The unfolded protein response and endoplasmic reticulum protein targeting machineries converge on the stress sensor IRE1.

Diego Acosta-Alvear,Han Li,Tobias C Walther,Peter Walter,G Elif Karagöz,Florian Fröhlich

doi:10.7554/elife.43036

Abstract

The protein folding capacity of the endoplasmic reticulum (ER) is tightly regulated by a network of signaling pathways, known as the unfolded protein response (UPR). UPR sensors monitor the ER folding status to adjust ER folding capacity according to need. To understand how the UPR sensor IRE1 maintains ER homeostasis, we identified zero-length crosslinks of RNA to IRE1 with single nucleotide precision in vivo. We found that IRE1 specifically crosslinks to a subset of ER-targeted mRNAs, SRP RNA, ribosomal and transfer RNAs. Crosslink sites cluster in a discrete region of the ribosome surface spanning from the A-site to the polypeptide exit tunnel. Moreover, IRE1 binds to purified 80S ribosomes with high affinity, indicating association with ER-bound ribosomes. Our results suggest that the ER protein translocation and targeting machineries work together with the UPR to tune the ER's protein folding load.

Highlights

Protein folding and maturation in the endoplasmic reticulum (ER) are essential for cell physiology, as most of the all secreted and transmembrane proteins are synthesized and folded in this organelle
By contrast to the XBP1 and BLOC1S1 mRNAs, which are processed by IRE1 with high efficiency, the PAR-CLIP captured mRNAs are cleaved by IRE1-KR at least one order-of-magnitude less efficiently, requiring longer incubation times and higher enzyme concentrations (Figure 2D and Peschek et al, 2015)
Among the transcripts found in the core-set of IRE1-interacting RNAs, we identified RN7SL, the RNA component of the signal recognition particle (SRP), as the most robust PAR-CLIP hit (Figure 2A)

Summary

Introduction

Protein folding and maturation in the endoplasmic reticulum (ER) are essential for cell physiology, as most of the all secreted and transmembrane proteins are synthesized and folded in this organelle. Perturbations leading to protein folding defects in the ER –collectively known as ER stress– activate an ensemble of transcriptional programs known as the unfolded protein response (UPR) (Karagoz and Acosta-Alvear D, 2018; Walter and Ron, 2011). Three ER membrane embedded protein folding sensors control the UPR: ATF6 (activating transcription factor 6), PERK (protein kinase R (PKR)-like kinase) and IRE1 (inositol requiring enzyme 1). Unfolded proteins serve as direct ligands for IRE1’s lumenal sensor domain, promoting its oligomerization and activation in the plane of the ER membrane (Aragon et al, 2009; Gardner and Walter, 2011; Karagoz et al, 2017; Kimata et al, 2007; Li et al, 2010). Active IRE1 responds to ER stress in two ways: (i) it cleaves an unconventional intron from the mRNA encoding the transcription factor XBP1 (X-box binding protein 1), initiating a spliceosome-independent mRNA splicing reaction that culminates in the production of XBP1s (‘s’ for spliced), a potent transcription activator

Methods

Results

Conclusion