SUMMARY When unfolded proteins accumulate to irremediably high levels within the endoplasmic reticulum (ER), intracellular signaling pathways called the unfolded protein response (UPR) become hyperactivated to cause programmed cell death. We discovered that thioredoxin-interacting protein (TXNIP) is a critical node in this “Terminal UPR.” TXNIP becomes rapidly induced by IRE1α, an ER bifunctional kinase/endoribonuclease (RNase). Hyperactivated IRE1α increases TXNIP mRNA stability by reducing levels of a TXNIP destabilizing micro-RNA, miR-17. In turn, elevated TXNIP protein activates the NLRP3 inflammasome, causing Caspase-1 cleavage and interleukin 1β (IL-1β) secretion. Txnip gene deletion reduces pancreatic β-cell death during ER stress, and suppresses diabetes caused by proinsulin misfolding in the Akita mouse. Finally, small molecule IRE1α RNase inhibitors suppress TXNIP production to block IL-1β secretion. In summary, the IRE1α-TXNIP pathway is used in the terminal UPR to promote sterile inflammation and programmed cell death, and may be targeted to develop effective treatments for cell degenerative diseases.

SUMMARY Depending on endoplasmic reticulum (ER) stress levels, the ER transmembrane multi-domain protein IRE1α promotes either adaptation or apoptosis. Unfolded ER proteins cause IRE1α lumenal domain homo-oligomerization, inducing trans auto-phosphorylation that further drives homo-oligomerization of its cytosolic kinase/ endoribonuclease (RNase) domains to activate mRNA splicing of adaptive XBP1 transcription factor. However, under high/chronic ER stress, IRE1α surpasses an oligomerization threshold that expands RNase substrate repertoire to many ER-localized mRNAs, leading to apoptosis. To modulate these effects, we developed ATP-competitive IRE1α Kinase Inhibiting RNase Attenuators—KIRAs—that allosterically inhibit IRE1α’s RNase by breaking oligomers. One optimized KIRA, KIRA6, inhibits IRE1α in vivo and promotes cell survival under ER stress. Intravitreally, KIRA6 preserves photoreceptor functional viability in rat models of ER stress-induced retinal degeneration. Systemically, KIRA6 preserves pancreatic β-cells, increases insulin, and reduces hyperglycemia in Akita diabetic mice. Thus, IRE1α powerfully controls cell fate, but can itself be controlled with small molecules to reduce cell degeneration.

Glutathione contributes to thiol-redox control and to extra-mitochondrial iron-sulphur cluster (ISC) maturation. To determine the physiological importance of these functions and sort out those that account for the GSH requirement for viability, we performed a comprehensive analysis of yeast cells depleted of or containing toxic levels of GSH. Both conditions triggered an intense iron starvation-like response and impaired the activity of extra-mitochondrial ISC enzymes but did not impact thiol-redox maintenance, except for high glutathione levels that altered oxidative protein folding in the endoplasmic reticulum. While iron partially rescued the ISC maturation and growth defects of GSH-depleted cells, genetic experiments indicated that unlike thioredoxin, glutathione could not support by itself the thiol-redox duties of the cell. We propose that glutathione is essential by its requirement in ISC assembly, but only serves as a thioredoxin backup in cytosolic thiol-redox maintenance. Glutathione-high physiological levels are thus meant to insulate its cytosolic function in iron metabolism from variations of its concentration during redox stresses, a model challenging the traditional view of it as prime actor in thiol-redox control.