bims-unfpre Biomed news
on Unfolded protein response
Issue of 2018‒12‒30
eight papers selected by
Susan Logue
Apoptosis Research Centre

  1. J Biol Chem. 2018 Dec 28. pii: jbc.RA118.006682. [Epub ahead of print]
    Wu CH, Silvers CR, Messing EM, Lee YF.
      The field cancerization effect has been proposed to explain bladder cancer's multifocal and recurrent nature, yet the mechanisms of this effect remain unknown. In this work, using cell biology, flow cytometry, and qPCR analyses, along with a xenograft mouse tumor model, we show that chronic exposure to tumor-derived extracellular vesicles (TEVs) results in the neoplastic transformation of non-malignant human SV-HUC urothelial cells. Inhibition of EV uptake prevented this transformation. Transformed cells not only possessed several oncogenic properties, such as increased genome instability, loss of cell-cell contact inhibition, and invasiveness, but also displayed altered morphology and cell structures, such as an enlarged cytoplasm with disrupted endoplasmic reticulum (ER) alignment and the accumulation of smaller mitochondria. Exposure of SV-HUC cells to TEVs provoked the unfolded protein response in the endoplasmic reticulum (UPR ER ). Prolonged induction of UPR ER signaling activated the survival branch of the UPR ER pathway, in which cells had elevated expression of inositol-requiring enzyme 1 (IRE1), NF-κB, and the inflammatory cytokine leptin, and incurred loss of the pro-apoptotic protein C/EBP homologous protein (CHOP). More importantly, inhibition of ER stress by docosahexaenoic acid prevented TEV-induced transformation. We propose that TEVs promote malignant transformation of predisposed cells by inhibiting pro-apoptotic signals and activating tumor-promoting ER stress-induced unfolded protein response and inflammation. This study provides detailed insight into the mechanisms underlying the bladder cancer field effect and tumor recurrence.
    Keywords:  bladder cancer; cell stress; cytokine; endoplasmic reticulum stress (ER stress); exosome (vesicle); extracellular vesicles; filed cancerization; inflammation; unfolded protein response (UPR); urothelial carconoma
  2. Elife. 2018 Dec 24. pii: e43036. [Epub ahead of print]7
    Acosta-Alvear D, Karagöz GE, Fröhlich F, Li H, Walther TC, Walter P.
      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.
    Keywords:  cell biology; human
  3. PLoS One. 2018 ;13(12): e0208993
    Liu C, Li X, Li C, Zhang Z, Gao X, Jia Z, Chen H, Jia Q, Zhao X, Liu J, Liu B, Xu Z, Tian Y, He K.
      Endoplasmic reticulum (ER) stress results from imbalances in unfolded/misfolded proteins, contributing to a wide variety of human diseases. To better understand the mechanisms involved in the cellular response to ER stress in cardiomyocytes, we previously conducted a genome-wide screening in an in vitro ER stress model of rat cardiomyocytes, which highlighted amino acid transporter heavy chain, member 2 (SLC3A2) as an important factor in ER stress. In the present study, we characterized the role of SLC3A2 during the unfolded protein response (UPR), as one of the primary pathways activated during ER stress. First, we confirmed the induction of Slc3a2 mRNA expression following treatment with various ER stress inducers in rat cardiomyocytes (H9C2) and neural cells (PC12). Knockdown of Slc3a2 expression with small interfering RNA (siRNA) revealed that the encoded protein functions upstream of three important UPR proteins: ATF4, ATF6, and XBP1. siRNA-mediated knockdown of both SLC3A2 and mammalian target of rapamycin 1 (mTOR1) revealed that mTOR1 acts as a mediator between SLC3A2 and the UPR. RNA sequencing was then performed to gain a more thorough understanding of the function of SLC3A2, which identified 23 highly differentially regulated genes between the control and knockdown cell lines, which were related to the UPR and amino acid transport. Notably, flow cytometry further showed that SLC3A2 inhibition also enhanced the apoptosis of rat cardiomyocytes. Taken together, these results highlight SLC3A2 as a complex, multifunctional signaling protein that acts upstream of well-known UPR proteins with anti-apoptotic properties, suggesting its potential as a therapeutic target for ER stress-related diseases.
  4. FEBS J. 2018 Dec 26.
    Park K, Lee SE, Shin KO, Uchida Y.
      Endoplasmic reticulum (ER) stress is a mechanism that allows to protect normal cellular functions in response to both internal perturbations, such as accumulation of unfolded proteins, and external perturbations, for example redox stress, UVB irradiation, and infection. A hallmark of ER stress is the accumulation of misfolded and unfolded proteins. Physiological levels of ER stress trigger the unfolded protein response (UPR) which is required to restore normal ER functions. However, the UPR can also initiate a cell death program/apoptosis pathway in response to excessive or persistent ER stress. Recently, it has become evident that chronic ER stress occurs in several diseases, including skin diseases like Darier's disease, rosacea, vitiligo, and melanoma; furthermore, it is suggested that ER stress is directly involved in the pathogenesis of these disorders. Here, we review the role of ER stress in skin function, and discuss its significance in skin diseases. This article is protected by copyright. All rights reserved.
    Keywords:  endoplasmic reticulum stress; skin disease; skin function; unfolded protein response
  5. Cell Death Discov. 2018 ;4 115
    Chidawanyika T, Sergison E, Cole M, Mark K, Supattapone S.
      Endoplasmic reticulum (ER) stress from accumulated misfolded proteins in the ER can activate the unfolded protein response (UPR). The UPR acts either to restore proteostasis or to activate cell death pathways if the stress cannot be resolved. The key downstream effectors in these pathways have been studied extensively. However, in comparison, stressor-specific key mediators are not as well characterized. In this study, we sought to identify and compare the genes that are necessary for cell death induced by three classic pharmacological ER stressors with different mechanisms of action: thapsigargin, tunicamycin, and brefeldin A. We conducted genome-wide CRISPR/Cas9-based loss-of-function screens against these agents in HAP1 cells, which are a near-haploid cell line. Our screens confirmed that MFSD2A and ARF4, which were identified in previous screens, are necessary for tunicamycin- and brefeldin A-induced cytotoxicity, respectively. We identified a novel gene, SEC24A, as an essential gene for thapsigargin-induced cytotoxicity in HAP1 cells. Further experiments showed that the ability of SEC24A to facilitate ER stress-induced cell death is specific to thapsigargin and that SEC24A acts upstream of the UPR. These findings show that the genes required for ER stress-induced cell death are specific to the agent used to induce ER stress and that the resident ER cargo receptor protein SEC24A is an essential mediator of thapsigargin-induced UPR and cell death.
  6. Int J Mol Sci. 2018 Dec 24. pii: E72. [Epub ahead of print]20(1):
    Wu MH, Lee CY, Huang TJ, Huang KY, Tang CH, Liu SH, Kuo KL, Kuan FC, Lin WC, Shi CS.
      Chondrosarcoma, a heterogeneous malignant bone tumor, commonly produces cartilage matrix, which generally has no response to conventional therapies. Studies have reported that MLN4924, a NEDD8-activating enzyme inhibitor, achieves antitumor effects against numerous malignancies. In this study, the suppressive effects of MLN4924 on human chondrosarcoma cell lines were investigated using in vitro and in vivo assays, which involved measuring cell viability, cytotoxicity, apoptosis, proliferation, cell cycles, molecule-associated cell cycles, apoptosis, endoplasmic reticulum (ER) stress, and tumor growth in a xenograft mouse model. Our results demonstrated that MLN4924 significantly suppressed cell viability, exhibited cytotoxicity, and stimulated apoptosis through the activation of caspase-3 and caspase-7 in chondrosarcoma cell lines. Furthermore, MLN4924 significantly inhibited cell proliferation by diminishing the phosphorylation of histone H3 to cause G2/M cell cycle arrest. In addition, MLN4924 activated ER stress⁻related apoptosis by upregulating the phosphorylation of c-Jun N-terminal kinase (JNK), enhancing the expression of GRP78 and CCAAT-enhancer-binding protein homologous protein (CHOP, an inducer of endoplasmic ER stress⁻related apoptosis) and activating the cleavage of caspase-4. Moreover, MLN4924 considerably inhibited the growth of chondrosarcoma tumors in a xenograft mouse model. Finally, MLN4924-mediated antichondrosarcoma properties can be accompanied by the stimulation of ER stress⁻related apoptosis, implying that targeting neddylation by MLN4924 is a novel therapeutic strategy for treating chondrosarcoma.
    Keywords:  MLN4924; NEDD8; chondrosarcoma; endoplasmic reticulum stress; neddylation; ubiquitin-proteasome system
  7. Onco Targets Ther. 2018 ;11 8995-9006
    Zhao H, Han L, Jian Y, Ma Y, Yan W, Chen X, Xu H, Li L.
      Background: Resveratrol is known as a natural phytoalexin found in grapes and wine, which has significant antitumor activity under in vitro and in vivo conditions. In recent years, great progress has been made in understanding the underlying mechanisms of resveratrol in inducing cellular apoptosis of melanoma cells. Our previous study has shown that the apoptosis regulation of resveratrol in melanoma cells was independent of activation of classical apoptosis-related protein p53.Materials and methods: MTT assay and 5-bromo-2'-deoxyuridine staining assay were used to analyze cell viability and proliferation. Immunofluorescence analysis of γ-H2AX was employed to clarify DNA damages. Annexin V-propidine iodide/fluorescein isothiocyanate assay was performed to evaluate the cell apoptosis. The mechanisms underlying the activation of M2-type pyruvate kinase (PKM2) by Erk1/2 to stabilize and maintain Bcl-2 signaling was investigated by subcellular fractionation analyses, immunofluorescence analysis, co-immunoprecipitation assay, ubiquitination assay, and glutathione S-transferase pull-down assay.
    Results: In the present study, we found that resveratrol dramatically inhibited melanoma cell proliferation and induced cell apoptosis through upregulation of p53 in a concentration-dependent manner. Conversely, p53 downregulation by short hairpin RNA couldn't rescue resveratrol-induced cell proliferation inhibition or apoptosis enlargement. Additionally, we found that resveratrol downregulated antiapoptotic protein Bcl-2 and activated Bax in the protein levels by promoting Bcl-2 degradation and cytochrome c release. Moreover, we discovered that PKM2, had a key role in cell apoptosis triggered by resveratrol through interacting with Bcl-2. Based on these results, we overexpressed PKM2 in melanoma cells and found that this prevented resveratrol-induced apoptosis by stabilizing the protein level of Bcl-2.
    Conclusion: Taken together, our results provided a novel mechanism accounting for the apoptosis induction of resveratrol in melanoma cells and suggested that downregulating Erk/PKM2/Bcl-2 axis appears to be a new approach for the prevention or treatment of melanoma.
    Keywords:  ER stress; antitumor; cytochrome c; ubiquitination
  8. Circ Res. 2018 Oct 19.
    Blackwood EA, Hofmann C, Santo Domingo M, Bilal AS, Sarakki A, Stauffer WT, Arrieta A, Thuerauf DJ, Kolkhorst FW, Müller OJ, Jakobi T, Dieterich C, Katus HA, Doroudgar S, Glembotski CC.
      RATIONALE: ER stress dysregulates ER proteostasis, which activates the transcription factor, ATF6, an inducer of genes that enhance protein folding and restore proteostasis. Due to increased protein synthesis, it is possible that protein folding and, thus, ER proteostasis are challenged during cardiac myocyte growth. However, it is not known whether ATF6 is activated, and if so, what its function is during hypertrophic growth of cardiac myocytes.OBJECTIVE: To examine the activity and function of ATF6 during cardiac hypertrophy.
    METHODS AND RESULTS: We found that ATF6 was activated and ATF6-target genes were induced in mice subjected to an acute model of trans-aortic constriction (TAC), or to free-wheel exercise, which promote adaptive cardiac myocyte hypertrophy with preserved cardiac function. Cardiac myocyte-specific deletion of Atf6 (ATF6 cKO) blunted TAC- and exercise-induced cardiac myocyte hypertrophy and impaired cardiac function, demonstrating a role for ATF6 in compensatory myocyte growth. Transcript profiling and chromatin immunoprecipitation identified RHEB as an ATF6-target gene in the heart. RHEB is an activator of mTORC1, a major inducer of protein synthesis and subsequent cell growth. Both TAC and exercise upregulated RHEB, activated mTORC1, and induced cardiac hypertrophy in WT mouse hearts, but not in ATF6 cKO hearts. Mechanistically, knockdown of ATF6 in neonatal rat ventricular myocytes blocked phenylephrine (PE)-, and insulin-like growth factor 1 (IGF1)-mediated Rheb induction, mTORC1 activation, and myocyte growth, all of which were restored by ectopic RHEB expression. Moreover, AAV9- RHEB restored cardiac growth to ATF6 cKO mice subjected to TAC. Finally, ATF6 induced RHEB in response to growth factors, but not in response to other activators of ATF6 that do not induce growth, indicating that ATF6 target gene induction is stress-specific.
    CONCLUSIONS: Compensatory cardiac hypertrophy activates ATF6, which induces Rheb and activates mTORC1. Thus, ATF6 is a previously unrecognized link between growth stimuli and mTORC1-mediated cardiac growth.
    Keywords:  ATF6; Rheb; cardiac protein folding; proteostasis