Pumilio protects Xbp1 mRNA from regulated Ire1-dependent decay

SUMMARYThe unfolded protein response (UPR) maintains homeostasis of the endoplasmic reticulum(ER). Residing in the ER membrane, the UPR mediator Ire1 deploys its cytoplasmic kinase-endoribonuclease domain to activate the key UPR transcription factor Xbp1 through non-conventional splicing of Xbp1 mRNA. Ire1 also degrades diverse ER-targeted mRNAs through regulated Ire1-dependent decay (RIDD), but how it spares Xbp1 mRNA from this decay is unknown. We identified binding sites for the RNA-binding protein Pumilio in the 3’UTR Drosophila Xbp1. In the developing Drosophila eye, Pumilio bound both the Xbp1unspliced and Xbp1spliced mRNAs, but only Xbp1spliced was stabilized by Pumilio. Furthermore, Pumilio displayed Ire1 kinase-dependent phosphorylation during ER stress, which was required for its stabilization of Xbp1spliced. Human IRE1 could directly phosphorylate Pumilio, and phosphorylated Pumilio protected Xbp1spliced mRNA against RIDD. Thus, Ire1-mediated phosphorylation enables Pumilio to shield Xbp1spliced from RIDD. These results uncover an important and unexpected regulatory link between an RNA-binding protein and the UPR.

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Staufen 1 amplifies pro-apoptotic activation of the unfolded protein response

10.1101/820225 ◽

2019 ◽

Author(s):

Mandi Gandelman ◽

Warunee Dansithong ◽

Karla P Figueroa ◽

Sharan Paul ◽

Daniel R Scoles ◽

...

Keyword(s):

Unfolded Protein Response ◽

Cortical Neurons ◽

Rna Binding ◽

Binding Protein ◽

Unfolded Protein ◽

Functional Consequences ◽

Exogenous Expression ◽

The Unfolded Protein Response ◽

Protein Response

AbstractStaufen-1 (STAU1) is an RNA binding protein that becomes highly overabundant in numerous neurodegenerative disease models, including those carrying mutations in presenilin1 (PSEN1), microtubule associated protein tau (MAPT), huntingtin (HTT), TAR DNA-binding protein-43 gene (TARDBP) or C9orf72. We previously reported that elevations in STAU1 determine autophagy defects. Additional functional consequences of STAU1 overabundance, however, have not been investigated. We studied the role of STAU1 in the chronic activation of the Unfolded Protein Response (UPR), a common feature among the neurodegenerative diseases where STAU1 is increased, and is directly associated with neuronal death. Here we report that STAU1 is a novel modulator of the UPR, and is required for apoptosis induced by activation of the PERK-CHOP pathway. STAU1 levels increased in response to multiple ER stressors and exogenous expression of STAU1 was sufficient to cause apoptosis through the PERK-CHOP pathway of the UPR. Cortical neurons and skin fibroblasts derived from Stau1−/− mice showed reduced UPR and apoptosis when challenged with thapsigargin. In fibroblasts from SCA2 patients or with ALS-causing TDP-43 and C9ORF72 mutations we found highly increased STAU1 and CHOP levels in basal conditions. STAU1 knockdown restored CHOP levels to normal. Taken together, these results show STAU1 overabundance reduces cellular resistance to ER stress and precipitates apoptosis.

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Induction of the FK506-binding protein, FKBP13, under conditions which misfold proteins in the endoplasmic reticulum

Biochemical Journal ◽

10.1042/bj3030705 ◽

1994 ◽

Vol 303 (3) ◽

pp. 705-708 ◽

Cited By ~ 42

Author(s):

K T Bush ◽

B A Hendrickson ◽

S K Nigam

Keyword(s):

Endoplasmic Reticulum ◽

Mdck Cells ◽

Binding Protein ◽

Significant Similarity ◽

Fk506 Binding Protein ◽

Unfolded Protein ◽

Immunoglobulin Binding Protein ◽

The Unfolded Protein Response ◽

Protein Response ◽

Immunoglobulin Binding

In order to determine whether the endoplasmic reticulum (ER) luminal FK506-binding protein, FKBP13, shares properties of ER molecular chaperones, MDCK cells were treated with either tunicamycin or Ca2+ ionophores. By Northern-blot analysis, tunicamycin resulted in a 2-fold rise in FKBP13 mRNA, whereas ionophores (A23187 and ionomycin) caused a more impressive rise in FKBP13 mRNA (up to 5-fold with ionomycin). Actinomycin D chase experiments in ionomycin-treated cells revealed no change in the half-life of FKBP13 mRNA, indicating that the increase in FKBP13 mRNA observed was not due to greater message stability. Moreover, sequencing of the 5′ flanking region of the gene for murine FKBP13 revealed significant similarity to similar regions in human BiP (immunoglobulin-binding protein) and the human glucose-regulated protein grp94, including a 37 bp sequence in FKBP13 with approximately 50% identity with the unfolded protein response element of the BiP gene. Together, these data suggest a role for FKBP13 in ER protein folding.

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Endoplasmic reticulum stress and the unfolded protein response are linked to synergistic IFN-β inductionvia X-box binding protein 1

European Journal of Immunology ◽

10.1002/eji.200737882 ◽

2008 ◽

Vol 38 (5) ◽

pp. 1194-1203 ◽

Cited By ~ 123

Author(s):

Judith A. Smith ◽

Matthew J. Turner ◽

Monica L. DeLay ◽

Erin I. Klenk ◽

Dawn P. Sowders ◽

...

Keyword(s):

Endoplasmic Reticulum ◽

Endoplasmic Reticulum Stress ◽

Unfolded Protein Response ◽

Binding Protein ◽

Unfolded Protein ◽

The Unfolded Protein Response ◽

Protein Response

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Staufen 1 amplifies proapoptotic activation of the unfolded protein response

Cell Death and Differentiation ◽

10.1038/s41418-020-0553-9 ◽

2020 ◽

Vol 27 (10) ◽

pp. 2942-2951 ◽

Cited By ~ 1

Author(s):

Mandi Gandelman ◽

Warunee Dansithong ◽

Karla P. Figueroa ◽

Sharan Paul ◽

Daniel R. Scoles ◽

...

Keyword(s):

Neurodegenerative Diseases ◽

Unfolded Protein Response ◽

Cortical Neurons ◽

Rna Binding ◽

Binding Protein ◽

Unfolded Protein ◽

Functional Consequences ◽

Exogenous Expression ◽

The Unfolded Protein Response ◽

Protein Response

AbstractStaufen-1 (STAU1) is an RNA-binding protein that becomes highly overabundant in numerous neurodegenerative disease models, including those carrying mutations in presenilin1 (PSEN1), microtubule-associated protein tau (MAPT), huntingtin (HTT), TAR DNA-binding protein-43 gene (TARDBP), or C9orf72. We previously reported that elevations in STAU1 determine autophagy defects and its knockdown is protective in models of several neurodegenerative diseases. Additional functional consequences of STAU1 overabundance, however, have not been investigated. We studied the role of STAU1 in the chronic activation of the unfolded protein response (UPR), a common feature among neurodegenerative diseases and often directly associated with neuronal death. Here we report that STAU1 is a novel modulator of the UPR, and is required for apoptosis induced by activation of the PERK–CHOP pathway. STAU1 levels increased in response to multiple endoplasmic reticulum (ER) stressors, and exogenous expression of STAU1 was sufficient to cause apoptosis through the PERK–CHOP pathway of the UPR. Cortical neurons and skin fibroblasts derived from Stau1−/− mice showed reduced UPR and apoptosis when challenged with thapsigargin. In fibroblasts from individuals with SCA2 or with ALS-causing TDP-43 and C9ORF72 mutations, we found highly increased STAU1 and CHOP levels in basal conditions, and STAU1 knockdown restored CHOP levels to normal. Taken together, these results show that STAU1 overabundance reduces cellular resistance to ER stress and precipitates apoptosis.

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Quercetin Affects Hsp70/IRE1αMediated Protection from Death Induced by Endoplasmic Reticulum Stress

Oxidative Medicine and Cellular Longevity ◽

10.1155/2015/645157 ◽

2015 ◽

Vol 2015 ◽

pp. 1-11 ◽

Cited By ~ 19

Author(s):

Antonello Storniolo ◽

Marisa Raciti ◽

Alessandra Cucina ◽

Mariano Bizzarri ◽

Livia Di Renzo

Keyword(s):

Endoplasmic Reticulum ◽

Binding Protein ◽

Human Leukemia ◽

Unfolded Protein ◽

Homologous Protein ◽

Dietary Flavonoid ◽

Induced Apoptosis ◽

The Unfolded Protein Response ◽

Shock Proteins ◽

Protein Response

Relative to their normal counterparts, tumor cells generally exhibit a greater “stress phenotype” and express heat shock proteins (Hsp) that represent candidate targets for anticancer therapy. Here we investigated the role of Hsp70 in survival induced by endoplasmic reticulum (ER) stressors in human leukemia U937 cells. Quercetin, a major dietary flavonoid, or specific silencing affected the expression level of Hsp70 and did not allow the upregulation of inositol-requiring kinase 1α(IRE1α), the prototype ER stress sensor regulating the unfolded protein response (UPR), that protects the cells against the stress of misfolded proteins in the ER. The reduction of Hsp70 prevented the upregulation of immunoglobulin heavy-chain binding protein (BiP), but not of CCAAT/enhancer-binding protein-homologous protein (CHOP), and induced apoptosis. Also specific silencing of IRE1αor inhibition of its endoribonuclease activity by 4μ8c hampered the upregulation of BiP, but not of CHOP, and induced apoptosis. These results suggest that drugs affecting the Hsp70-IRE1αaxis, like quercetin, or affecting directly IRE1αmay represent an effective adjuvant antileukemia therapy.

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Enhanced Biosynthesis of Coagulation Factor VIII through Diminished Engagement of the Unfolded Protein Response

Journal of Biological Chemistry ◽

10.1074/jbc.m111.238758 ◽

2011 ◽

Vol 286 (27) ◽

pp. 24451-24457 ◽

Cited By ~ 24

Author(s):

Harrison C. Brown ◽

Bagirath Gangadharan ◽

Christopher B. Doering

Keyword(s):

Endoplasmic Reticulum ◽

Factor Viii ◽

Unfolded Protein Response ◽

Binding Protein ◽

Coagulation Factor ◽

Coagulation Factor Viii ◽

Luciferase Reporter ◽

Unfolded Protein ◽

The Unfolded Protein Response ◽

Protein Response

Human and porcine coagulation factor VIII (fVIII) display a biosynthetic efficiency differential that is being exploited for the development of new protein and gene transfer-based therapies for hemophilia A. The cellular and/or molecular mechanism(s) responsible for this phenomenon have yet to be uncovered, although it has been temporally localized to post-translational biosynthetic steps. The unfolded protein response (UPR) is a cellular adaptation to structurally distinct (e.g. misfolded) or excess protein in the endoplasmic reticulum and is known to be induced by heterologous expression of recombinant human fVIII. Therefore, it is plausible that the biosynthetic differential between human and porcine fVIII results from differential UPR activation. In the current study, UPR induction was examined in the context of ongoing fVIII expression. UPR activation was greater during human fVIII expression when compared with porcine fVIII expression as determined by ER response element (ERSE)-luciferase reporter activity, X-box-binding protein 1 (XBP1) splicing, and immunoglobulin-binding protein (BiP) up-regulation. Immunofluorescence microscopy of fVIII expressing cells revealed that human fVIII was notably absent in the Golgi apparatus, confirming that endoplasmic reticulum to Golgi transport is rate-limiting. In contrast, a significant proportion of porcine fVIII was localized to the Golgi indicating efficient transit through the secretory pathway. Overexpression of BiP, an integral UPR protein, reduced the secretion of human fVIII by 50%, but had no effect on porcine fVIII biosynthesis. In contrast, expression of BiP shRNA increased human fVIII expression levels. The current data support the model of differential engagement of UPR by human and porcine fVIII as a non-traditional mechanism for regulation of gene product biosynthesis.

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The Unfolded Protein Response (UPR) Is Activated in Human Acute Myeloid Leukemia (AML) and Suppresses Translation of the CCAAT/Enhancer Binding Protein-Alpha (CEBPA) by Induction of the RNA-Binding Protein Calreticulin

Blood ◽

10.1182/blood.v112.11.2934.2934 ◽

2008 ◽

Vol 112 (11) ◽

pp. 2934-2934

Author(s):

Julian Schardt ◽

Marianne Eyholzer ◽

Beatrice U Mueller ◽

Thomas Pabst

Keyword(s):

Transcription Factor ◽

Er Stress ◽

Rna Binding ◽

Binding Protein ◽

Leukemic Cells ◽

Protein Levels ◽

Unfolded Protein ◽

The Unfolded Protein Response ◽

Protein Response ◽

Spliced Variant

Abstract Deregulation of the myeloid key transcription factor CCAAT/enhancer binding protein alpha (CEBPA) is a common event in AML patients. We previously reported that the RNA-binding protein calreticulin efficiently blocks CEBPA translation and is specifically induced in core binding factor (CBF) leukemias. In addition, calreticulin is a crucial component of the unfolded protein response (UPR) which is triggered by the accumulation of misfolded proteins within the endoplasmic reticulum (ER). In vitro studies suggested that the UPR is activated in some solid cancers and thereby involved in tumor development. The role of the UPR during leukemogenesis has not been addressed so far. Here, we investigated the induction of the spliced variant of the X-box binding protein 1 (XBP1s) as a marker for activated ER stress and determined the expression of key mediators of the UPR such as calreticulin and the 78-kDa glucose-regulated protein (GRP78) in leukemic cells from 92 consecutive AML patients. Increased expression of the XBP1 spliced variant was detected in 16 of 92 AML patients. Consistently, this group also had increased mRNA and protein levels of calreticulin and GRP78. In patients expressing the XBP1 spliced variant, CEBPA protein was hardly detectable in contrast to AML patients not expressing the XBP1 spliced variant. Moreover, treatment of myeloid leukemic cells with compounds activating the UPR – such as thapsigargin - confirmed rapid induction of XBP1s, GRP78 and calreticulin in myeloid cells whereas CEBPA protein levels decreased. In addition, conditional expression of calreticulin in U937 cells suppressed CEBPA protein. At the molecular level, we identified two functional ER stress response elements (ERSE) in the calreticulin promoter, and both elements were found to be necessary for full induction of calreticulin following ER stress. The presence of the tripartite nuclear transcription factor Y (NFY) and activating transcription factor 6 (ATF6), as well as an intact binding site for the YY1 transcription factor (YY1) within these ERSE motifs appeared to be crucial for mediating sensitivity to ER stress. Finally, chromatin-immunoprecipitation assays indicated that binding of NFY and YY1 to the ERSE motifs is induced after induction of ER stress in vivo. Therefore, we conclude that the UPR is activated in a subgroup of AML patients. Activation of the UPR involves induction of calreticulin expression by the ATF6 pathway and ultimately leads to suppressed CEBPA translation, thus contributing to the block in myeloid differentiation in these leukemias.

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