SnapShot: The Unfolded Protein ResponseR. Luke Wiseman,1 Cole M. Haynes,2 and David Ron3,4
1The Scripps Research Institute, La Jolla, CA, USA; 2Memorial Sloan-Kettering Cancer Center, New York, NY, USA; 3NYU School of Medicine, New York, NY, USA; 4Institute of Metabolic Science, University of Cambridge, Cambridge, UK
The Unfolded Protein ResponseProtein folding capacity in the endoplasmic reticulum (ER) is matched to the flux of newly synthesized proteins passing through the secretory pathway by the activation of intra-cellular signaling pathways collectively referred to as the unfolded protein response (UPR). The UPR responds directly to the accumulation of unfolded proteins in the ER lumen. Accumulation of unfolded proteins activates signaling cascades in the cytosol via transmembrane sensor proteins in the ER membrane. Activation of the UPR results in both translational attenuation of new protein synthesis and transcriptional activation of stress response genes to relieve ER stress. Homeostasis of protein folding in the ER plays a key role in development and pathophysiology. Here, we present a SnapShot of key events in the UPR in yeast and metazoans.
UPR Activation in YeastThe UPR in the budding yeast Saccharomyces cerevisiae is triggered by a single transmembrane protein in the ER called Ire1. Ire1 monitors the folding status of newly synthe-sized proteins in the ER lumen through direct interactions with the ER chaperone proteins Kar2/BiP (reviewed in Ron and Walter, 2007). Upon accumulation of unfolded proteins, Ire1 is activated by dissociation from Kar2/BiP and direct interaction with the unfolded proteins (dotted arrow), resulting in the oligomerization of Ire1’s lumenal domain. Signal-ing is transduced to the cytosolic domain of Ire1 through the sequential steps of autophosphorylation, binding of ADP, and dimerization (followed by oligomerization) of the Ire1 endoribonuclease domain. This leads to the activation of the sequence-specific Ire1 ribonuclease (RNase) (Korennykh et al., 2009; Lee et al., 2008). The active Ire1 RNase and the tRNA ligase, Rlg1, collaborate to remove an intron from HAC1 mRNA (reviewed in Bernales et al., 2006). The resulting spliced mRNA is translated into Hac1p, a bZip transcrip-tion factor, which moves to the nucleus and upregulates genes encoding ER chaperones, components of the ER-associated degradation (ERAD) pathway, and proteins involved in ER expansion (e.g., lipid synthesis enzymes) (Travers et al., 2000). Through this Hac1-dependent transcriptional response, Ire1 signaling increases the folding capacity of the yeast ER and reduces the number of unfolded proteins, thus relieving ER stress.
UPR Activation in MetazoansThe metazoan UPR consists of three branches, each regulated by distinct transmembrane proteins in the ER: IRE1, PERK, and ATF6. The UPR integrates all three pathways to produce an ER stress response that uses both translational and transcriptional mechanisms (reviewed in Schroder and Kaufman, 2005).IRE1Metazoans have two orthologs of yeast Ire1, IRE1α and IRE1β, which are both activated by an evolutionarily conserved mechanism. This entails dimerization, phosphorylation, nucleotide binding, and, likely, the formation of a higher-order oligomer of undefined stoichiometry. Activation of IRE1 in metazoans leads to the splicing of XBP1 mRNA, which encodes a bZIP transcription factor that regulates UPR target genes. Unspliced XBP1 mRNA (XBP1U) encodes a short-lived transcription factor that represses the expression of these target genes. Excision of an intron from XBP1 mRNA through an IRE1-dependent splicing mechanism results in a frameshift in the XBP1 transcript (XBP1S). This transcript is translated into the UPR-activating transcription factor XBP1S, which upregulates genes similar to those upregulated by Hac1 in yeast.
In addition to the transcriptional remodeling of the ER folding environment, IRE1 is also involved in the activation of cell death pathways in response to prolonged ER stress. Phosphorylated IRE1 associates with tumor necrosis associated factor 2 (TRAF2) to modulate the activity of the c-Jun N-terminal kinase (JNK) pathway via the apoptosis sig-naling-regulating kinase (ASK1), which controls apoptosis through caspase 12 activation (reviewed in Ron and Walter, 2007). Additionally, IRE1 is linked to a nonspecific RNase activity that degrades mRNA localized to the ER membrane (Hollien et al., 2009), leading to a global reduction in proteins imported into the ER lumen (dotted arrow).PERKThe activation of PKR-like ER-localized eIF2α kinase (PERK) is also mediated by interaction with BiP. Under conditions of stress, BiP dissociates from the PERK lumenal domain. This triggers PERK dimerization, subsequent autophosphorylation, and then activation of the cytosolic kinase domain (reviewed in Ron and Harding, 2007). PERK phospho-rylates the heterotrimeric translation initiation factor eIF2 on its α subunit (eIF2α), which attenuates global protein translation through inhibition of the eIF2B complex, leading to a rapid reduction in the abundance of folding proteins in the ER. At high levels of eIF2α phosphorylation, NF-κB signaling is also activated (through relief of IκB inhibition), potentially promoting cell survival and an inflammatory response.
Paradoxically, eIF2α phosphorylation selectively increases translation of the activating transcription factor 4 (ATF4) mRNA through conserved upstream opening reading frames (uORFs) in the ATF4 transcript. The ATF4 protein moves to the nucleus and upregulates genes encoding amino acid transporters, redox enzymes that promote protein folding within the ER lumen (e.g., ERO1), and a proapoptotic transcription factor called the CCAAT enhancer-binding homologous protein (CHOP). ATF4 also increases transcrip-tion of XBP1 mRNA and, thus, provides a mechanism to amplify UPR signaling. PERK activity is balanced by the constitutively expressed repressor of eIF2α phosphorylation (CReP) and the stress-inducible GADD34 (an ATF4 target). Both CReP and GADD34 associate with protein phosphatase I (PPI) to dephosphorylate eIF2α and relieve PERK-dependent translation attenuation in a negative-feedback circuit.ATF6Activating transcription factor 6 (ATF6) is also regulated by interaction with BiP in the ER lumen. Dissociation of BiP from ATF6 permits packaging of ATF6 into COPII vesicles for trafficking to the Golgi. In the Golgi, ATF6 is sequentially cleaved by the site-1-protease (S1P) and site-2-protease (S2P), with release of the 50 kDa cytosolic domain of ATF6. This portion of ATF6 may then heterodimerize with XBP1 before trafficking to the nucleus to activate genes encoding ER chaperones (Yoshida et al., 2000). As activation of ATF6 is posttranslational, its target genes may represent the initial transcriptional response to ER stress, which is subsequently integrated with the IRE1 pathway once spliced XBP1 tran-scripts accumulate (Yoshida et al., 2001). ATF6 shares homology with two tissue-specific proteins: CREB-H in hepatocytes and OASIS in astrocytes (not shown), which activate the stress-dependent transcription of genes with promoters containing inflammatory response elements and cAMP-responsive elements, respectively (Bernales et al., 2006).
AbbreviationsADP, adenosine diphosphate; ERO1, ER oxidase 1; GADD34, growth arrest and DNA damage-inducible gene 34; IκB, inhibitor of kappa-B; IRE1, inositol requiring gene 1; NF-κB, nuclear factor-kappa-B; PP1, protein phosphatase 1; XBP1, X-box binding protein 1.
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SnapShot: The Unfolded Protein ResponseR. Luke Wiseman,1 Cole M. Haynes,2 and David Ron3,4
1The Scripps Research Institute, La Jolla, CA, USA; 2Memorial Sloan-Kettering Cancer Center, New York, NY, USA; 3NYU School of Medicine, New York, NY, USA; 4Institute of Metabolic Science, University of Cambridge, Cambridge, UK
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