Structural and Functional Significance of the Endoplasmic Reticulum Unfolded Protein Response Transducers and Chaperones at the Mitochondria-ER Contacts: A Cancer Perspective

Abstract

In the last decades, the endoplasmic reticulum (ER) has emerged as a key coordinator of cellular homeostasis, thanks to its physical interconnection to almost all intracellular organelles. In particular, an intense and mutual crosstalk between the ER and mitochondria occurs at the mitochondria-ER contacts (MERCs). MERCs ensure a fine-tuned regulation of fundamental cellular processes, involving cell fate decision, mitochondria dynamics, metabolism, and proteostasis, which plays a pivotal role in the tumorigenesis and therapeutic response of cancer cells. Intriguingly, recent studies have shown that different components of the unfolded protein response (UPR) machinery, including PERK, IRE1α, and ER chaperones, localize at MERCs. These proteins appear to exhibit multifaceted roles that expand beyond protein folding and UPR transduction and are often related to the control of calcium fluxes to the mitochondria, thus acquiring relevance to cell survival and death. In this review, we highlight the novel functions played by PERK, IRE1α, and ER chaperones at MERCs focusing on their impact on tumor development.

Keywords: cancer; endoplasmic reticulum; mitochondria–ER contacts; molecular chaperones; unfolded protein response.

FIGURE 1

The unfolded protein response (UPR) signaling pathway. In unstressed conditions (upper box), the activation of inositol-requiring enzyme 1a (IRE1α), pancreatic ER kinase (PERK), and activating transcription factor 6a (ATF6α) is inhibited by the binding of Bip/GRP78. During the endoplasmic reticulum (ER) stress, binding of glucose regulated protein 78-kDa/binding immunoglobulin protein (GRP78/BiP) to misfolded proteins allows the activation of IRE1α, PERK, and ATF6α (lower box). Activated IRE1α cleaves 26-nucleotides from the X-box binding protein 1 (XBP1) mRNA allowing the translation of XBP1; hyper-oligomerized IRE1α executes regulated IRE1a-dependent decay (RIDD) activity on selected cytosolic mRNAs (Cox and Walter, 1996; Hollien et al., 2009). Activated PERK phosphorylates the eukaryotic initiation factor 2a (eIF2α) leading to attenuation of protein synthesis and to the preferential translation of activating transcription factor 4 (ATF4) mRNA (Harding et al., 2000). ATF6α activation is achieved in the Golgi complex where it undergoes to intramembrane proteolysis-specific cleavage by site-1 protease (S1P) and S2P to produce a transcriptionally active fragment (pATF6α). XBP1, ATF4, and pATF6α are responsible for the execution of the UPR transcriptional program in the nucleus (Walter and Ron, 2011).

FIGURE 2

Schematic representation of relevant events involving IRE1α and PERK proteins at mitochondria–ER contacts (MERCs). (A) PERK-involving events at MERCs: (i) PERK tethers the ER to mitochondria promoting the rapid transfer of reactive oxygen species (ROS) signals, likely under the form of lipid hydroperoxides (Verfaillie et al., 2012); (ii) Mitofusin 2 (Mfn2) lies upstream of PERK and under basal conditions maintains PERK inactive (Munoz et al., 2013); the PERK/eIF2α/ATF4 signaling pathway upregulates the expression of SERCA1 truncated proteins (S1T) (iii) and sigma-1 receptor (σ1R) (iv) at MERCs (Chami et al., 2008; Mitsuda et al., 2011). (B) Events involving IRE1α at MERCs: (i) σ1R-dependent stabilization of IRE1α oligomerization at MERCs (Mori et al., 2013); (ii) mitochondrial ubiquitin ligase (MITOL)-dependent ubiquitylation of IRE1α at MERCs prevents ER stress induced apoptosis (Takeda et al., 2019); (iii) the AKT-mammalian target of rapamycin (mTOR) signaling attenuates IRE1 RNase activity by promoting the re-establishment of ER-mitochondria contacts (Sanchez-Alvarez et al., 2017); (iv) IRE1α scaffolds IP₃R at MERCs to sustain calcium transfer and mitochondrial bioenergetics (Carreras-Sureda et al., 2019). The putative pro-survival or pro-apoptotic outputs of the depicted events are reported.

FIGURE 3

Schematic representation of Ca²⁺ handling at MERCs in healthy cell and cancer cell. (A) Healthy cell: The ER Ca²⁺ load is determined by ER Ca²⁺ uptake systems sarco-ER Ca2 + transport ATPases (SERCAs) and Ca²⁺-binding protein calnexin (CLNX), which is targeted to MERCs following a phosphofurin-acidic cluster sorting protein 2 (PACS2)-dependent palmitoylation to maintain high Ca²⁺ levels with the ER. Ca²⁺ is released from the ER through the activation of inositol 1,4,5-tris phosphate receptors (IP3Rs) Ca²⁺ channels that are physically connected to voltage-dependent anion channel (VDAC) located at the outer mitochondrial membrane (OMM) via glucose-regulated protein 75-kDa (GRP75). The Ca²⁺ ions transferred through the IP3Rs–GRP75–VDAC complex are imported into the mitochondrial matrix via the mitochondrial Ca²⁺ uniporter (MCU) complex. The activity of IP3Rs is controlled by several chaperones. The oxidoreductase ER oxidoreductin 1a (Ero1α) interacts with IP3Rs and modulates Ca²⁺ flux in a redox-sensitive manner. In addition, the σ1R is released from the Ca²⁺-dependent chaperone GRP78/BiP and promotes prolonged ER calcium release by stabilizing IP3R3. Regulation of Ca2 + efflux from ER is regulated, also, through the Sec61 channel that acts as a passive ER calcium leak channel. (B) Mitochondrial Ca²⁺ uptake affects cell death pathways. Following apoptotic stimuli, at MERCs, calcium uptake by the IP3R–GRP75–VDAC complex favors Ca²⁺ overload that promotes OMM permeabilization, inner mitochondrial membrane (IMM) cristae remodeling, and the opening of the permeability transition pore (PTP). This causes a rapid collapse of the membrane potential and the swelling of mitochondria, with consequent loss of cytochrome c that is released into the cytosol to trigger apoptosis. (C) At MERCs, ER chaperones regulate the Ca²⁺ homeostasis. Overexpression of GRP78/BiP at MERCs increases the Ca²⁺ storage capacity of the ER and attenuates apoptosis. Red arrows indicate inhibitory effect of GRP78/78 on the passive Ca²⁺ efflux through the Sec61 channel. Under chronic ER stress, involving prolonged ER Ca²⁺ depletion, σ1R translocates to the peripheral ER and attenuates cellular damage; in this way, σ1R is no more able to stabilize IP3R3, thereby preventing cell death. CLNX could be considered as a novel biomarker as its upregulation is related to the increased activity of sarco/endoplasmic reticulum Ca²⁺ATPase (SERCA) pump, that concentrates Ca²⁺ into the ER lumen, allowing resistance to cell death. Ero1-α and GRP75 contribution to the tumorigenesis is still unclear.