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Title: Post-transcriptional regulation of Xbp1 mRNA by Pumilio under ER stress

Speaker: Fátima Cairrão

Affiliation: Laboratory of Cell Signaling in Drosophila, ITQB NOVA

Abstract: 

Various environmental stresses and changes in physiological conditions can result in the accumulation of unfolded proteins in the endoplasmic reticulum (ER) lumen, a cellular condition named ER stress. To counteract ER stress and restore normal protein folding capacity, cells activate the unfolding protein response (UPR). Three signaling pathways are involved in the UPR activated by different ER transmembrane protein sensors: the inositol requiring enzyme 1 (IRE1a), the PKR-like ER kinase (PERK) and the activating transcription factor 6 (ATF6).

IRE1a plays a major role in the initiation of UPR through the activation of the downstream transcriptional factor XBP1. Homeostasis in cells during the Unfolded Protein Response (UPR) requires Xbp1 mRNA to be properly localized to the ER and modulated to respond adequately to changes in demand for protein folding and ER stress. We noted that the Drosophila Xbp1 3’UTR contains putative binding sites for the RNA binding protein Pumilio, matching the 8nt core consensus sequence previously described. We therefore hypothesized that Xbp1 mRNA may be a target for post-transcriptional regulation by Pumilio.

Our studies demonstrate that Pumilio stabilizes Xbp1spliced mRNA and its expression acting as a protector factor of Xbp1spliced mRNA stability. Also TAP-RNA affinity purification of Xbp1 mRNAs from adult drosophila eyes showed association with the RNA binding domain of Pumilio. Furthermore, Pumilio’s phosphorylation state affected Xbp1 regulation during the UPR. Indeed, Drosophila Pumilio showed an increase in phosphorylation during ER stress in S2 cells. Kinase assays using a recombinant human IRE1a and purified recombinant forms of Pumilio revealed that Ire1, which harbors a kinase moiety, mediates phosphorylation of both drosophila and human Pumilio proteins. These results suggest an evolutionarily conserved mechanism involving Ire1-mediated phosphorylation of Pumillio to regulate Xbp1 mRNA stability.

Fatima Cairrão, PhD
Lab. Cell Signaling in Drosophila, piso 4