ORIGINAL PAPER

New directions in ER stress-induced cell death

Susan E. Logue • Patricia Cleary • Svetlana Saveljeva •

Afshin Samali

� Springer Science+Business Media New York 2013

Abstract Endoplasmic reticulum (ER) stress has been

implicated in the pathophysiology of many diseases

including heart disease, cancer and neurodegenerative

diseases such as Alzheimer’s and Huntington’s. Prolonged

or excessive ER stress results in the initiation of signaling

pathways resulting in cell death. Over the past decade

much research investigating the onset and progression of

ER stress-induced cell death has been carried out. Owing to

this we now have a better understanding of the signaling

pathways leading to ER stress-mediated cell death and

have begun to appreciate the importance of ER localized

stress sensors, IRE1a, ATF6 and PERK in this process. In

this article we provide an overview of the current thinking

and concepts concerning the various stages of ER stress-

induced cell death, focusing on the role of ER localized

proteins in sensing and triggering ER stress-induced death

signals with particular emphasis on the contribution of

calcium signaling and Bcl-2 family members to the exe-

cution phase of this process. We also highlight new and

emerging directions in ER stress-induced cell death

research particularly the role of microRNAs, ER-mito-

chondria cross talk and the prospect of mitochondria-

independent death signals in ER stress-induced cell death.

Keywords Endoplasmic reticulum � Stress � Unfolded

protein response � Cell death

Introduction

ER stress is triggered due to a loss of homeostasis in the

ER causing accumulation of misfolded proteins within

the ER lumen. Examples of such physiological stresses

include hypoxia, glucose deprivation and oxidative stress,

conditions which can also often be found associated

with tumor microenvironments. Three ER transmembrane

receptors IRE1a (inositol requiring enzyme/endonuclease

1), PERK (double stranded RNA-activated protein Kinase

(PKR)-like ER kinase) and ATF6 (activating transcription

factor 6) constantly monitor the ‘‘health’’ of the ER.

Under normal conditions each receptor is maintained

in an inactive state through binding, via their luminal

domain, with the ER chaperone protein Grp78 (Bip,

HspA5). Accumulation of unfolded proteins triggers

dissociation of Grp78 (owing to a higher affinity for

unfolded proteins) from IRE1a, PERK and ATF6 facili-

tating their activation. Upon Grp78 release, IRE1adimerizes and autophosphorylates activating its kinase

and endonuclease functions [1]. Likewise, PERK dimer-

izes and autophosphorylates, activating its kinase domain

[1], while ATF6 translocates to the Golgi where site 1

protease (S1P) and site 2 protease (S2P) process it to

generate an active transcription factor which subsequently

translocates to the nucleus [2]. The collective signaling

pathways initiated by these ER stress receptors are

commonly referred to as the unfolded protein response

(UPR). The UPR is a highly conserved stress pathway

which functions as a short term adaptive mechanism

aimed at reducing levels of unfolded proteins and

restoring balance to the ER. However, if the UPR is

insufficient to deal with chronic exposure to ER stress-

inducing stimuli then a switch to ER stress-induced death

signaling commences.

S. E. Logue � P. Cleary � S. Saveljeva � A. Samali (&)

Apoptosis Research Centre, NUI Galway, Galway, Ireland

e-mail: afshin.samali@nuigalway.ie

123

Apoptosis

DOI 10.1007/s10495-013-0818-6

ER stress-mediated death initiation

IRE1a

IRE1a is an ER transmembrane protein containing a kinase

and endoribonuclease (RNase) domain on its cytosolic

portion [3]. Oligomerization of IRE1a by Grp78 dissocia-

tion juxtaposes the kinase domains causing trans-

autophosphorylation which in addition to activating its

kinase activity also triggers the endoribonuclease activity

of IRE1a. By virtue of its RNase activity, IRE1a splices a

26 nucleotide intron from XBP1 mRNA causing a frame

shift enabling translation and generation of a basic leucine

zipper family transcription factor, spliced XBP1 (XBP1s)

[4]. XBP1s has a diverse range of target genes which share

the common aim of short term adaption and ultimately

restoration of ER function. The majority of XBP1 target

genes are involved in either increasing the folding capacity

of the ER or associated with the degradation of accumu-

lated proteins with the aim of reducing ER protein load.

Temporal analysis of IRE1a activation in response to ER

stress found it to be an early event which diminished upon

prolonged stress [5]. Moreover, expression of a mutant

form of IRE1a, in which RNase activity can be selectively

activated, lead to an enhancement in cell survival upon

treatment with ER stress inducers indicating pro-survival

functions [5]. Recent reports also suggest XBP1s signaling

may be able to modulate apoptotic signaling. Upon IL-3

deprivation, BaF3 cells stably expressing XBP1s exhibited

increased survival which was in part attributed to modu-

lation of Bcl-2 family members including Bim [6]. Fur-

thermore, overexpression of XBP1s in MCF-7 cells

increased Bcl-2 levels following stimulation with Tamox-

ifen or ethanol [7]. Currently, it remains unknown whether

XBP1s can modulate Bcl-2 family member expression

during ER stress. It is possible XBP1s targets stretch

beyond proteins directly involved in ER function and it

may be actively involved in the suppression of apoptosis

through the modulation of Bcl-2 family members however

future studies will be needed to verify this.

Owing to the pro-survival targets of XBP1s, IRE1asignaling is generally regarded as an adaptive response.

Indeed work by Lin and colleagues, investigating the tem-

poral activation of UPR stress sensors, found IRE1a sig-

naling to be attenuated in cells undergoing prolonged ER

stress supporting the hypothesis that this pathway does not

actively participate in pro-apoptotic signals [5]. However,

overexpression of IRE1a in HEK293T cells has been

reported to induce death indicating there must be pro-

apoptotic signaling components [3]. Indeed the recruitment

of TNF receptor associated factor 2 (TRAF2) to IRE1a has

been linked to several pro-apoptotic pathways the most well

defined being the IRE1a-TRAF2-JNK axis [8]. The

association of IRE1a with TRAF2 triggers phosphorylation

cascades involving ASK1 and culminating in JNK activa-

tion. JNK-mediated phosphorylation has been demonstrated

to modulate Bcl-2 family member function. For example,

phosphorylation of Bcl-2/Bcl-xL by JNK can reduce their

anti-apoptotic ability while phosphorylation of Bid and Bim

by JNK has been demonstrated to increase their pro-apop-

totic ability [9–12]. Therefore, IRE1a-mediated JNK acti-

vation may represent a mechanism through which IRE1acan manipulate relative levels of pro- and anti-apoptotic

Bcl-2 family members thus tipping the balance in favor of

apoptosis (Fig. 1). IRE1a signaling has also been suggested

to modulate cellular release of TNFa which can feedback in

an autocrine manner and activate death receptor signaling.

Again this pathway is mediated by the adapter protein

TRAF2 which recruits IKK to the IRE1a complex where it

is phosphorylated and activated. Activated IKK phosphor-

ylates IjB tagging it for degradation thereby permitting

NF-jB translocation to the nucleus and upregulation of

target genes such as TNFa [13] (Fig. 1). TNF receptor 1

(TNFR1) has also been implicated in IRE1a-mediated JNK

signaling with JNK activation found to be deficient in

TNFR1-/- MEFs exposed to ER stress [14]. It is proposed

that upon ER stress TNFR1 co-localizes with RIP1 and

IRE1a at the membrane of the ER and that this complex is

necessary for the optimal JNK signaling upon ER stress and

execution of apoptosis [14].

RIDD

The RNase activity of IRE1a has recently been linked to a

process referred to as regulated IRE1-dependent decay of

mRNAs (RIDD). RIDD was first described in D. melano-

gaster where IRE1a activity was shown to mediate the

rapid decay of ER localized mRNAs [15]. Subsequent

studies have also verified the existence of RIDD in mam-

malian cells [16]. While this process is reliant upon IRE1aRNase activity it is distinct from XBP1 splicing and is

reported to selectively target and degrade mRNAs encod-

ing secretory proteins involved in protein folding within

the ER. Initial activation of RIDD would be expected to aid

cell survival by reducing the protein load on the ER.

However, prolonged RIDD signaling has been reported to

correlate with increased apoptosis [16]. The switch

between anti-apoptotic XBP1s signaling and pro-apoptotic

RIDD may be dependent upon the conformational state of

IRE1a. Administration of a peptide domain derived from

the kinase domain of IRE1a triggered IRE1a oligomeri-

zation and XBP1 cleavage but diminished RIDD and JNK

activation [16]. IRE1a mediated RIDD activation is a new

phenomenon in the field of ER stress and further studies are

required to identify RIDD targets and appreciate the

mechanisms controlling its activation.

Apoptosis

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Regulation of IRE1a signaling

Since IRE1a can elicit pro-survival and pro-apoptotic

signals, mechanisms controlling the switch between the

two must exist. Recent studies have revealed that IRE1asignaling is indeed finely controlled by a complex array of

protein interactions with IRE1a as the central core com-

ponent [17]. Pro-apoptotic Bcl-2 family members Bax and

Bak positively modulate the amplitude of IRE1a signaling

by interacting at the ER with the cytoplasmic domains of

IRE1a resulting in increased XBP1s and JNK phosphory-

lation [18]. Binding of Bax and Bak to IRE1 is negatively

regulated by Bax Inhibitor 1 (BI-1), a transmembrane

protein localized to the ER and nuclear envelope. Nor-

mally, BI-1 is ubiquitinated by bi-functional apoptosis

regulator (BAR) leading to proteosomal degradation.

Under prolonged ER stress BAR expression is downregu-

lated, BI-1 expression maintained and IRE1a signaling

attenuated [19]. Recent work by Hetz and colleagues has

proposed another layer of regulation mediated by BH3-

only Bcl-2 family members. Under mild ER stress BH3-

only proteins Bim and PUMA bind IRE1a, via their

BH3 domain, and stimulate its RNase activity. Indeed upon

induction of mild ER stress an in vivo reduction in XBP1s

levels was determined in Bim-/- mice. However, upon

sustained or chronic ER stress BH3-only proteins resume

their pro-apoptotic function and target mitochondrial

mediated pathways committing the cell to death [20].

Hsp72 has also recently been demonstrated to bind to and

regulate IRE1a signaling. Gupta and colleagues reported

binding of Hsp72 to the cytosolic domain of IRE1a, an

interaction which increased the RNase activity of IRE1aresulting in an increase in XBP1 splicing [21]. Based on the

current data it appears that numerous mechanisms may

regulate the amplitude of IRE1a signaling, and through this

mechanism control the switch from pro-survival to pro-

apoptotic signaling. Future studies are required to fully

determine the complexity of IRE1a regulation.

PERK

Following dissociation from Grp78, PERK dimerizes,

autophosphorylates and signals for a general translational

inhibition by phosphorylating elongation initiation factor

2a (eIF2a) [22] (Fig. 1). This general block in translation

promotes cell survival by providing the cell with a window

of opportunity to reduce the backlog of unfolded proteins

thereby alleviating ER stress. The importance of this

translational block is clearly evident in the hypersensitivity

to ER stress-induced death of PERK-/- MEFs and knock-

in non-phosphorylatable eIF2a cells [23, 24]. However, the

translational block is not absolute as genes with particular

regulatory sequences in their 50 untranslated region, such as

Fig. 1 Unfolded protein response: IRE1, PERK and ATF6 activation.

Cells cope with stressful conditions by activating the unfolded protein

response. This response is mediated via the dissociation of Grp78 from

three ER transmembrane proteins IRE1a, PERK and ATF6. a Following

dissociation of Grp78, IRE1a oligomerizes and autophosphorylates

facilitating its activation. Active IRE1a induces splicing of XBP1 mRNA

to XBP1s and also activates JNK via TRAF2 and ASK1. Furthermore,

active IRE1a has been linked to downstream NF-jB activation and also

RIDD, which can lead to the degradation of pro-survival mRNA. b Like

IRE1a, PERK dimerizes and autophosphorylates following Grp78

dissociation. Active PERK mediates its response via phosphorylation

of eIF2a leading to a translational block and cap independent translation

of ATF4. ATF4 induces CHOP which has multiple downstream targets.

c Following Grp78 dissociation, ATF6 is transported to the Golgi where it

is cleaved from its membrane anchor. Little is known about ATF6

regulated pathways but it is involved in the upregulation of UPR

associated genes, XBP1, CHOP, Grp78, PDI and EDEM1

Apoptosis

123

an internal ribosome entry site (IRES) can bypass the

translational block with activating transcription factor 4

(ATF4) being one such example [25]. ATF4 is a member of

the CCAAT/enhancer binding protein family (C/EBP)

family of transcription factors. The majority of transcrip-

tional targets of ATF4 are associated with cell survival and

include genes involved in amino acid metabolism, redox

reactions, protein secretion and stress responses [26]. As

such transcription of this subset of genes in conjunction

with translation inhibition should reduce levels of ER

stress. However, when stress cannot be alleviated ATF4

helps push the cell towards death by upregulating tran-

scription factor C/EBP homologous protein (CHOP) [23].

CHOP

CHOP upregulation is a common point of convergence for

all 3 arms of the UPR with binding sites for ATF6, ATF4

and XBP1s present within its promoter clearly illustrating

the importance of this transcription factor. CHOP signaling

is thought to mediate cell death signaling by firstly altering

the transcription of genes involved in apoptosis and oxi-

dative stress and secondly by relieving PERK mediated

translational inhibition [27]. Pro-apoptotic targets of CHOP

include BH3-only members of the Bcl-2 family. Puthala-

kath and colleagues demonstrated Bim upregulation in

MCF-7 cells specifically in response to ER stress-inducing

agents. Furthermore, knockdown of Bim in MCF-7 cells

significantly attenuated ER stress-induced cell death

clearly highlighting a role of Bim in the execution of ER

stress-induced apoptosis. Dissection of the specific path-

ways regulating Bim revealed that a combination of tran-

scriptional upregulation via CHOP and post translational

modification namely protein phosphatase 2a (PP2a)-medi-

ated dephosphorylation enabled sustained Bim expression

[28]. CHOP has also been reported to regulate expression

of BH3 only proteins by interacting with FOXO3A (in

neuronal cells treated with tunicamycin) [29] and AP-1

complex protein c-Jun leading to its phosphorylation (in

saturated fatty acid treated hepatocytes) [30].

CHOP-mediated downregulation of Bcl-2 has also been

reported in response to ER stress suggesting that this

transcription factor may shift the balance of Bcl-2 family

members in favor of pro-apoptotic thus ensuring propaga-

tion and execution of the apoptotic signal [31]. Other

transcriptional targets of CHOP include endoplasmic

reticulum oxidoreductin 1 (ERO1a) and tibbles related

protein 3 (TBR3). Increased ERO1a expression results in a

hyperoxidizing environment within the ER which may

promote cell death [32]. Additionally ERO1a has been

reported to activate the inositol trisphoshate receptor

(IP3R) stimulating calcium release from the ER [33],

concurrent uptake by the mitochondria may lead to calcium

overload and apoptosis. TBR3 is an intracellular pseudo-

kinase that modulates the activity of several signal trans-

duction kinases. Overexpression of TRB3 has been linked

to cell death onset while knockdown of TRB3 in 293 and

HeLa cells was reported to attenuate tunicamycin induced

death [34]. The mechanism through which TRB3 mediates

death signals is not understood but it has been suggested

that TRB3 promotes apoptosis through binding AKT,

preventing its phosphorylation and reducing its kinase

activity [35, 36]. In addition to transcriptional control of

pro- and anti-apoptotic genes CHOP activation also lifts

translational inhibition mediated by PERK phosphorylation

of eIF2a. CHOP mediated enhancement of GADD34 per-

mits protein phosphatase 1 (PP1) dephosphorylation of

eIF2a thus lifting translational inhibition [37]. Release of

this translational block permits production of pro-apoptotic

proteins further committing the cell to death. Inhibition of

eIF2a dephosphorylation, by treatment with salubrinal,

inhibited ER stress-induced apoptosis underscoring the

contribution of releasing translational inhibition to pro-

gression of cell death [38]. Indeed, the importance of

GADD34 signaling for ER stress-induced apoptosis is

clearly evident in knockout mice which displayed resis-

tance to ER stress-induced kidney damage [32].

The importance of CHOP-mediated signaling to ER

stress-induced apoptosis is clearly illustrated by the pres-

ence of binding sites for ATF6, ATF4 and XBP1s in its

promoter region. However important CHOP signaling is to

the ER stress-induced apoptosis, the requirement for it is

not absolute as CHOP-/- MEF cells still undergo apoptosis

in response to prolonged ER stress albeit with much slower

kinetics [32].

ATF6

Owing to the presence of an ER targeted hydrophobic

sequence ATF6 is an ER tethered protein. Following dis-

sociation of Grp78, ATF6 translocates to the Golgi where

SP1 and SP2 proteases cleave it releasing active ATF6 into

the cytosol [2]. This bZip transcription factor family

member upregulates expression of genes mainly involved

in adapting to ER stress such as Grp78, Protein Disulphide

Isomerase (PDI) and ER degradation-enhancing a-man-

nosidase-like protein 1 (EDEM1) [39]. ATF6 also increa-

ses transcription of XBP1 mRNA, an important IRE1atarget [40]. ATF6 signaling is largely pro-survival and

adaptive, however it can also be pro-apoptotic. ATF6 has

been demonstrated to upregulate levels of CHOP during

sustained ER stress [40]. Although not in an ER stress

context selective activation of ATF6 apoptotic myoblasts

during the differentiation process has been reported and

linked to the downregulation of the anti-apoptotic protein

Mcl-1 highlighting a potential pro-apoptotic role for ATF6

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[41]. Whether ATF6 can mediate downregulation of Mcl-1

or other anti-apoptotic Bcl-2 family members during ER

stress is currently unknown.

Mitochondria-mediated death signaling

As discussed above sustained activation of UPR signals

can result in the upregulation of pro-apoptotic Bcl-2

family members such as CHOP-mediated activation of

Bim. Other BH3 only proteins transcriptionally regulated

by ER stress include PUMA and Noxa. Puma and Noxa

are pro-apoptotic BH3 family members often referred

to as ‘sensitizers’ of apoptosis, with Noxa reported to

interact with Mcl-1 and A1 while Puma is thought to

interact with various members of the pro-survival Bcl-2

family leading to subsequent MOMP induction [42,

43].Transcriptional activation of both Puma and Noxa in

response to ER stress has been reported in a p53-

dependent manner [44]. Partial suppression of ER stress-

induced apoptosis has been reported in p53-/- cells and

attributed to defective induction of Puma and Noxa [44].

The mechanism facilitating p53 activation during ER

stress has not been fully elucidated. Recent studies sug-

gest p53 upregulation during ER stress occurs in a NF-jB

dependent manner [45]. Interestingly, IRE1a, PERK and

ATF6 have all been linked to the activation NF-jB sig-

nals under various circumstances. PERK-mediated trans-

lational inhibition has been reported to lower levels of the

short half-life protein IjB, permitting NF-jB transloca-

tion to the nucleus [46]. IRE1a signals have also been

implicated in NF-jB activation via TRAF2 recruitment of

IKK permitting translocation of NF-jB [47], while ATF6

signaling has been implicated in NF-jB activation during

shiga toxin treatment of rat Nrk52e renal proximal

tubular cells [48]. Knockdown of p53 exerted protective

effects against ER stress induced by tunicamycin or

brefeldin A in MCF-7 cells indicating it may have an

important role in mediating death signals [45]. Given the

diverse targets of NF-jB it is likely that its activation

increases expression of pro-apoptotic proteins such as

BH3-only proteins thus committing the cell to apoptosis.

CHOP and ATF4 have also been implicated in PUMA

and Noxa induction respectively [30, 44]. The importance

of BH3-only protein induction is illustrated by PUMA

and Noxa null MEFs which like Bim null MEFs exhibit

partial resistance to ER stress-induced apoptosis [44].

Work by Futami and colleagues in which they carried out

a siRNA screen for genes regulating ER stress-induced

apoptosis confirmed a functional role for Noxa and Puma

[49]. In neuronal cells, Puma transcriptional induction

alone is crucial for the execution of apoptosis in response

to ER stress [29]. The combination of increased BH3-

only protein expression, via predominately transcriptional

but also post-translation modifications in the case of Bim,

and repression of anti-apoptotic proteins such as Bcl-2

shifts the balance in favor of apoptosis permitting Bax-

Bak homo-oligomerization and mitochondrial outer

membrane permeabilization causing cytochrome c release

and subsequent apoptosome formation. Overexpression of

Bcl-2 reduces loss of mitochondrial membrane potential

and protects cells against ER stress inducers such as

thapsigargin underscoring the importance of mitochon-

drial mediated signals in the propagation of ER stress-

induced apoptosis [50].

ER/Mito Calcium cross talk and death

In addition to mediating death signals by triggering Bax-

Bak oligomerization and mitochondrial outer membrane

permeabilization, Bcl-2 family members have also been

implicated in the regulation of ER mitochondria calcium

signaling. The ER sequesters high concentrations of cal-

cium (1-3 mM) through a dynamic process of active uptake

via sarco/endoplasmic reticulum calcium transport ATPase

(SERCA) pumps and release through calcium channels

inositol trisphosphate receptor (IP3R) and ryanodine

receptors [51]. The maintenance of sufficient ER calcium

concentrations is imperative for ER function as many

chaperone proteins, such as Grp78, require calcium binding

to function at their optimum capacity [52]. Therefore, low

ER calcium concentrations reduce chaperone function and

disrupt the protein folding capacity of the ER resulting in a

backlog of unfolded proteins and ER stress.

In addition to their specialized cell death functions

recent work has demonstrated Bcl-2 family Bax, Bak, Bcl-

xL and Bcl-2 can associate with the ER both under basal

and stress conditions [53–55]. Reports indicate that ER

specific overexpression of Bcl-2 and Bcl-xL lower free ER

calcium concentration and increase protection against

apoptosis [54, 56]. The mechanism facilitating reduced free

calcium levels is thought to involve Bcl-2 Bcl-xL inter-

actions with IP3R possibly controlling channel opening.

The protective role of Bcl-2 in regulating calcium release

can be inhibited by the activation of kinases such as JNK.

Phosphorylation of Bcl-2 within an unstructured loop

region diminishes its anti-apoptotic protection by firstly

inhibiting its ability to bind and neutralize BH3-only pro-

teins and secondly by causing increased calcium release

from the ER (presumably by an inability to bind and reg-

ulate IP3R) which associated with an increase in mito-

chondria calcium uptake and pro-apoptotic signals [57].

Studies have also implicated the Bcl-2/Bcl-xL binding

partner BI-1 in regulation of ER calcium concentra-

tion. Overexpression of BI-1 in HT1080 cells reduced

Bax translocation, mitochondrial depolarization and ER

calcium release in response to thapsigargin treatment.

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A similar dysregulation in calcium release was present in

cells derived from BI-1-/- mice which displayed enhanced

calcium release and increased sensitivity to tunicamycin

compared to wild type BI-1?/? counterparts suggesting BI-

1 is important in the transmission of the death signal from

the ER to the mitochondria [58, 59].

Pro-apoptotic Bcl-2 family members Bax and Bak also

localize to the ER where they function to antagonize Bcl-2

and Bcl-xL increasing ER calcium concentration and

enhancing apoptotic sensitivity. The function of Bak and

Bax is nicely illustrated by Bax/Bak double knockout

MEFs which exhibit lower ER calcium concentrations and

increased resistance to calcium dependent apoptotic signals

[60]. Bax and Bak, analogous to their role in release of

mitochondrial intramembrane space proteins, can oligo-

merize at the ER during ER stress-induced apoptosis [55].

Recently it has been demonstrated that Bax-Bak oligo-

merization and insertion into the ER induces pore forma-

tion facilitating release of luminal proteins Grp78 and PDI

[61]. Whether calcium can be released by this mechanism

has not been determined.

Localization of BH3-only members of the Bcl-2 family

to the ER has also been described. For example, Bik a

primarily ER localized BH3-only protein can mediate Bax-

Bak dependent calcium release that has been shown to

participate in intrinsic apoptotic signals. Surprisingly Bik

upregulation in response to ER stress signals has not been

reported and it appears to be an event solely associated

with genotoxic stress [62]. Other members of the BH3-only

subfamily regulated by ER stress signals include Puma and

Noxa [44, 63]. Increased Puma expression has been linked

to depletion of ER calcium levels via Bax activation [64].

Bcl-2 family members help regulate both ER calcium

levels and release in response to pro-apoptotic signals.

Surprisingly, mitochondrial calcium transporters have a

low affinity for calcium and therefore require high levels to

stimulate mitochondrial uptake. Within the cell this is

achieved by contact sites between the ER and mitochondria

with high calcium concentrations enabling mitochondrial

calcium uptake. Such regions are referred to as mito-

chondria associated ER membranes (MAMs). MAMs

ensure the efficient shuttling of calcium between the ER

and mitochondria and as a consequence of this function are

enriched in IP3 receptors which are linked to voltage

dependent anion channel 1 (VDAC1) by the mitochondrial

chaperone protein Grp75 [65]. The importance of Grp75 in

this interaction has been demonstrated by knockdown of

Grp75 resulting in reduced mitochondrial calcium uptake

following agonist stimulation [66]. Regulation of MAM

signaling in response to ER stress has been reported. The

Sigma receptor 1 (Sig1-R) is an ER localized, calcium

sensitive, transmembrane chaperone which complexes with

Grp78 at MAMs. Calcium depletion from the ER causes

Grp78 dissociation from Sig1-R increasing their respective

chaperone activities and Sig1-R binding to and stabiliza-

tion of IP3 receptors. Upon conditions of chronic ER stress

Sig-1R redistribute from MAMs to the entire ER (via an

unknown mechanism) where presumably they attempt, via

their chaperone activity, to alleviate ER stress [67]. Indeed

overexpression of Sig-1Rs reduces ER stress responses

whereas knockdown of Sig1-R enhances apoptosis [67].

Another ER stress-induced MAM localized protein

recently implicated ER stress-induced apoptosis is sarco-

plasmic reticulum calcium ATPase 1 (S1T). Upon induction

of ER stress S1T expression is enhanced via PERK-eIF2a-

ATF4-CHOP signaling. Increased S1T expression increases

ER calcium depletion through a combination of increased

ER calcium leak, increased ER mitochondria contact sites

and inhibition of mitochondria movement [68]. Knockdown

of S1T expression reduced ER stress, mitochondrial cal-

cium overload and apoptosis highlighting an important role

for S1T in ER stress-induced apoptosis [68]. Aside from

facilitating increased ER mitochondria contact sites by

controlling S1T expression recent work has proposed that

PERK itself is an essential MAM component. Verfaille and

colleagues recently demonstrated PERK-/- cells have

weaker ER mitochondria contact sites resulting in dysreg-

ulated ER mitochondria calcium signaling [69]. Given that

PERK signaling is required for the regulation of many genes

upon induction of ER stress including S1T it would be

expected that the kinase domain of PERK is required for

maintenance of ER mitochondria interaction sites. Sur-

prisingly expression of a kinase dead mutant of PERK was

able to restore ER mitochondria interaction in PERK-/-

MEFs suggesting that PERK may, in addition to regulating

downstream effectors, also function as a scaffold protein

[69]. Indeed significant enrichment of PERK at MAMs was

identified; as yet the exact function of PERK at MAMs sites

is unknown.

Alternate modes of ER stress-induced cell death

Overexpression of Bcl-2 is able to inhibit ER stress-induced

apoptosis indicating an important role for mitochondrial

death signals in this process. Bcl-2 overexpression antago-

nizes ER stress-induced regulation of BH3-only proteins

preventing mitochondrial cytochrome c release and caspase

activation [70]. Additionally, Bcl-2 family members are

inherently important in ER mitochondria calcium signaling

with overexpression of Bcl-2 lowering ER calcium levels

thereby preventing mitochondrial calcium overload and

apoptosis. Likewise Bax-/- Bak-/- deficient cells exhibit

resistance to ER stress-induced apoptosis presumably

through a combination of mitochondrial and calcium medi-

ated processes. Several reports, using Bax-/- Bak-/- cells,

have demonstrated cell death upon prolonged exposure to

Apoptosis

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ER stress-inducing conditions/agents [71, 72]. In vitro

studies examining important regulators of ER stress-

induced apoptosis such as Bax-/- Bak-/- or caspase-9-/-

MEFs rarely extend ER stress treatment times beyond 48 h

as wild type cells have succumbed to death at this point

and inhibition in the knockout cells is evident. However,

prolonged ER stress conditions can initiate cell death in

mitochondrial-mediated apoptosis compromised cells such

as Bax-/- Bak-/- MEFs. This in itself is not an unex-

pected result as exposure to prolonged stress will at some

point trigger death via an alternate mechanism. Indeed

Bax-/- Bak-/- MEFs exhibit features of autophagy and

cell death in response to prolonged ER stress [73]. Studies

within our laboratory have demonstrated that cells defi-

cient in the mitochondrial pathway undergo an alternate

form of cell death involving aspects of autophagy (LC3 I

to II conversion) and apoptosis (caspase activation) when

exposed to prolonged stresses including ER stress

(unpublished results). Moreover, our data indicates that

caspase activation is dependent upon ATG5 indicating

cross-talk between autophagy and cell death pathways in

response to prolonged ER stress (unpublished results).

These findings are of considerable interest when we take

into account that many cancer cells are resistant to death

signals propagated via the mitochondrial pathway. Fur-

thermore in vivo such cells are exposed to stresses such as

sustained glucose deprivation or hypoxia that are known to

induce a robust ER stress response. Therefore, in the future

it will be important, in the context of diseases such as

cancer to understand how cells devoid of conventional

apoptotic signaling pathway retain susceptibility to ER

stress-induced death and in particular the role that

autophagy may play.

microRNAs and ER stress

The regulation of ER stress-induced death pathways by

microRNAs is a recent area of research with studies indi-

cating miRNAs can either directly modulate the ER stress

response or themselves be regulated by ER stress. For

example, Yang and colleagues demonstrated miR-122

overexpression downregulated ER stress responses in

HepG2 cells [74]. This observation is particularly inter-

esting in the context of hepatocellular cancer where

repression of miR-122 is frequently observed. The down-

regualtion of miR-122 would presumably lift repression on

UPR responses increasing the adaptive ability of the cancer

cell. Indeed in cisplatin treated Huh7 cells inhibition of

miR-122 decreased cell death highlighting the benefit of

miR-122 repression to cancer cells. [74]. ER stress-medi-

ated downregulation of miR-221/222 has been reported in

hepatocellular carcinoma cells where it associated with a

resistance to cell death [75]. Addition of miR-221/222

mimetics restored sensitivity to ER stress-induced apop-

tosis via a mechanism involving upregulation of p27kip1

and G1 phase arrest suggesting mimetics directed against

miR-122 or miR-221/222 maybe of therapeutic benefit

particularly in hepatocellular cancer [75].

Direct regulation of miRNA expression by ER stress

sensors particularly PERK has been reported and may

regulate the delicate balance that exists between pro-and

anti-apoptotic signaling during ER stress. PERK mediated

induction of miR-30c-2* has been reported during ER

stress and linked to a downregulation in XBP1 mRNA

reducing pro-survival signaling and aiding commitment to

cell death [76]. Additionally PERK mediated repression of

the mir-106b-25 cluster and its host gene MCM-7 has been

reported to result in increased Bim expression and apop-

tosis [77]. Conversely, recent work from Chitnis and col-

leagues implicates PERK facilitated miRNA regulation in

pro-survival signaling. miR-211 was identified as a PERK

target and demonstrated to repress CHOP expression

allowing a temporal window for the pro-survival response.

However, upon sustained ER stress miR-211 expression

was silenced, permitting CHOP accumulation and induc-

tion of the pro-apoptotic response [78]. Based on the cur-

rent literature it seems that miRNA regulation help shift the

balance between survival and cell death during ER stress.

Further research into ER stress-mediated regulation of

miRNAs is required to fully elucidate their role and

determine if they represent a viable therapeutic target.

Conclusions

ER stress-induced cell death is a complex and highly reg-

ulated process carefully controlled by ER localized stress

receptors. Initial signaling from each stress receptor aims to

reduce levels of unfolded proteins and restore cellular

homeostasis. However, following sustained or excessive ER

stress a switch in signaling from survival to death occurs

sealing the fate of the cell. Based on the current data IRE1aand PERK signals are important in cell death commitment.

Signals from each of these receptors have important roles in

regulating Bcl-2 family member expression particularly the

expression of BH3-only proteins. By tipping the balance in

favour of pro-apoptotic Bcl-2 family members pro-apop-

totic mitochondria-mediated signals are activated commit-

ting the cell to death. In addition to triggering Bax/Bak

oligomerization and cytochrome c release Bcl-2 family

members have recently been shown to function in ER

mitochondria cross talk thereby controlling calcium

movement between these two organelles. Recent studies

have highlighted the complexity of ER mitochondria cal-

cium signaling particularly the importance of MAMs in this

process. In the last few years the role of ER localised

Apoptosis

123

proteins Sigma 1 receptor and the calcium ATPase S1T in

ER mitochondria cross talk has emerged. The role of

MAMs and the proteins which regulate cross talk during ER

stress is one obvious area of research for the future. The role

of microRNAs in regulation of ER stress-induced cell death

also merits future research. It is only in the past few years

that we have started to appreciate the function of microRNAs

in ER dependent death signaling. Further work is required to

unmask the array of microRNA targets and determine their

function in ER stress-induced death.

Over the past 10 years the field of ER stress-induced

death has yielded much information concerning the basic

signaling mechanisms triggered. It is only now that we are

beginning to both understand the delicate balance of

interplay between pro-survival and pro-death signals.

Acknowledgments Our research is supported by grants from Sci-

ence Foundation Ireland (09/RFP/BIC2371), Breast Cancer Campaign

(2010NovPR13). P Cleary is funded by an Irish Cancer Society

Scholarship (CRS11CLE).

References

1. Bertolotti A, Zhang Y, Hendershot LM, Harding HP, Ron D

(2000) Dynamic interaction of BiP and ER stress transducers in

the unfolded-protein response. Nat Cell Biol 2(6):326–332

2. Ye J, Rawson RB, Komuro R, Chen X, Dave UP, Prywes R, Brown

MS, Goldstein JL (2000) ER stress induces cleavage of membrane-

bound ATF6 by the same proteases that process SREBPs. Mol Cell

6(6):1355–1364. doi:10.1016/s1097-2765(00)00133-7

3. Wang XZ, Harding HP, Zhang Y, Jolicoeur EM, Kuroda M, Ron

D (1998) Cloning of mammalian Ire1 reveals diversity in the ER

stress responses. EMBO J 17(19):5708–5717

4. Yoshida H, Matsui T, Yamamoto A, Okada T, Mori K (2001)

XBP1 mRNA is induced by ATF6 and spliced by IRE1 in

response to ER stress to produce a highly active transcription

factor. Cell 107(7):881–891

5. Lin JH, Li H, Yasumura D, Cohen HR, Zhang C, Panning B,

Shokat KM, LaVail MM, Walter P (2007) IRE1 signaling affects

cell fate during the unfolded protein response. Science

318(5852):944–949. doi:10.1126/science.1146361

6. Kurata M, Yamazaki Y, Kanno Y, Ishibashi S, Takahara T,

Kitagawa M, Nakamura T (2011) Anti-apoptotic function of

Xbp1 as an IL-3 signaling molecule in hematopoietic cells. Cell

Death Dis 10(2):e118. doi:10.1038/cddis

7. Gomez BP, Riggins RB, Shajahan AN, Klimach U, Wang A,

Crawford AC, Zhu Y, Zwart A, Wang M, Clarke R (2007)

Human X-Box binding protein-1 confers both estrogen indepen-

dence and antiestrogen resistance in breast cancer cell lines.

FASEB J 21(14):4013–4027

8. Urano F, Wang X, Bertolotti A, Zhang Y, Chung P, Harding HP,

Ron D (2000) Coupling of stress in the ER to activation of JNK

protein kinases by transmembrane protein kinase IRE1. Science

287(5453):664–666

9. Yamamoto K, Ichijo H, Korsmeyer SJ (1999) BCL-2 Is phos-

phorylated and inactivated by an ASK1/Jun N-terminal protein

kinase pathway normally activated at G2/M. Mol Cell Biol

19(12):8469–8478

10. Donovan N, Becker EBE, Konishi Y, Bonni A (2002)

JNK phosphorylation and activation of BAD couples the

stress-activated signaling pathway to the cell death machinery.

J Biol Chem 277(43):40944–40949. doi:10.1074/jbc.M206113200

11. Lei K, Davis RJ (2003) JNK phosphorylation of Bim-related

members of the Bcl2 family induces Bax-dependent apoptosis.

Proc Natl Acad Sci USA 100(5):2432–2437

12. Maundrell K, Antonsson B, Magnenat E, Camps M, Muda M,

Chabert C, Gillieron C, Boschert U, Vial-Knecht E, Martinou J-C,

Arkinstall S (1997) Bcl-2 undergoes phosphorylation by c-Jun

N-terminal kinase/stress-activated protein kinases in the presence

of the constitutively active GTP-binding protein Rac1. J Biol

Chem 272(40):25238–25242. doi:10.1074/jbc.272.40.25238

13. Hu P, Han Z, Couvillon AD, Kaufman RJ, Exton JH (2006)

Autocrine tumor necrosis factor alpha links endoplasmic reticu-

lum stress to the membrane death receptor pathway through

IRE1a-mediated NF-jB activation and down-regulation of

TRAF2 expression. Mol Cell Biol 26(8):3071–3084. doi:

10.1128/mcb.26.8.3071-3084.2006

14. Yang Q, Kim YS, Lin Y, Lewis J, Neckers L, Liu ZG (2006)

Tumour necrosis factor receptor 1 mediates endoplasmic reticu-

lum stress-induced activation of the MAP kinase JNK. EMBO

Rep 7(6):622–627

15. Hollien J, Weissman JS (2006) Decay of endoplasmic reticulum-

localized mRNAs during the unfolded protein response. Science

313(5783):104–107. doi:10.1126/science.1129631

16. Han D, Lerner AG, Walle LV, Upton JP, Xu W, Hagen A, Backes

BJ, Oakes SA, Papa FR (2009) IRE1a kinase activation modes

control alternate endoribonuclease outputs to determine divergent

cell fates. Cell 138(3):562–575

17. Woehlbier U, Hetz C (2011) Modulating stress responses by the

UPRosome: a matter of life and death. Trends Biochem Sci

36(6):329–337

18. Hetz C, Bernasconi P, Fisher J, Lee AH, Bassik MC, Antonsson

B, Brandt GS, Iwakoshi NN, Schrinzel A, Glimcher LH, Kors-

meyer SJ (2006) Proapoptotic BAX and BAK modulate the

unfolded protein response by a direct interaction with IRE1a.

Science 312(5773):572–576

19. Rong J, Chen L, Toth JI, Tcherpakov M, Petroski MD, Reed JC

(2011) Bifunctional apoptosis regulator (BAR), an endoplasmic

reticulum (ER)-associated E3 ubiquitin ligase, modulates BI-1

protein stability and function in ER stress. J Biol Chem

286(2):1453–1463. doi:10.1074/jbc.M110.175232

20. Rodriguez DA, Zamorano S, Lisbona F, Rojas-Rivera D, Urra H,

Cubillos-Ruiz JR, Armisen R, Henriquez DR, Cheng HE, Letek

M, Vaisar T, Irrazabal T, Gonzalez-Billault C, Letai A, Pimentel-

Muinos FX, Kroemer G, Hetz C (2012) BH3-only proteins are

part of a regulatory network that control the sustained signalling

of the unfolded protein response sensor IRE1[alpha]. EMBO J

31(10):2322–2335

21. Gupta S, Deepti A, Deegan S, Lisbona F, Hetz C, Samali A

(2010) HSP72 protects cells from ER stress-induced apoptosis via

enhancement of IRE1a-XBP1 signaling through a physical

interaction. PLoS Biol 8(7):e1000410. doi:10.1371/journal.pbio.

1000410

22. Harding HP, Zhang Y, Ron D (1999) Protein translation and

folding are coupled by an endoplasmic-reticulum-resident kinase.

Nature 397(6716):271–274

23. Harding HP, Novoa I, Zhang Y, Zeng H, Wek R, Schapira M,

Ron D (2000) Regulated translation initiation controls stress-

induced gene expression in mammalian cells. Mol Cell

6(5):1099–1108

24. Scheuner D, Song B, McEwen E, Liu C, Laybutt R, Gillespie P,

Saunders T, Bonner-Weir S, Kaufman RJ (2001) Translational

control is required for the unfolded protein response and in vivo

glucose homeostasis. Mol Cell 7(6):1165–1176

25. Lu PD, Harding HP, Ron D (2004) Translation reinitiation at

alternative open reading frames regulates gene expression in an

Apoptosis

123

integrated stress response. J Cell Biol 167(1):27–33. doi:

10.1083/jcb.200408003

26. Harding HP, Zhang Y, Zeng H, Novoa I, Lu PD, Calfon M, Sadri

N, Yun C, Popko B, Paules R, Stojdl DF, Bell JC, Hettmann T,

Leiden JM, Ron D (2003) An integrated stress response regulates

amino acid metabolism and resistance to oxidative stress. Mol

Cell 11(3):619–633. doi:10.1016/s1097-2765(03)00105-9

27. Oyadomari S, Mori M (2003) Roles of CHOP//GADD153

in endoplasmic reticulum stress. Cell Death Differ 11(4):

381–389

28. Puthalakath H, O’Reilly LA, Gunn P, Lee L, Kelly PN, Hun-

tington ND, Hughes PD, Michalak EM, McKimm-Breschkin J,

Motoyama N, Gotoh T, Akira S, Bouillet P, Strasser A (2007) ER

stress triggers apoptosis by activating BH3-only protein BIM.

Cell 129(7):1337–1349. doi:10.1016/j.cell.2007.04.027

29. Ghosh AP, Klocke BJ, Ballestas ME, Roth KA (2012) CHOP

potentially co-operates with FOXO3a in neuronal cells to regulate

PUMA and BIM expression in response to ER stress. PLoS ONE

7(6):e39586. doi:10.1371/journal.pone.0039586

30. Cazanave SC, Elmi NA, Akazawa Y, Bronk SF, Mott JL, Gores

GJ (2010) CHOP and AP-1 cooperatively mediate PUMA

expression during lipoapoptosis. Am J Physiol Gastrointest Liver

Physiol 299(1):G236–G243. doi:10.1152/ajpgi.00091.2010

31. McCullough KD, Martindale JL, Klotz LO, Aw TY, Holbrook NJ

(2001) Gadd153 sensitizes cells to endoplasmic reticulum stress

by down-regulating Bcl2 and perturbing the cellular redox state.

Mol Cell Biol 21(4):1249–1259

32. Marciniak SJ, Yun CY, Oyadomari S, Novoa I, Zhang Y, Jung-

reis R, Nagata K, Harding HP, Ron D (2004) CHOP induces

death by promoting protein synthesis and oxidation in the stressed

endoplasmic reticulum. Genes Dev 18(24):3066–3077

33. Li G, Mongillo M, Chin KT, Harding H, Ron D, Marks AR,

Tabas I (2009) Role of ERO1-a-mediated stimulation of inositol

1,4,5-triphosphate receptor activity in endoplasmic reticulum

stress-induced apoptosis. J Cell Biol 186(6):783–792

34. Ohoka N, Yoshii S, Hattori T, Onozaki K, Hayashi H (2005)

TRB3, a novel ER stress-inducible gene, is induced via ATF4-

CHOP pathway and is involved in cell death. EMBO J

24(6):1243–1255

35. Du K, Herzig S, Kulkarni RN, Montminy M (2003) TRB3: a

tribbles homolog that inhibits Akt/PKB activation by insulin in

liver. Science 300(5625):1574–1577

36. Zou CG, Cao XZ, Zhao YS, Gao SY, Li SD, Liu XY, Zhang Y,

Zhang KQ (2009) The molecular mechanism of endoplasmic

reticulum stress-induced apoptosis in PC-12 neuronal cells: the

protective effect of insulin-like growth factor I. Endocrinology

150(1):277–285

37. Novoa I, Zeng H, Harding HP, Ron D (2001) Feedback inhibition

of the unfolded protein response by GADD34-mediated dephos-

phorylation of eIF2a. J Cell Biol 153(5):1011–1021

38. Boyce M, Bryant KF, Jousse C, Long K, Harding HP, Scheuner

D, Kaufman RJ, Ma D, Coen DM, Ron D, Yuan J (2005) A

selective inhibitor of elF2a dephosphorylation protects cells from

ER stress. Science 307(5711):935–939

39. Adachi Y, Yamamoto K, Okada T, Yoshida H, Harada A, Mori K

(2008) ATF6 is a transcription factor specializing in the regula-

tion of quality control proteins in the endoplasmic reticulum. Cell

Struct Funct 33(1):75–89

40. Yoshida H, Okada T, Haze K, Yanagi H, Yura T, Negishi M,

Mori K (2000) ATF6 activated by proteolysis binds in the pres-

ence of NF-Y (CBF) directly to the cis-acting element responsible

for the mammalian unfolded protein response. Mol Cell Biol

20(18):6755–6767

41. Morishima N, Nakanishi K, Nakano A (2011) Activating tran-

scription factor-6 (ATF6) mediates apoptosis with reduction of

myeloid cell leukemia sequence 1 (Mcl-1) protein via induction

of WW domain binding protein. J Biol Chem 286(40):35227–

35235

42. Chen L, Willis SN, Wei A, Smith BJ, Fletcher JI, Hinds MG,

Colman PM, Day CL, Adams JM, Huang DCS (2005) Differ-

ential targeting of prosurvival Bcl-2 proteins by their BH3-only

ligands allows complementary apoptotic function. Mol Cell

17(3):393–403

43. Letai A, Bassik MC, Walensky LD, Sorcinelli MD, Weiler S,

Korsmeyer SJ (2002) Distinct BH3 domains either sensitize or

activate mitochondrial apoptosis, serving as prototype cancer

therapeutics. Cancer Cell 2(3):183–192. doi:10.1016/s1535-

6108(02)00127-7

44. Li J, Lee B, Lee AS (2006) Endoplasmic reticulum stress-induced

apoptosis: multiple pathways and activation of p53-UP-regulated

modulator of apoptosis (PUMA) and NOXA by p53. J Biol Chem

281(11):7260–7270

45. Lin W-C, Chuang Y-C, Chang Y-S, Lai M-D, Teng Y-N, Su I-J,

Wang CCC, Lee K-H, Hung J-H (2012) Endoplasmic reticulum

stress stimulates p53 expression through NF-jB activation. PLoS

ONE 7(7):e39120. doi:10.1371/journal.pone.0039120

46. Deng J, Lu PD, Zhang Y, Scheuner D, Kaufman RJ, Sonenberg

N, Harding HP, Ron D (2004) Translational repression mediates

activation of nuclear factor kappa B by phosphorylated transla-

tion initiation factor 2. Mol Cell Biol 24(23):10161–10168

47. Kaneko M, Niinuma Y, Nomura Y (2003) Activation signal of

nuclear factor-jB in response to endoplasmic reticulum stress is

transduced via IRE1 and tumor necrosis factor receptor-associ-

ated factor 2. Biol Pharm Bull 26(7):931–935

48. Yamazaki H, Hiramatsu N, Hayakawa K, Tagawa Y, Okamura

M, Ogata R, Huang T, Nakajima S, Yao J, Paton AW, Paton JC,

Kitamura M (2009) Activation of the Akt-NF-jB pathway by

subtilase cytotoxin through the ATF6 branch of the unfolded

protein response. J Immunol 183(2):1480–1487

49. Futami T, Miyagishi M, Taira K (2005) Identification of a net-

work involved in thapsigargin-induced apoptosis using a library

of small interfering RNA expression vectors. J Biol Chem 280(1):

826–831

50. Heath-Engel HM, Chang NC, Shore GC (2008) The endoplasmicreticulum in apoptosis and autophagy: role of the BCL-2 protein

family. Oncogene 27(50):6419–6433

51. Szegezdi E, MacDonald DC, Chonghaile TN, Gupta S, Samali A

(2009) Bcl-2 family on guard at the ER. Am J Physiol Cell

Physiol 296(5):C941–C953

52. Gaut JR, Hendershot LM (1993) The modification and assembly

of proteins in the endoplasmic reticulum. Curr Opin Cell Biol

5(4):589–595

53. Chen R, Valencia I, Zhong F, McColl KS, Roderick HL, Boot-

man MD, Berridge MJ, Conway SJ, Holmes AB, Mignery GA,

Velez P, Distelhorst CW (2004) Bcl-2 functionally interacts with

inositol 1,4,5-trisphosphate receptors to regulate calcium release

from the ER in response to inositol 1,4,5-trisphosphate. J Cell

Biol 166(2):193–203

54. White C, Li C, Yang J, Petrenko NB, Madesh M, Thompson CB,

Foskett JK (2005) The endoplasmic reticulum gateway to apop-

tosis by Bcl-XL modulation of the InsP3R. Nat Cell Biol

7(10):1021–1028

55. Zong WX, Li C, Hatzivassiliou G, Lindsten T, Yu QC, Yuan J,

Thompson CB (2003) Bax and Bak can localize to the endo-

plasmic reticulum to initiate apoptosis. J Cell Biol 162(1):59–69

56. Pinton P, Ferrari D, Magalhaes P, Schulze-Osthoff K, Di Virgilio

F, Pozzan T, Rizzuto R (2000) Reduced loading of intracellular

Ca2 ? stores and downregulation of capacitative Ca2 ? influx in

Bcl-2-overexpressing cells. J Cell Biol 148(5):857–862

57. Bassik MC, Scorrano L, Oakes SA, Pozzan T, Korsmeyer SJ

(2004) Phosphorylation of BCL-2 regulates ER Ca2 ? homeo-

stasis and apoptosis. EMBO J 23(5):1207–1216

Apoptosis

123

58. Chae HJ, Kim HR, Xu C, Bailly-Maitre B, Krajewska M, Kra-

jewski S, Banares S, Cui J, Digicaylioglu M, Ke N, Kitada S,

Monosov E, Thomas M, Kress CL, Babendure JR, Tsien RY,

Lipton SA, Reed JC (2004) BI-1 regulates an apoptosis pathway

linked to endoplasmic reticulum stress. Mol Cell 15(3):355–366

59. Bailly-Maitre B, Fondevila C, Kaldas F, Droin N, Luciano F,

Ricci JE, Croxton R, Krajewska M, Zapata JM, Kupiec-Weg-

linski JW, Farmer D, Reed JC (2006) Cytoprotective gene bi-1 is

required for intrinsic protection from endoplasmic reticulum

stress and ischemia-reperfusion injury. Proc Natl Acad Sci USA

103(8):2809–2814

60. Oakes SA, Scorrano L, Opferman JT, Bassik MC, Nishino M,

Pozzan T, Korsmeyer SJ (2005) Proapoptotic BAX and BAK

regulate the type 1 inositol trisphosphate receptor and calcium

leak from the endoplasmic reticulum. Proc Natl Acad Sci USA

102(1):105–110

61. Wang X, Olberding KE, White C, Li C (2011) Bcl-2 proteins

regulate ER membrane permeability to luminal proteins during

ER stress-induced apoptosis. Cell Death Differ 18(1):38–47

62. Mathai JP, Germain M, Shore GC (2005) BH3-only BIK regu-

lates BAX, BAK-dependent release of Ca2 ? from endoplasmic

reticulum stores and mitochondrial apoptosis during stress-

induced cell death. J Biol Chem 280(25):23829–23836

63. Reimertz C, Kogel D, Rami A, Chittenden T, Prehn JHM (2003)

Gene expression during ER stress-induced apoptosis in neurons:

induction of the BH3-only protein Bbc3/PUMA and activation of

the mitochondrial apoptosis pathway. J Cell Biol 162(4):587–597

64. Luo X, He Q, Huang Y, Sheikh MS (2005) Transcriptional

upregulation of PUMA modulates endoplasmic reticulum cal-

cium pool depletion-induced apoptosis via Bax activation. Cell

Death Differ 12(10):1310–1318

65. Grimm S (2012) The ER-mitochondria interface: the social net-

work of cell death. Biochim Biophys Acta 1823(2):327–334

66. Szabadkai G, Bianchi K, Varnai P, De Stefani D, Wieckowski

MR, Cavagna D, Nagy AI, Balla T, Rizzuto R (2006) Chaperone-

mediated coupling of endoplasmic reticulum and mitochondrial

Ca2 ? channels. J Cell Biol 175(6):901–911

67. Hayashi T, Su TP (2007) Sigma-1 receptor chaperones at the

ER-mitochondrion interface regulate Ca2 ? signaling and cell

survival. Cell 131(3):596–610

68. Chami M, Oules B, Szabadkai G, Tacine R, Rizzuto R, Paterlini-

Brechot P (2008) Role of SERCA1 truncated isoform in the

proapoptotic calcium transfer from ER to mitochondria during

ER stress. Mol Cell 32(5):641–651. doi:10.1016/j.molcel.2008.

11.014

69. Verfaillie T, Rubio N, Garg AD, Bultynck G, Rizzuto R,

Decuypere JP, Piette J, Linehan C, Gupta S, Samali A, Agostinis

P (2012) PERK is required at the ER-mitochondrial contact sites

to convey apoptosis after ROS-based ER stress. Cell Death Differ

11:1880–1891. doi:10.1038/cdd.2012.74

70. Samali A, Gupta S, Cuffe L, Szegezdi E, Logue SE, Neary C,

Healy S (2010) Mechanisms of ER stress-mediated mitochondrial

membrane permeabilization. Int J Cell Biol 2010:830307–

830318. doi:10.1155/2010/830307

71. Shimizu S, Kanaseki T, Mizushima N, Mizuta T, Arakawa-

Kobayashi S, Thompson CB, Tsujimoto Y (2004) Role of Bcl-2

family proteins in a non-apoptopic programmed cell death

dependent on autophagy genes. Nat Cell Biol 6(12):1221–1228

72. Buytaert E, Callewaert G, Vandenheede JR, Agostinis P (2006)

Deficiency in apoptotic effectors Bax and Bak reveals an auto-

phagic cell death pathway initiated by photodamage to the

endoplasmic reticulum. Autophagy 2(3):238–240

73. Ullman E, Fan Y, Stawowczyk M, Chen HM, Yue Z, Zong WX

(2008) Autophagy promotes necrosis in apoptosis-deficient cells

in response to ER stress. Cell Death Differ 15(2):422–425

74. Yang F, Zhang L, Wang F, Wang Y, Huo X, Yin Y, Sun SH

(2011) Modulation of the unfolded protein response is the core of

microRNA-122-involved sensitivity to chemotherapy in hepato-

cellular carcinoma 1,2. Neoplasia 13(7):590–600

75. Dai R, Li J, Liu Y, Yan D, Chen S, Duan C, Liu X, He T, Li H

(2010) MiR-221/222 suppression protects against endoplasmic

reticulum stress-induced apoptosis via p27 Kip1- and MEK/ERK-

mediated cell cycle regulation. Biol Chem 391(7):791–801

76. Byrd AE, Aragon IV, Brewer JW (2012) MicroRNA-30c-2*

limits expression of proadaptive factor XBP1 in the unfolded

protein response. J Cell Biol 196(6):689–698

77. Gupta S, Read DE, Deepti A, Cawley K, Gupta A, Oommen D,

Verfaillie T, Matus S, Smith MA, Mott JL, Agostinis P, Hetz C,

Samali A (2012) Perk-dependent repression of miR-106b-25

cluster is required for ER stress-induced apoptosis. Cell Death

Dis 3:e333. doi:10.1038/cddis.2012.74

78. Chitnis NS, Pytel D, Bobrovnikova-Marjon E, Pant D, Zheng H,

Maas NL, Frederick B, Kushner J, Chodosh L, Koumenis C,

Fuchs S, Diehl J (2012) miR-211 Is a prosurvival MicroRNA that

regulates chop expression in a PERK-dependent manner. Mol

Cell 48(3):353–364. doi:10.1016/j.molcel.2012.08.025

Apoptosis

123