
mammalian apoptotic regulators. In 1999, the antiapoptotic role of the human
Cdc48p ortholog VCP/p97 and a related protein in C. elegans was described
(Shirogane et al., 1999; Wu, D. et al., 1999). Intriguingly, VCP was found in a screen
for new antiapoptotic proteins. A mutated form, similar to the cdc48S565G mutation
in yeast, induces apoptosis dominantly in B cells. This makes CDC48/VCP the first
apoptotic regulator to be discovered by its function in yeast apoptosis. Recently,
another AAA protein has been described that modifies Bax-induced apoptosis in yeast.
According to Manon et al. (2001), expression of bax in yeast induces not only a release
REGULATORS AND APPLICATIONS OF YEAST APOPTOSIS 143