ISRIB Blunts the Integrated Stress Response by Allosterically Antagonising the Inhibitory Effect of Phosphorylated eIF2 on eIF2B

Abstract

Summary The small molecule ISRIB antagonizes the activation of the integrated stress response (ISR) by phosphorylated translation initiation factor 2, eIF2(αP). ISRIB and eIF2(αP) bind distinct sites in their common target, eIF2B, a guanine nucleotide exchange factor for eIF2. We have found that ISRIB-mediated acceleration of eIF2B’s nucleotide exchange activity in vitro is observed preferentially in the presence of eIF2(αP) and is attenuated by mutations that desensitize eIF2B to the inhibitory effect of eIF2(αP). ISRIB’s efficacy as an ISR inhibitor in cells also depends on presence of eIF2(αP). Cryoelectron microscopy (cryo-EM) showed that engagement of both eIF2B regulatory sites by two eIF2(αP) molecules remodels both the ISRIB-binding pocket and the pockets that would engage eIF2α during active nucleotide exchange, thereby discouraging both binding events. In vitro , eIF2(αP) and ISRIB reciprocally opposed each other’s binding to eIF2B. These findings point to antagonistic allostery in ISRIB action on eIF2B, culminating in inhibition of the ISR.

Manipulation of the unfolded protein response: A pharmacological strategy against coronavirus infection

“…Subsequently, we evaluated the inhibitory effect of four different UPR inhibitors (UPRi) on each one of the UPR branches in cells infected with MHV for 8 h at MOI 5 (S5 Fig) . GSK-2606414 (henceforth referred to as PERKi) is a specific inhibitor of PERK [43,44]. As expected, PERKi treatment prevented autophosphorylation of PERK and reduced phosphorylation of its substrate, eIF2α (S5A Integrated stress response inhibitor (ISRIB) acts downstream of eIF2α in the PERK pathway by preventing p-eIF2α from binding and inhibiting eIF2B [45]. Therefore, eIF2B can recycle eIF2-GDP to active eIF2-GTP, and translation initiation can still occur, despite the levels of p-eIF2α remaining unchanged.…”

“…Integrated stress response inhibitor (ISRIB) acts downstream of eIF2α in the PERK pathway by preventing p-eIF2α from binding and inhibiting eIF2B [ 45 ]. Therefore, eIF2B can recycle eIF2-GDP to active eIF2-GTP, and translation initiation can still occur, despite the levels of p-eIF2α remaining unchanged.…”

PERK Pathway and Neurodegenerative Disease: To Inhibit or to Activate?

“…Another approach was the development of compounds that inhibit the pathway downstream of P-eIF2α, partially restoring protein synthesis and inhibiting ATF4 translation. ISRIB was, thus, identified in a high throughput screen and turned out to bind eIF2B causing it to be resistant to P-eIF2α inhibition [203][204][205][206]. ISRIB showed protective effects in a model of prion disease [207], without pancreatic toxicity.…”

Sugar phosphate activation of the stress sensor eIF2B

“…To identify potential changes in protein conformation upon sugar phosphate binding, we performed pairwise comparisons of eIF2B-F6P and eIF2Bα-M6P with the reported structures of apo eIF2B ( PDB 7D46 22 ) and eIF2Bα ( PDB 3ECS 27 ). eIF2Bα-M6P aligned to apo eIF2Bα with r.m.s.d.…”

“…However, these natural metabolites exert their effect by engaging the eIF2Bα 2 dimer, rather than by bridging the eIF2B(β/δ) 2 tetramer interface like ISRIB. Unlike the recently demonstrated activity of ISRIB as an antagonist of phosphorylated eIF2 binding 21 , 22 , the sugar phosphate mechanism is primarily driven by its ability to decamerize eIF2B. We generate point mutations in the eIF2Bα sugar-binding pocket, one of which is known to cause VWM disease.…”