An RNA zipper promotes the unfolded protein response

Cells adjust the protein folding capacity of the endoplasmic reticulum by a network of stress signaling pathways known as the unfolded protein response (UPR). The most conserved branch of the UPR relies on the cytoplasmic splicing of a specific mRNA. We identified an RNA zipper mechanism as a unique and conserved feature promoting this non-conventional splicing mechanism.

HFSP Long-Term Fellow Jirka Peschek and colleagues
authored on Fri, 04 December 2015

Most transmembrane and secretory proteins are modified, folded and assembled in the endoplasmic reticulum (ER). When the ER-folding machinery becomes overwhelmed, misfolded polypeptides accumulate, leading to ER-stress. The unfolded protein response (UPR) is a network of intracellular signaling pathways that adjusts the protein folding capacity of the ER according to the demands of the cell. The most conserved branch is mediated by the transmembrane sensor/transducer protein IRE1. ER stress activates its RNase domain, which initiates the UPR signal by a unique mechanism: in a non-conventional, cytoplasmic splicing reaction, IRE1 removes an intron from XBP1 mRNA followed by exon-exon ligation by tRNA ligase, allowing the production of the XBP1 transcription factor that drives the transcriptional response. How the splicing reaction, requiring two cleavage events and the subsequent ligation of the correct exon ends, is orchestrated with fidelity remains an outstanding question.

To study the IRE1-mediated splicing of XBP1 mRNA, we reconstituted the reaction biochemically in vitro. We found that cleavage of the RNA substrate by IRE1 at the splice sites resulted in autonomous removal of the intron while the exons remained base-paired. Guided by computational analyses, we identified a unique feature of the RNA: conserved base‐pairing of the exons serves as a zipper‐like mechanism to bring the correct ends of the cleaved XBP1 mRNA together for subsequent ligation. Furthermore, this RNA‐intrinsic conformational change promotes intron‐ejection and is important to complete the splicing reaction by tRNA ligase. Finally, we could show that exon zippering also ensures the efficiency and fidelity of XBP1 splicing in living cells.

Figure: A conformational RNA zipper promotes the IRE1-mediated non-conventional splicing of XBP1 mRNA. The unspliced (left) and spliced (right) RNA sequences proximal to the splice sites are depicted in the conserved secondary structures. After cleavage by the RNase domain of IRE1, the exons zipper up to (i) eject the intron (red) and to (ii) place the correct ends in proximity for subsequent ligation by tRNA ligase. The graphic is taken from the original reference. © 2015 The Authors. Published under the terms of the CC BY NC ND 4.0 license.

The UPR is a key stress response in eukaryotic cells that can determine the cell’s fate in a life-or-death decision. In order to understand how the UPR functions in normal cells and how it may be rewired in diseases, such as cancer or diabetes, a detailed mechanistic understanding of its signaling reactions is fundamentally important. We have uncovered an evolutionarily conserved structural RNA rearrangement within the mRNA substrate of the IRE1 branch of the UPR. Intriguingly, the intron is not just a passive bystander in the proposed splicing mechanism but undergoes active and essential conformational changes.

Reference

A conformational RNA zipper promotes intron ejection during non-conventional XBP1 mRNA splicing. Peschek J, Acosta-Alvear D, Mendez AS, Walter P. EMBO Reports (2015), DOI: 10.15252/embr.201540955.

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