Genetics of Apoptosis

3.1

Endoplasmic reticulum targeting

Lee et al. (1999) selectively targeted Bcl-2 to mitochondria or endoplasmic reticulum
(ER) with COOH-terminal fusions to organelle-targeting sequences from ActA and
cytochrome b5. Certain apoptotic stimuli in Rat-1 fibroblasts were susceptible to
both ER—and mitochondria-targeted Bcl-2 inhibition (serum-starvation plus c-myc
overexpression, taxol, and ceramide), but others (etoposide) could be rescued only
with mitochondria-targeted or wild-type Bcl-2. Annis et al. (2001) postulated that
ER-targeted Bcl-2 rescued a subset of apoptotic deaths characterized by early
mitochondrial depolarization prior to cytochrome c release.
Speculation about ER-specific functions of Bcl-2 has centered on calcium
transport. Lam et al. (1994) initially reported in 1993 that Bcl-2 inhibited the efflux
of ER calcium induced by thapsigargin (TG), an irreversible inhibitor of the ER Ca^2
+-ATPase pump (SERCA). This result has not been consistently confirmed, with two

groups in agreement (Foyouzi-Youssefi et al., 2000; Nutt et al., 2002), but other
groups showing that Bcl-2 increases TG-induced ER Ca2+ efflux (Kuo et al., 1998;
Williams et al., 2000) or has no effect (Ichimiya et al., 1998). A detailed analysis of
altered Ca2+ flux in Bcl-2-expressing cells has suggested increased ER Ca2+ uptake
(SERCA-dependent or—independent) (He et al., 1997; Kuo et al., 1998; Kobrinsky
and Kirchberger, 2001), increased mitochondrial Ca2+ buffering capacity (Ichimiya
et al., 1998; Zhu et al., 1999), or increased permeability of ER membranes to Ca2+
(Foyouzi-Youssefi et al., 2000; Pinton et al., 2000) as mechanisms. Foyouzi-Youssefi
et al. and Pinton et al. observed lower ER calcium concentrations in Bcl-2 compared
to control cells by direct measurements with the Ca2+-sensitive fluorescent probes,
cameleons and aequorins, respectively.
One logical explanation for the antiapoptotic activity of ER-localized Bcl-2 is the
binding of proapoptotic BH proteins and prevention of their translocation to
mitochondria. Alternatively, Bcl-2 may interact with known ER proteins to carry out
an organelle-specific antiapoptotic function. Ng et al. (1997) identified binding of
Bcl-xL/Bcl-2 to Bap31, an integral ER membrane protein. In the model of adenovirus
E1A-induced apoptosis, Bap31 is cleaved by caspase-8 at two sites to yield a
proapoptotic NH 2 -terminal p20 fragment. The association of Bcl-xL and Bap31,
stabilized by procaspase-8 and the C. elegans Apaf-1 homolog Ced-4, prevented
proteolysis of Bap31 (Ng and Shore, 1998).
Hacki et al. (2000) demonstrated a connection between ER stress pathways,
induced by brefeldin A or tunicamycin, and mitochondrial cytochrome c release. This
inter-organellar cross-talk was blocked by ER-localized Bcl-2. In a possibly related
finding, Zuppini et al. (2000) related proteolytic cleavage of Bap31 cleavage to ER
stress, and noted reduced processing in cells deficient in the ER lectin chaperone,
calnexinS.

64 GENETICS OF APOPTOSIS