Genetics of Apoptosis

In contrast to Bax, Bak is found inserted into the outer mitochondrial membrane
of normal cells (Griffiths et al., 1999). Similar to Bax, activation of Bak has been
shown to be associated with changes in its tertiary and quaternary structure. When
Jurkat or CEM-C7A cells were exposed to the apoptosis inducers staurosporine,
etoposide, or dexamethasone, changes in the tertiary structure of Bak were detected
(Griffiths et al., 1999). Antibodies to the N-terminal domain of Bak were reactive
only after apoptotic stimulation, suggesting that changes had occurred in this domain
of the protein. Furthermore, although Bcl-XL coimmunoprecipitated with Bak before
stimulation, no coimmunoprecipitation was detected after stimulation. This result
suggests that the proapoptotic activity of Bak is neutralized by binding to Bcl-XL in
the mitochondrial membrane, and that after apoptotic stimulation conformational
changes in Bak result in dissociation from Bcl-XL, leaving Bak free to exert its
proapoptotic activity (Griffiths et al., 1999). Another study showed that tcBid can
activate Bak. tcBid translocated to the mitochondria and induced conformational
changes in Bak, resulting in Bak oligomerization in the mitochondrial membrane
and release of cytochrome c from the mitochondria (Wei et al., 2000). Combined,
these results show that conformational changes appear to be a major mechanism for
activation of proapoptotic members of the multidomain group. So far, only one
protein, Bid, has been shown to activate directly the multidomain proteins. However,
it would be surprising if additional proteins were not also able to activate the
multidomain proteins through direct interactions.

7.

‘BH3-domain-only’ proteins as multidomain protein activators

The ‘BH3-domain-only proteins’ appear to function mainly through activation of
the multidomain proteins. Bid was shown a few years ago to provide a shunt between
the two apoptotic pathways, the death receptor and the mitochondrial pathway
(Schmitz et al., 1999). In some cell types (so-called type 2 cells), Fas-induced apoptosis
results in a low activation of caspase-8 at the membrane receptor. This appears to be
insufficient for activation of the direct caspase pathway. In these cells, caspase-8
cleaves Bid, generating a C-terminal fragment (tcBid) that interacts with Bax or Bak,
and results in activation of these proteins and the mitochondrial pathway. This
mechanism functions as an amplification loop to enhance the apoptotic intracellular
signaling cascade. tcBid has been shown to translocate to the mitochondria, where it
is found in apoptotic cells (Gross et al., 1999b). Whether the protein colocalizes only
with Bax or Bak or whether it has an independent activity at the mitochondrial
membrane still remains unclear. Fibroblasts from Bax and Bak double-deficient mice
were completely resistant to tcBid-induced cell death. The mice also showed a
pronounced increased resistance to hepatocyte damage after treatment with anti-Fas
antibodies (Wei et al., 2001). At least in these two systems, Bid acts through activation
of the multidomain proteins. However, in artificial membrane systems, tcBid has a
membrane-destabilizing effect (Kudla et al., 2000). Whether such an activity is also
present under physiologic conditions remains to be elucidated. It is conceivable that

130 GENETICS OF APOPTOSIS