The unfolded protein response (UPR) is an intracellular signaling pathway that counteracts variable stresses that impair protein folding in the endoplasmic reticulum (ER). As such, the UPR is thought to be a homeostat that finely tunes ER protein folding capacity and ER abundance according to need. The mechanism by which the ER stress sensor Ire1 is activated by unfolded proteins and the role that the ER chaperone protein BiP plays in Ire1 regulation have remained unclear. Here we show that the UPR matches its output to the magnitude of the stress by regulating the duration of Ire1 signaling. BiP binding to Ire1 serves to desensitize Ire1 to low levels of stress and promotes its deactivation when favorable folding conditions are restored to the ER. We propose that, mechanistically, BiP achieves these functions by sequestering inactive Ire1 molecules, thereby providing a barrier to oligomerization and activation, and a stabilizing interaction that facilitates de-oligomerization and deactivation. Thus BiP binding to or release from Ire1 is not instrumental for switching the UPR on and off as previously posed. By contrast, BiP provides a buffer for inactive Ire1 molecules that ensures an appropriate response to restore protein folding homeostasis to the ER by modulating the sensitivity and dynamics of Ire1 activity. Secreted and membrane-spanning proteins constitute one of every three proteins produced by a eukaryotic cell. Many of these proteins initially fold and assemble in the endoplasmic reticulum (ER). A variety of physiological and environmental conditions can increase the demands on the ER, overwhelming the ER protein folding machinery. To restore homeostasis in response to ER stress, cells activate an intracellular signaling pathway called the unfolded protein response (UPR) that adjusts the folding capacity of the ER according to need. Its failure impairs cell viability and has been implicated in numerous disease states. In this study, we quantitatively interrogate the homeostatic capacity of the UPR. We arrive at a mechanistic model for how the ER stress sensor Ire1 cooperates with its binding partner BiP, a highly redundant ER chaperone, to fine-tune UPR activity. Moving between a predictive computational model and experiments, we show that BiP release from Ire1 is not the switch that activates Ire1; rather, BiP modulates Ire1 activation and deactivation dynamics. BiP binding to Ire1 and its dissociation in an ER stress-dependent manner buffers the system against mild stresses. Furthermore, BiP binding accelerates Ire1 deactivation when stress is removed. We conclude that BiP binding to Ire1 serves to fine-tune the dynamic behavior of the UPR by modulating its sensitivity and shutoff kinetics. This function of the interaction between Ire1 and BiP may be a general paradigm for other systems in which oligomer formation and disassembly must be finely regulated.

The accumulation of misfolded proteins activates the unfolded protein response in the endoplasmic reticulum. The transmembrane protein Ire1 is a central player in this pathway, acting as a kinase and an endoribonuclease. It excises an intron on HAC1 mRNA resulting in translation of the transcription factor Hac1 that in turn activates target genes. This issue reports two studies on Ire1. Korennykh et al. solve the crystal structure of Ire1 kinase and show that it spontaneously assembles into a rod-shaped oligomer. This positions the kinase domains for trans-phosphorylation, orders the RNase domains and creates an interaction site for mRNA substrate binding. Aragón et al. show that on activation, Ire1 molecules cluster into discrete foci containing high-order oligomers on the endoplasmic reticulum membrane. HAC1 mRNA is recruited to these foci by means of a sequence in its 3′ untranslated region and is processed at these sites. In this way the HAC1 mRNA is delivered to a site where it is processed, ensuring that it is translated only when the unfolded protein response is on. It is shown that after activation, Ire1 molecules cluster into discrete foci containing high order oligomers on the endoplasmic reticulum membrane. HAC1 mRNA is recruited to these foci by means of a sequence in its 3′ UTR and is processes at these sites. Disruption of either Ire1 clustering or HAC1 mRNA recruitment impairs UPR signalling. Hence HAC1 mRNA is delivered to a site where it is processed thus ensuring that it is only translated when UPR is on. Deficiencies in the protein-folding capacity of the endoplasmic reticulum (ER) in all eukaryotic cells lead to ER stress and trigger the unfolded protein response (UPR)1,2,3. ER stress is sensed by Ire1, a transmembrane kinase/endoribonuclease, which initiates the non-conventional splicing of the messenger RNA encoding a key transcription activator, Hac1 in yeast or XBP1 in metazoans. In the absence of ER stress, ribosomes are stalled on unspliced HAC1 mRNA. The translational control is imposed by a base-pairing interaction between the HAC1 intron and the HAC1 5′ untranslated region4. After excision of the intron, transfer RNA ligase joins the severed exons5,6, lifting the translational block and allowing synthesis of Hac1 from the spliced HAC1 mRNA to ensue4. Hac1 in turn drives the UPR gene expression program comprising 7–8% of the yeast genome7 to counteract ER stress. Here we show that, on activation, Ire1 molecules cluster in the ER membrane into discrete foci of higher-order oligomers, to which unspliced HAC1 mRNA is recruited by means of a conserved bipartite targeting element contained in the 3′ untranslated region. Disruption of either Ire1 clustering or HAC1 mRNA recruitment impairs UPR signalling. The HAC1 3′ untranslated region element is sufficient to target other mRNAs to Ire1 foci, as long as their translation is repressed. Translational repression afforded by the intron fulfils this requirement for HAC1 mRNA. Recruitment of mRNA to signalling centres provides a new paradigm for the control of eukaryotic gene expression.

Autophagy is a cytosolic quality control process that recognizes substrates through receptor‐mediated mechanisms. Procollagens, the most abundant gene products in Metazoa, are synthesized in the endoplasmic reticulum (ER), and a fraction that fails to attain the native structure is cleared by autophagy. However, how autophagy selectively recognizes misfolded procollagens in the ER lumen is still unknown. We performed siRNA interference, CRISPR‐Cas9 or knockout‐mediated gene deletion of candidate autophagy and ER proteins in collagen producing cells. We found that the ER‐resident lectin chaperone Calnexin (CANX) and the ER‐phagy receptor FAM134B are required for autophagy‐mediated quality control of endogenous procollagens. Mechanistically, CANX acts as co‐receptor that recognizes ER luminal misfolded procollagens and interacts with the ER‐phagy receptor FAM134B. In turn, FAM134B binds the autophagosome membrane‐associated protein LC3 and delivers a portion of ER containing both CANX and procollagen to the lysosome for degradation. Thus, a crosstalk between the ER quality control machinery and the autophagy pathway selectively disposes of proteasome‐resistant misfolded clients from the ER.

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The endoplasmic reticulum (ER) is dedicated to import, folding and assembly of all proteins that travel along or reside in the secretory pathway of eukaryotic cells. Folding in the ER is special. For instance, newly synthesized proteins are N-glycosylated and by default form disulfide bonds in the ER, but not elsewhere in the cell. In this review, we discuss which features distinguish the ER as an efficient folding factory, how the ER monitors its output and how it disposes of folding failures.