scholarly journals The Down-Regulation of Clusterin Expression Enhances the αSynuclein Aggregation Process

2020 ◽  
Vol 21 (19) ◽  
pp. 7181 ◽  
Author(s):  
Chiara Lenzi ◽  
Ileana Ramazzina ◽  
Isabella Russo ◽  
Alice Filippini ◽  
Saverio Bettuzzi ◽  
...  
Keyword(s):  
Proteasome Inhibition ◽  
Strong Increase ◽  
Aggregation Process ◽  
Down Regulation ◽  
Unfolded Protein ◽  
Aβ Aggregation ◽  
Important Player ◽  
The Unfolded Protein Response ◽  
Protein Response ◽  
Clusterin Expression

Parkinson’s Disease (PD) is a progressive neurodegenerative disease characterized by the presence of proteinaceous aggregates of αSynuclein (αSyn) in the dopaminergic neurons. Chaperones are key components of the proteostasis network that are able to counteract αSyn’s aggregation, as well as its toxic effects. Clusterin (CLU), a molecular chaperone, was consistently found to interfere with Aβ aggregation in Alzheimer’s Disease (AD). However, its role in PD pathogenesis has yet to be extensively investigated. In this study, we assessed the involvement of CLU in the αSyn aggregation process by using SH-SY5Y cells stably overexpressing αSyn (SH-Syn). First, we showed that αSyn overexpression caused a strong increase in CLU expression without affecting levels of Hsp27, Hsp70, and Hsp90, which are the chaperones widely recognized to counteract αSyn burden. Then, we demonstrated that αSyn aggregation, induced by proteasome inhibition, determines a strong increase of CLU in insoluble aggregates. Remarkably, we revealed that CLU down-regulation results in an increase of αSyn aggregates in SH-Syn without significantly affecting cell viability and the Unfolded Protein Response (UPR). Furthermore, we demonstrated the direct molecular interaction between CLU and αSyn via a co-immunoprecipitation (co-IP) assay. All together, these findings provide incontrovertible evidence that CLU is an important player in the response orchestrated by the cell to cope with αSyn burden.

Activation of ATF4 mediates unwanted Mcl-1 accumulation by proteasome inhibition

Blood ◽  
2012 ◽  
Vol 119 (3) ◽  
pp. 826-837 ◽  
Author(s):  
Jinsong Hu ◽  
Nana Dang ◽  
Eline Menu ◽  
Elke De Bryune ◽  
Dehui Xu ◽  
...  
Keyword(s):  
Myeloid Cell ◽  
Proteasome Inhibition ◽  
Endoplasmic Reticulum Stress Response ◽  
Unfolded Protein ◽  
Activating Transcription Factor 4 ◽  
Activating Transcription Factor ◽  
Induced Apoptosis ◽  
The Unfolded Protein Response ◽  
Protein Response ◽  
Transcription Factor 4

Abstract Myeloid cell leukemia-1 (Mcl-1) protein is an anti-apoptotic Bcl-2 family protein that plays essential roles in multiple myeloma (MM) survival and drug resistance. In MM, it has been demonstrated that proteasome inhibition can trigger the accumulation of Mcl-1, which has been shown to confer MM cell resistance to bortezomib-induced lethality. However, the mechanisms involved in this unwanted Mcl-1 accumulation are still unclear. The aim of the present study was to determine whether the unwanted Mcl-1 accumulation could be induced by the unfolded protein response (UPR) and to elucidate the role of the endoplasmic reticulum stress response in regulating Mcl-1 expression. Using quantitative RT-PCR and Western blot, we found that the translation of activating transcription factor-4 (ATF4), an important effector of the UPR, was also greatly enhanced by proteasome inhibition. ChIP analysis further revealed that bortezomib stimulated binding of ATF4 to a regulatory site (at position −332 to −324) at the promoter of the Mcl-1 gene. Knocking down ATF4 was paralleled by down-regulation of Mcl-1 induction by bortezomib and significantly increased bortezomib-induced apoptosis. These data identify the UPR and, more specifically, its ATF4 branch as an important mechanism mediating up-regulation of Mcl-1 by proteasome inhibition.

scholarly journals Mutational analysis of the human Xbp1 translational arrest peptide and construction of arrest-enhanced variants

2017 ◽  
Author(s):  
Nina Schiller ◽  
Anastasia Magoulopoulou ◽  
Florian Cymer ◽  
Gunnar Von Heijne
Keyword(s):  
Mutational Analysis ◽  
Strong Increase ◽  
E Coli ◽  
Unfolded Protein ◽  
Nascent Chain ◽  
Translational Arrest ◽  
Critical Residues ◽  
The Unfolded Protein Response ◽  
Natural Amino Acids ◽  
Protein Response

AbstractXbp1, a protein involved in the unfolded protein response, is a rare example of a mammalian protein that contains a well-defined translational arrest peptide (AP). In order to define the critical residues in the Xbp1u AP, and to search for variants with stronger arrest potency than the wildtype Xbp1u AP, we have carried out a full mutagenesis scan where each residue in the AP was replaced by the other 19 natural amino acids. We find that 10 of the 21 mutagenized positions are optimal already in the wildtype Xbp1 AP, while certain mutations in the remaining residues lead to a strong increase in the arrest potency. Xbp1 has thus evolved to induce an intermediate level of translational arrest, and versions with much stronger arrest efficiency exist. We further show Xbp1-induced translational arrest is reduced in response to increased tension in the nascent chain, making it possible to carry out studies in mammalian systems of cotranslational processes such as membrane protein assembly and protein folding by using suitable Xbp1 AP variants as “force sensors”, as has been done previously in E. coli using bacterial APs.

scholarly journals Different Induction of GRP78 and CHOP as a Predictor of Sensitivity to Proteasome Inhibitors in Thyroid Cancer Cells

Endocrinology ◽  
2007 ◽  
Vol 148 (7) ◽  
pp. 3258-3270 ◽  
Author(s):  
Hua-Qin Wang ◽  
Zhen-Xian Du ◽  
Hai-Yan Zhang ◽  
Da-Xin Gao
Keyword(s):  
Thyroid Cancer ◽  
Cancer Cells ◽  
Cell Lines ◽  
Proteasome Inhibitors ◽  
Proteasome Inhibition ◽  
Sensitive Cell ◽  
Unfolded Protein ◽  
The Unfolded Protein Response ◽  
Protein Response ◽  
Thyroid Cancer Cells

Proteasome inhibitors represent a novel class of antitumor agents with preclinical and clinical evidence of activity against hematological malignancies and solid tumors. Emerging lines of evidence suggest that the unfolded protein response is implicated in proteasome inhibitors-induced apoptosis. Glucose-regulated protein 78 kDa (GRP78) and CCAAT/enhancer-binding protein homologous protein (CHOP) as part of the unfolded protein response play critical roles in cell survival or death. Here we demonstrate that induction of GRP78 and CHOP are differently regulated upon proteasome inhibition in different thyroid cancer cell lines, and GRP78 levels as well as preferential induction of GRP78 or CHOP appears to be involved in the responsiveness. Insensitive ARO, 8305C, and 8505C cell lines inherently express relatively high levels of GRP78 compared with sensitive cell lines, and its levels are further up-regulated upon treatment with proteasome inhibitors. CHOP levels are dramatically induced in sensitive cell lines until 24 h after proteasome inhibition. On the other hand, only a slight increase is observed at 4 h in insensitive cell lines, and this increase is unable to be detected after 8 h. Insensitive cells are sensitized to proteasome inhibition by suppression of GRP78. Furthermore, suppression of CHOP induction or overexpression of GRP78 partially prevents proteasome inhibition-mediated cell death. Our study indicates a molecular mechanism by which the sensitivity of thyroid cancer cells is regulated by the level of GRP78 as well as preferential induction of GRP78 or CHOP upon treatment with proteasome inhibitors. Our experiments therefore suggest a novel approach toward sensitization of thyroid cancer cells to proteasome inhibitors.

scholarly journals Translational and posttranslational regulation of XIAP by eIF2α and ATF4 promotes ER stress–induced cell death during the unfolded protein response

2014 ◽  
Vol 25 (9) ◽  
pp. 1411-1420 ◽  
Author(s):  
Nobuhiko Hiramatsu ◽  
Carissa Messah ◽  
Jaeseok Han ◽  
Matthew M. LaVail ◽  
Randal J. Kaufman ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Cell Death ◽  
Er Stress ◽  
Unfolded Protein Response ◽  
Protein Misfolding ◽  
Down Regulation ◽  
Unfolded Protein ◽  
Activity Loss ◽  
The Unfolded Protein Response ◽  
Protein Response

Endoplasmic reticulum (ER) protein misfolding activates the unfolded protein response (UPR) to help cells cope with ER stress. If ER homeostasis is not restored, UPR promotes cell death. The mechanisms of UPR-mediated cell death are poorly understood. The PKR-like endoplasmic reticulum kinase (PERK) arm of the UPR is implicated in ER stress–induced cell death, in part through up-regulation of proapoptotic CCAAT/enhancer binding protein homologous protein (CHOP). Chop−/− cells are partially resistant to ER stress–induced cell death, and CHOP overexpression alone does not induce cell death. These findings suggest that additional mechanisms regulate cell death downstream of PERK. Here we find dramatic suppression of antiapoptosis XIAP proteins in response to chronic ER stress. We find that PERK down-regulates XIAP synthesis through eIF2α and promotes XIAP degradation through ATF4. Of interest, PERK's down-regulation of XIAP occurs independently of CHOP activity. Loss of XIAP leads to increased cell death, whereas XIAP overexpression significantly enhances resistance to ER stress–induced cell death, even in the absence of CHOP. Our findings define a novel signaling circuit between PERK and XIAP that operates in parallel with PERK to CHOP induction to influence cell survival during ER stress. We propose a “two-hit” model of ER stress–induced cell death involving concomitant CHOP up-regulation and XIAP down-regulation both induced by PERK.

scholarly journals Targeting Multiple Myeloma through the Biology of Long-Lived Plasma Cells

Cancers ◽  
2020 ◽  
Vol 12 (8) ◽  
pp. 2117 ◽  
Author(s):  
Adam Utley ◽  
Brittany Lipchick ◽  
Kelvin P. Lee ◽  
Mikhail A. Nikiforov
Keyword(s):  
Multiple Myeloma ◽  
Bone Marrow ◽  
Plasma Cells ◽  
Standard Of Care ◽  
Proteasome Inhibition ◽  
Unfolded Protein ◽  
Survival Pathways ◽  
Disease Induction ◽  
The Unfolded Protein Response ◽  
Protein Response

Multiple myeloma (MM) is a hematological malignancy of terminally differentiated bone marrow (BM) resident B lymphocytes known as plasma cells (PC). PC that reside in the bone marrow include a distinct population of long-lived plasma cells (LLPC) that have the capacity to live for very long periods of time (decades in the human population). LLPC biology is critical for understanding MM disease induction and progression because MM shares many of the same extrinsic and intrinsic survival programs as LLPC. Extrinsic survival signals required for LLPC survival include soluble factors and cellular partners in the bone marrow microenvironment. Intrinsic programs that enhance cellular fidelity are also required for LLPC survival including increased autophagy, metabolic fitness, the unfolded protein response (UPR), and enhanced responsiveness to endoplasmic reticulum (ER) stress. Targeting LLPC cell survival mechanisms have led to standard of care treatments for MM including proteasome inhibition (Bortezomib), steroids (Dexamethasone), and immunomodulatory drugs (Lenalidomide). MM patients that relapse often do so by circumventing LLPC survival pathways targeted by treatment. Understanding the mechanisms by which LLPC are able to survive can allow us insight into the treatment of MM, which allows for the enhancement of therapeutic strategies in MM both at diagnosis and upon patient relapse.

scholarly journals BET Inhibitors Synergize with Carfilzomib to Induce Cell Death in Cancer Cells via Impairing Nrf1 Transcriptional Activity and Exacerbating the Unfolded Protein Response

Biomolecules ◽  
2020 ◽  
Vol 10 (4) ◽  
pp. 501 ◽  
Author(s):  
Janakiram R. Vangala ◽  
Ajay Potluri ◽  
Senthil K. Radhakrishnan
Keyword(s):  
Unfolded Protein Response ◽  
Solid Tumors ◽  
Cell Lines ◽  
Proteasome Inhibitors ◽  
Proteasome Inhibition ◽  
Therapeutic Modality ◽  
Unfolded Protein ◽  
Bet Inhibitors ◽  
The Unfolded Protein Response ◽  
Protein Response

Currently, proteasome inhibitors bortezomib, carfilzomib, and ixazomib are successfully used in clinics to treat multiple myeloma. However, these agents show limited efficacy against solid tumors. Identification of drugs that can potentiate the action of proteasome inhibitors could help expand the use of this therapeutic modality to solid tumors. Here, we found that bromodomain extra-terminal (BET) family protein inhibitors such as JQ1, I-BET762, and I-BET151 synergize with carfilzomib in multiple solid tumor cell lines. Mechanistically, BET inhibitors attenuated the ability of the transcription factor Nrf1 to induce proteasome genes in response to proteasome inhibition, thus, impeding the bounce-back response of proteasome activity, a critical pathway by which cells cope with proteotoxic stress. Moreover, we found that treatment with BET inhibitors or depletion of Nrf1 exacerbated the unfolded protein response (UPR), signaling that was initiated by proteasome inhibition. Taken together, our work provides a mechanistic explanation behind the synergy between proteasome and BET inhibitors in cancer cell lines and could prompt future preclinical and clinical studies aimed at further investigating this combination.

scholarly journals Disruption of ceramide synthesis by CerS2 down-regulation leads to autophagy and the unfolded protein response

2009 ◽  
Vol 424 (2) ◽  
pp. 273-283 ◽  
Author(s):  
Stefka D. Spassieva ◽  
Thomas D. Mullen ◽  
Danyelle M. Townsend ◽  
Lina M. Obeid
Keyword(s):  
Unfolded Protein Response ◽  
Stress Responses ◽  
Apoptotic Cell ◽  
Sphingolipid Metabolism ◽  
Ceramide Synthase ◽  
Down Regulation ◽  
Unfolded Protein ◽  
The Unfolded Protein Response ◽  
Protein Response ◽  
Long Chain

Ceramide metabolism has come under recent scrutiny because of its role in cellular stress responses. CerS2 (ceramide synthase 2) is one of the six mammalian isoforms of ceramide synthase and is responsible for the synthesis of VLC (very-long-chain) ceramides, e.g. C24, C24:1. To study the role of CerS2 in ceramide metabolism and cellular homoeostasis, we down-regulated CerS2 using siRNA (small interfering RNA) and examined several aspects of sphingolipid metabolism and cell stress responses. CerS2 down-regulation had a broad effect on ceramide homoeostasis, not just on VLC ceramides. Surprisingly, CerS2 down-regulation resulted in significantly increased LC (long-chain) ceramides, e.g. C14, C16, and our results suggested that the increase was due to a ceramide synthase-independent mechanism. CerS2-down-regulation-induced LC ceramide accumulation resulted in growth arrest which was not accompanied by apoptotic cell death. Instead, cells remained viable, showing induction of autophagy and activation of PERK [PKR (double-stranded-RNA-dependent protein kinase)-like endoplasmic reticulum kinase] and IRE1 (inositol-requiring 1) pathways [the latter indicating activation of the UPR (unfolded protein response)].

scholarly journals Dysfunction of the unfolded protein response increases neurodegeneration in aged rat hippocampus following proteasome inhibition

Aging Cell ◽  
2009 ◽  
Vol 8 (6) ◽  
pp. 654-665 ◽  
Author(s):  
María Paz Gavilán ◽  
Cristina Pintado ◽  
Elena Gavilán ◽  
Sebastián Jiménez ◽  
Rosa M. Ríos ◽  
...  
Keyword(s):  
Unfolded Protein Response ◽  
Proteasome Inhibition ◽  
Rat Hippocampus ◽  
Aged Rat ◽  
Unfolded Protein ◽  
The Unfolded Protein Response ◽  
Protein Response

Oestrogen-driven changes in the bovine uterus are associated with activation of the unfolded protein response

2014 ◽  
Author(s):  
Mohammed A Alfattah ◽  
Paul Anthony McGettigan ◽  
John Arthur Browne ◽  
Khalid M Alkhodair ◽  
Katarzyna Pluta ◽  
...  
Keyword(s):  
Unfolded Protein Response ◽  
Unfolded Protein ◽  
The Unfolded Protein Response ◽  
Protein Response

Native Ubiquitin Structural Changes Resulting from Complexation with β-methylamino-L-alanine (BMAA)

2020 ◽  
Author(s):  
Katie Mae Wilson ◽  
Aurora Burkus-Matesevac ◽  
Samuel Maddox ◽  
Christopher Chouinard
Keyword(s):  
Structural Changes ◽  
Noncovalent Complex ◽  
Unfolded Protein ◽  
Protein Size ◽  
High Concentrations ◽  
The Unfolded Protein Response ◽  
Critical Binding ◽  
Protein Response ◽  
The Brain ◽  
Lateral Sclerosis

β-methylamino-L-alanine (BMAA) has been linked to the development of neurodegenerative (ND) symptoms following chronic environmental exposure through water and dietary sources. The brains of those affected by this condition, often referred to as amyotrophic lateral sclerosis-parkinsonism-dementia complex (ALS-PDC), have exhibited the presence of plaques and neurofibrillary tangles (NFTs) from protein aggregation. Although numerous studies have sought to better understand the correlation between BMAA exposure and onset of ND symptoms, no definitive link has been identified. One prevailing hypothesis is that BMAA acts a small molecule ligand, complexing with critical proteins in the brain and reducing their function. The objective of this research was to investigate the effects of BMAA exposure on the native structure of ubiquitin. We hypothesized that formation of a Ubiquitin+BMAA noncovalent complex would alter the protein’s structure and folding and ultimately affect the ubiquitinproteasome system (UPS) and the unfolded protein response (UPR). Ion mobility-mass spectrometry revealed that at sufficiently high concentrations BMAA did in fact form a noncovalent complex with ubiquitin, however similar complexes were identified for a range of additional amino acids. Collision induced unfolding (CIU) was used to interrogate the unfolding dynamics of native ubiquitin and these Ubq-amino acid complexes and it was determined that complexation with BMAA led to a significant alteration in native protein size and conformation, and this complex required considerably more energy to unfold. This indicates that the complex remains more stable under native conditions and this may indicate that BMAA has attached to a critical binding location.

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