scholarly journals ER stress signaling has an activating transcription factor 6α (ATF6)-dependent “off-switch”

2018 ◽  
Vol 293 (47) ◽  
pp. 18270-18284 ◽  
Author(s):  
Franziska Walter ◽  
Aisling O'Brien ◽  
Caoimhín G. Concannon ◽  
Heiko Düssmann ◽  
Jochen H. M. Prehn
Keyword(s):  
Transcription Factor ◽  
Er Stress ◽  
Neuroblastoma Cells ◽  
Stress Signaling ◽  
Signaling Proteins ◽  
Protein Levels ◽  
Activating Transcription Factor ◽  
Screening Platform ◽  
The Unfolded Protein Response ◽  
Activating Transcription Factor 6Α

In response to an accumulation of unfolded proteins in the endoplasmic reticulum (ER) lumen, three ER transmembrane signaling proteins, inositol-requiring enzyme 1 (IRE1), PRKR-like ER kinase (PERK), and activating transcription factor 6α (ATF6α), are activated. These proteins initiate a signaling and transcriptional network termed the unfolded protein response (UPR), which re-establishes cellular proteostasis. When this restoration fails, however, cells undergo apoptosis. To investigate cross-talk between these different UPR enzymes, here we developed a high-content live cell screening platform to image fluorescent UPR-reporter cell lines derived from human SH-SY5Y neuroblastoma cells in which different ER stress signaling proteins were silenced through lentivirus-delivered shRNA constructs. We observed that loss of ATF6 expression results in uncontrolled IRE1-reporter activity and increases X box–binding protein 1 (XBP1) splicing. Transient increases in both IRE1 mRNA and IRE1 protein levels were observed in response to ER stress, suggesting that IRE1 up-regulation is a general feature of ER stress signaling and was further increased in cells lacking ATF6 expression. Moreover, overexpression of the transcriptionally active N-terminal domain of ATF6 reversed the increases in IRE1 levels. Furthermore, inhibition of IRE1 kinase activity or of downstream JNK activity prevented an increase in IRE1 levels during ER stress, suggesting that IRE1 transcription is regulated through a positive feed-forward loop. Collectively, our results indicate that from the moment of activation, IRE1 signaling during ER stress has an ATF6-dependent “off-switch.”

Pancreatic β-cells depend on basal expression of active ATF6α-p50 for cell survival even under nonstress conditions

2012 ◽  
Vol 302 (7) ◽  
pp. C992-C1003 ◽  
Author(s):  
Tracy Teodoro ◽  
Tanya Odisho ◽  
Elena Sidorova ◽  
Allen Volchuk
Keyword(s):  
Endoplasmic Reticulum ◽  
Er Stress ◽  
Cell Survival ◽  
Protein Levels ◽  
Unfolded Protein ◽  
Basal Expression ◽  
Steady State Levels ◽  
Activating Transcription Factor ◽  
The Unfolded Protein Response ◽  
Insulinoma Cells

Activating transcription factor 6 (ATF6) is one of three principle endoplasmic reticulum (ER) stress response proteins and becomes activated when ER homeostasis is perturbed. ATF6 functions to increase ER capacity by stimulating transcription of ER-resident chaperone genes such as GRP78. Using an antibody that recognizes active ATF6α-p50, we found that active ATF6α was detected in insulinoma cells and rodent islets even under basal conditions and the levels were further increased by ER stress. To examine the function of ATF6α-p50, we depleted endogenous ATF6α-p50 levels using small interfering RNA in insulinoma cells. Knockdown of endogenous ATF6α-p50 levels by ∼60% resulted in a reduction in the steady-state levels of GRP78 mRNA and protein levels in nonstressed cells. Furthermore, ATF6α knockdown resulted in an apoptotic phenotype. We hypothesized that removal of the ATF6α branch of the unfolded protein response (UPR) would result in ER stress. However, neither the PKR-like endoplasmic reticulum kinase (PERK), nor the inositol requiring enzyme 1 (IRE1) pathways of the UPR were significantly activated in ATF6α knockdown cells, although these cells were more sensitive to ER stress-inducing compounds. Interestingly, phosphorylation of JNK, p38, and c-Jun were elevated in ATF6α knockdown cells and inhibition of JNK or p38 kinases prevented apoptosis. These results suggest that ATF6α may have a role in maintaining β-cell survival even in the absence of ER stress.

scholarly journals Divest Yourself of a Preconceived Idea: Transcription Factor ATF6 Is Not a Soluble Protein!

2010 ◽  
Vol 21 (9) ◽  
pp. 1435-1438 ◽  
Author(s):  
Kazutoshi Mori
Keyword(s):  
Transcription Factor ◽  
Er Stress ◽  
Transcriptional Activation ◽  
Intracellular Signaling ◽  
Leucine Zipper ◽  
Transmembrane Protein ◽  
Basic Leucine Zipper ◽  
Induction Program ◽  
Activating Transcription Factor ◽  
The Unfolded Protein Response

The unfolded protein response (UPR), an evolutionarily conserved transcriptional induction program that is coupled with intracellular signaling from the endoplasmic reticulum (ER) to the nucleus, is activated to cope with ER stress and to maintain the homeostasis of the ER. In 1996, we isolated a basic leucine zipper protein, which had been previously named activating transcription factor (ATF)6, as a candidate transcription factor responsible for the mammalian UPR. Subsequent analysis, however, was confounding. The problem was eventually tracked down to an unusual property of ATF6: rather than being a soluble nuclear protein, as expected for an active transcription factor, ATF6 was instead synthesized as a transmembrane protein embedded in the ER, which was activated by ER stress-induced proteolysis. ATF6 was thus unique: an ER stress sensor/transducer that is involved in all steps of the UPR, from the sensing step in the ER to the transcriptional activation step in the nucleus.

scholarly journals Sledgehammer to Scalpel: Broad Challenges to the Heart and Other Tissues Yield Specific Cellular Responses via Transcriptional Regulation of the ER-Stress Master Regulator ATF6α

2020 ◽  
Vol 21 (3) ◽  
pp. 1134 ◽  
Author(s):  
Winston T. Stauffer ◽  
Adrian Arrieta ◽  
Erik A. Blackwood ◽  
Christopher C. Glembotski
Keyword(s):  
Transcription Factor ◽  
Protein Folding ◽  
Er Stress ◽  
Molecular Mechanisms ◽  
Transmembrane Protein ◽  
Fine Tuning ◽  
Cellular Functions ◽  
Similar Mode ◽  
Activating Transcription Factor ◽  
Activating Transcription Factor 6Α

There are more than 2000 transcription factors in eukaryotes, many of which are subject to complex mechanisms fine-tuning their activity and their transcriptional programs to meet the vast array of conditions under which cells must adapt to thrive and survive. For example, conditions that impair protein folding in the endoplasmic reticulum (ER), sometimes called ER stress, elicit the relocation of the ER-transmembrane protein, activating transcription factor 6α (ATF6α), to the Golgi, where it is proteolytically cleaved. This generates a fragment of ATF6α that translocates to the nucleus, where it regulates numerous genes that restore ER protein-folding capacity but is degraded soon after. Thus, upon ER stress, ATF6α is converted from a stable, transmembrane protein, to a rapidly degraded, nuclear protein that is a potent transcription factor. This review focuses on the molecular mechanisms governing ATF6α location, activity, and stability, as well as the transcriptional programs ATF6α regulates, whether canonical genes that restore ER protein-folding or unexpected, non-canonical genes affecting cellular functions beyond the ER. Moreover, we will review fascinating roles for an ATF6α isoform, ATF6β, which has a similar mode of activation but, unlike ATF6α, is a long-lived, weak transcription factor that may moderate the genetic effects of ATF6α.

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 ◽  
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.

The effect of baicalein on endoplasmic reticulum stress and autophagy on liver damage

2021 ◽  
pp. 096032712110036
Author(s):  
MC Üstüner ◽  
C Tanrikut ◽  
D Üstüner ◽  
UK Kolaç ◽  
Z Özdemir Köroğlu ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Transcription Factor ◽  
Er Stress ◽  
Liver Damage ◽  
Albino Rats ◽  
Tem Analysis ◽  
Stress Markers ◽  
Activating Transcription Factor 4 ◽  
Activating Transcription Factor ◽  
Wistar Albino Rats

Carbon tetrachloride (CCl4) is a toxic chemical that causes liver injury. CCl4 triggers endoplasmic reticulum (ER) stress and unfolded protein response (UPR). UPR triggers autophagy to deal with the damage. The aim of this study was to investigate the effect of baicalein, derived from Scutellaria baicalensis, on CCl4-induced liver damage concerning ER stress and autophagy. Two groups of Wistar albino rats (n = 7/groups) were treated with 0.2 ml/kg CCl4 for 10 days with and without baicalein. Histological and transmission electron microscopy (TEM) analysis, autophagy, and ER stress markers measurements were carried out to evaluate the effect of baicalein. Histological examinations showed that baicalein reduced liver damage. TEM analysis indicated that baicalein inhibited ER stress and triggered autophagy. CCl4-induced elevation of C/EBP homologous protein (CHOP), glucose-regulating protein 78 (GRP78), activating transcription factor 4 (ATF4), activating transcription factor 6 (ATF6), inositol requiring enzyme 1 (IRE1), pancreatic ER kinase (PERK), and active/spliced form of X-box-binding protein 1 (XBP1s) ER stress markers were decreased by baicalein. Baicalein also increased the autophagy-related 5 (ATG5), Beclin1, and Microtubule-associated protein 1A/1B-light chain 3-phosphatidylethanolamine-conjugated form (LC3-II) autophagy marker levels. In conclusion, baicalein reduced the CCl4-induced liver damage by inhibiting ER stress and the trigger of autophagy.

scholarly journals Mammalian ECD Protein Is a Novel Negative Regulator of the PERK Arm of the Unfolded Protein Response

2017 ◽  
Vol 37 (18) ◽  
Author(s):  
Appolinaire A. Olou ◽  
Aniruddha Sarkar ◽  
Aditya Bele ◽  
C. B. Gurumurthy ◽  
Riyaz A. Mir ◽  
...  
Keyword(s):  
Er Stress ◽  
Translation Initiation Factor ◽  
Negative Regulator ◽  
Initiation Factor ◽  
Mrna Levels ◽  
Produce Cell ◽  
Stress Induction ◽  
Content Type ◽  
Protein Levels ◽  
The Unfolded Protein Response

ABSTRACT Mammalian Ecdysoneless (ECD) is a highly conserved ortholog of the Drosophila Ecd gene product whose mutations impair the synthesis of Ecdysone and produce cell-autonomous survival defects, but the mechanisms by which ECD functions are largely unknown. Here we present evidence that ECD regulates the endoplasmic reticulum (ER) stress response. ER stress induction led to a reduced ECD protein level, but this effect was not seen in PKR-like ER kinase knockout (PERK-KO) or phosphodeficient eukaryotic translation initiation factor 2α (eIF2α) mouse embryonic fibroblasts (MEFs); moreover, ECD mRNA levels were increased, suggesting impaired ECD translation as the mechanism for reduced protein levels. ECD colocalizes and coimmunoprecipitates with PERK and GRP78. ECD depletion increased the levels of both phospho-PERK (p-PERK) and p-eIF2α, and these effects were enhanced upon ER stress induction. Reciprocally, overexpression of ECD led to marked decreases in p-PERK, p-eIF2α, and ATF4 levels but robust increases in GRP78 protein levels. However, GRP78 mRNA levels were unchanged, suggesting a posttranscriptional event. Knockdown of GRP78 reversed the attenuating effect of ECD overexpression on PERK signaling. Significantly, overexpression of ECD provided a survival advantage to cells upon ER stress induction. Taken together, our data demonstrate that ECD promotes survival upon ER stress by increasing GRP78 protein levels to enhance the adaptive folding protein in the ER to attenuate PERK signaling.

scholarly journals Activating Transcription Factor 4 Is Translationally Regulated by Hypoxic Stress

2004 ◽  
Vol 24 (17) ◽  
pp. 7469-7482 ◽  
Author(s):  
Jaime D. Blais ◽  
Vasilisa Filipenko ◽  
Meixia Bi ◽  
Heather P. Harding ◽  
David Ron ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Microarray Analysis ◽  
Initiation Factor ◽  
Hypoxic Stress ◽  
Unfolded Protein ◽  
Activating Transcription Factor 4 ◽  
Activating Transcription Factor ◽  
The Unfolded Protein Response ◽  
Protein Response ◽  
Transcription Factor 4

ABSTRACT Hypoxic stress results in a rapid and sustained inhibition of protein synthesis that is at least partially mediated by eukaryotic initiation factor 2α (eIF2α) phosphorylation by the endoplasmic reticulum (ER) kinase PERK. Here we show through microarray analysis of polysome-bound RNA in aerobic and hypoxic HeLa cells that a subset of transcripts are preferentially translated during hypoxia, including activating transcription factor 4 (ATF4), an important mediator of the unfolded protein response. Changes in mRNA translation during the unfolded protein response are mediated by PERK phosphorylation of the translation initiation factor eIF2α at Ser-51. Similarly, PERK is activated and is responsible for translational regulation under hypoxic conditions, while inducing the translation of ATF4. The overexpression of a C-terminal fragment of GADD34 that constitutively dephosphorylates eIF2α was able to attenuate the phosphorylation of eIF2α and severely inhibit the induction of ATF4 in response to hypoxic stress. These studies demonstrate the essential role of ATF4 in the response to hypoxic stress, define the pathway for its induction, and reveal that GADD34, a target of ATF4 activation, negatively regulates the eIF2α-mediated inhibition of translation. Taken with the concomitant induction of additional ER-resident proteins identified by our microarray analysis, this study suggests an important integrated response between ER signaling and the cellular adaptation to hypoxic stress.

scholarly journals Role of Activating Transcription Factor 4 in Murine Choroidal Neovascularization Model

2021 ◽  
Vol 22 (16) ◽  
pp. 8890
Author(s):  
Hiroto Yasuda ◽  
Miruto Tanaka ◽  
Anri Nishinaka ◽  
Shinsuke Nakamura ◽  
Masamitsu Shimazawa ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Transcription Factor ◽  
Er Stress ◽  
Choroidal Neovascularization ◽  
Retinal Pigment ◽  
Age Related Macular Degeneration ◽  
Activating Transcription Factor 4 ◽  
Age Related ◽  
Activating Transcription Factor ◽  
Transcription Factor 4

Neovascular age-related macular degeneration (nAMD) featuring choroidal neovascularization (CNV) is the principal cause of irreversible blindness in elderly people in the world. Integrated stress response (ISR) is one of the intracellular signals to be adapted to various stress conditions including endoplasmic reticulum (ER) stress. ISR signaling results in the upregulation of activating transcription factor 4 (ATF4), which is a mediator of ISR. Although recent studies have suggested ISR contributes to the progression of some age-related disorders, the effects of ATF4 on the development of CNV remain unclear. Here, we performed a murine model of laser-induced CNV and found that ATF4 was highly expressed in endothelial cells of the blood vessels of the CNV lesion site. Exposure to integrated stress inhibitor (ISRIB) reduced CNV formation, vascular leakage, and the upregulation of vascular endothelial growth factor (VEGF) in retinal pigment epithelium (RPE)-choroid-sclera complex. In human retinal microvascular endothelial cells (HRMECs), ISRIB reduced the level of ATF4 and VEGF induced by an ER stress inducer, thapsigargin, and recombinant human VEGF. Moreover, ISRIB decreased the VEGF-induced cell proliferation and migration of HRMECs. Collectively, our findings showed that pro-angiogenic effects of ATF4 in endothelial cells may be a potentially therapeutic target for patients with nAMD.

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