finding provides an explanation of how CHOP sensitizes cells to ER stress and why
CHOP-/- cells are more resistant to tunicamycin-induced apoptosis. Other potential
transcriptional targets of CHOP have been identified, but at present it is unclear how
they could affect cell growth and apoptosis (Wang et al., 1998b).
2.4
Caspase-12The mechanism by which stress in the ER is coupled to activation of caspases was for
the most part a mystery until caspase-12 was characterized by Nakagawa and Yuan.
Caspase-12 is ubiquitously expressed and, like all caspases, synthesized as an inactive
proenzyme consisting of a regulatory prodomain, a large p20 subunit, and a small
p10 subunit (Van de Craen et al., 1997; Nakagawa and Yuan, 2000). However, unlike
other caspases, caspase-12 is remarkably specific to ER stress induced apoptosis since
it is activated (as measured by proteolysis of the precursor into distinct cleavage
fragments) by insults that elicit ER stress, but not by agents, such as staurosporin or
TNFα/anti-Fas, that activate mitochondrial or death receptor pathways, respectively
(Nakagawa et al., 2000). Accordingly, caspase-12-null MEFs are partially resistant to
apoptosis induced by Brefeldin A, tunicamycin, and thapsigargin, but not to
staurosporin, anti-Fas, or TNFα. In addition, mice deficient in caspase-12 show
increased survival and greater resistance to renal tubular epithelial cell death following
intraperitoneal injection of tunicamycin (Nakagawa et al., 2000). The partial
protection observed in caspase-12-deficient mice clearly indicates the importance of
this caspase in ER stress apoptosis, but also suggests that caspase-12 does not function
alone to initiate apoptosis. This finding is not surprising given the redundancy of
signaling in the UPR between Irel and ATF-6.
Caspase-12 is localized at the cytosolic face of the ER, placing it in a position to
respond to ER stress as a proximal signaling molecule. Several groups have proposed
different models of caspase-12 activation. Nakagawa and Yuan (2000) showed that
when mouse glial cells undergo oxygen and glucose deprivation (OGD), a condition
that induces ER stress, caspase-12 is cleaved from about 50-kDa size to two ~35-kDa
fragments in a calpain-, but not caspase-, dependent manner. Calpains are calcium-
activated cytosolic proteases that have been implicated in several forms of apoptosis
(Wang, 2000). In addition to agents such as A-23187 and thapsigargin that directly
mobilize ER calcium stores, other ER stress agents, such as tunicamycin, have also
been found to increase immediately the levels of cytosolic Ca2+ (Carlberg et al., 1996).
Release of Ca2+ may be a conserved feature in the ER stress response that triggers
calpain activation at the ER membrane and cleavage of caspase-12, or activation of
other Ca2+-dependent cell-death pathways (see below). m-Calpain can directly cleave
caspase-12 in vitro at T132 and K158, releasing the prodomain from the p20/p10
subunits and generating the two ~35-kDa fragments observed in vivo with antibody
against the p20 subunit. Calpain-cleaved caspase-12 may then autoactivate in trans,
by self-cleavage at D316, separating the p20 and p10 subunits; therefore, they could
form the active heterotetrameric p20/p10 complex characteristic of all caspases
THE ROLE OF THE ENDOPLASMIC RETICULUM IN APOPTOSIS 101