bims-ershed Biomed News
on ER Stress in Health and Diseases
Issue of 2021‒06‒06
eleven papers selected by
Matías Eduardo González Quiroz
Worker’s Hospital

  1. Cell Death Discov. 2021 Jun 03. 7(1): 131
      Altered protein homeostasis is associated with neurodegenerative diseases and acute brain injury induced under energy depletion conditions such as ischemia. The accumulation of damaged or unfolded proteins triggers the unfolded protein response (UPR), which can act as a homeostatic response or lead to cell death. However, the factors involved in turning and adaptive response into a cell death mechanism are still not well understood. Several mechanisms leading to brain injury induced by severe hypoglycemia have been described but the contribution of the UPR has been poorly studied. Cell responses triggered during both the hypoglycemia and the glucose reinfusion periods can contribute to neuronal death. Therefore, we have investigated the activation dynamics of the PERK and the IRE1α branches of the UPR and their contribution to neuronal death in a model of glucose deprivation (GD) and glucose reintroduction (GR) in cortical neurons. Results show a rapid activation of the PERK/p-eIF2α/ATF4 pathway leading to protein synthesis inhibition during GD, which contributes to neuronal adaptation, however, sustained blockade of protein synthesis during GR promotes neuronal death. On the other hand, IRE1α activation occurs early during GD due to its interaction with BAK/BAX, while ASK1 is recruited to IRE1α activation complex during GR promoting the nuclear translocation of JNK and the upregulation of Chop. Most importantly, results show that IRE1α RNase activity towards its splicing target Xbp1 mRNA occurs late after GR, precluding a homeostatic role. Instead, IRE1α activity during GR drives neuronal death by positively regulating ASK1/JNK activity through the degradation of 14-3-3 θ mRNA, a negative regulator of ASK and an adaptor protein highly expressed in brain, implicated in neuroprotection. Collectively, results describe a novel regulatory mechanism of cell death in neurons, triggered by the downregulation of 14-3-3 θ mRNA induced by the IRE1α branch of the UPR.
  2. Int J Mol Sci. 2021 May 21. pii: 5436. [Epub ahead of print]22(11):
      The KDEL receptor retrieval pathway is essential for maintaining resident proteins in the endoplasmic reticulum (ER) lumen. ER resident proteins serve a variety of functions, including protein folding and maturation. Perturbations to the lumenal ER microenvironment, such as calcium depletion, can cause protein misfolding and activation of the unfolded protein response (UPR). Additionally, ER resident proteins are secreted from the cell by overwhelming the KDEL receptor retrieval pathway. Recent data show that KDEL receptors are also activated during the UPR through the IRE1/XBP1 signaling pathway as an adaptive response to cellular stress set forth to reduce the loss of ER resident proteins. This review will discuss the emerging connection between UPR activation and KDEL receptors as it pertains to ER proteostasis and disease states.
    Keywords:  ER resident proteins; KDEL receptor; disease; endoplasmic reticulum; exodosis; unfolded protein response
  3. Biochem Biophys Res Commun. 2021 May 28. pii: S0006-291X(21)00845-7. [Epub ahead of print]563 8-14
      Although accumulating evidence indicates participation of endoplasmic reticulum (ER) stress pathway or the unfolded protein response (UPR) in immunity, there still exists little information linking ER stress to regulatory T cells (Tregs). To evaluate the potential contribution of the UPR, we tested the effects of thapsigargin (TG), an ER stress inducer, on the function of Tregs. Here we reported that TG stimulation induced the up-regulation of the endoplasmic reticulum (ER)-stress chaperon Glucose-Regulated Protein 78 (GRP78), which is a master regulator of the UPR, the phosphorylation of eukaryotic initiation factor 2 alpha (elF2α) and the induction of activating transcription factor 4 (ATF4), which are both protein kinase R (PKR)-like ER kinase (PERK) downstream targets in Tregs. Simultaneously, we demonstrated that, under ER stress conditions, Tregs presented enhanced functional activity upon TCR stimulation, as illustrated with forkhead box transcription factor (Foxp3) expression, interleukin-10 (IL-10) and transforming growth factor-β (TGF-β) production and suppressive functional analysis. Notably, pretreatment with GSK2656157, a potent and selective PERK inhibitor, markedly diminished the TG-induced hyperresponsiveness of Tregs upon T cell antigen receptor (TCR) stimulation. Therefore, our findings illustrated the inter-connection and coordination of the evolutionarily conserved ER stress response and TCR signaling in Tregs and uncover a critical new role of the PERK branch of UPR in the regulation of Tregs function.
    Keywords:  ER stress; GRP78; PERK; Tregs; UPR
  4. Nature. 2021 Jun 02.
      Compartmentalization is a defining characteristic of eukaryotic cells, and partitions distinct biochemical processes into discrete subcellular locations. Microscopy1 and biochemical fractionation coupled with mass spectrometry2-4 have defined the proteomes of a variety of different organelles, but many intracellular compartments have remained refractory to such approaches. Proximity-dependent biotinylation techniques such as BioID provide an alternative approach to define the composition of cellular compartments in living cells5-7. Here we present a BioID-based map of a human cell on the basis of 192 subcellular markers, and define the intracellular locations of 4,145 unique proteins in HEK293 cells. Our localization predictions exceed the specificity of previous approaches, and enabled the discovery of proteins at the interface between the mitochondrial outer membrane and the endoplasmic reticulum that are crucial for mitochondrial homeostasis. On the basis of this dataset, we created as a community resource that provides online tools for localization analysis of user BioID data, and demonstrate how this resource can be used to understand BioID results better.
  5. Front Cell Dev Biol. 2021 ;9 669379
      Mitochondria are double membrane organelles in eukaryotic cells that provide energy by generating adenosine triphosphate (ATP) through oxidative phosphorylation. They are crucial to many aspects of cellular metabolism. Mitochondria contain their own DNA that encodes for essential proteins involved in the execution of normal mitochondrial functions. Compared with nuclear DNA, the mitochondrial DNA (mtDNA) is more prone to be affected by DNA damaging agents, and accumulated DNA damages may cause mitochondrial dysfunction and drive the pathogenesis of a variety of human diseases, including neurodegenerative disorders and cancer. Therefore, understanding better how mtDNA damages are repaired will facilitate developing therapeutic strategies. In this review, we focus on our current understanding of the mtDNA repair system. We also discuss other mitochondrial events promoted by excessive DNA damages and inefficient DNA repair, such as mitochondrial fusion, fission, and mitophagy, which serve as quality control events for clearing damaged mtDNA.
    Keywords:  DNA repair; mitochondrial DNA; mitochondrial fission; mitochondrial fusion; mitophagy
  6. Molecules. 2021 May 23. pii: 3113. [Epub ahead of print]26(11):
      Enzymes, receptors, and other binding molecules in biological processes can recognize enantiomers as different molecular entities, due to their different dissociation constants, leading to diverse responses in biological processes. Enantioselectivity can be observed in drugs pharmacodynamics and in pharmacokinetic (absorption, distribution, metabolism, and excretion), especially in metabolic profile and in toxicity mechanisms. The stereoisomers of a drug can undergo to different metabolic pathways due to different enzyme systems, resulting in different types and/or number of metabolites. The configuration of enantiomers can cause unexpected effects, related to changes as unidirectional or bidirectional inversion that can occur during pharmacokinetic processes. The choice of models for pharmacokinetic studies as well as the subsequent data interpretation must also be aware of genetic factors (such as polymorphic metabolic enzymes), sex, patient age, hepatic diseases, and drug interactions. Therefore, the pharmacokinetics and toxicity of a racemate or an enantiomerically pure drug are not equal and need to be studied. Enantioselective analytical methods are crucial to monitor pharmacokinetic events and for acquisition of accurate data to better understand the role of the stereochemistry in pharmacokinetics and toxicity. The complexity of merging the best enantioseparation conditions with the selected sample matrix and the intended goal of the analysis is a challenge task. The data gathered in this review intend to reinforce the importance of the enantioselectivity in pharmacokinetic processes and reunite innovative enantioselective analytical methods applied in pharmacokinetic studies. An assorted variety of methods are herein briefly discussed.
    Keywords:  chiral analyses; chiral stationary phases; chirality; enantiomers; metabolism
  7. DNA Repair (Amst). 2021 May 23. pii: S1568-7864(21)00096-3. [Epub ahead of print]104 103140
      A broad spectrum of spontaneous and genotoxin-induced DNA lesions impede replication fork progression. The DNA damage response that acts to promote completion of DNA replication is associated with dynamic changes in chromatin structure that include two distinct processes which operate genome-wide during S-phase. The first, often referred to as histone recycling or parental histone segregation, is characterized by the transfer of parental histones located ahead of replication forks onto nascent DNA. The second, known as de novo chromatin assembly, consists of the deposition of new histone molecules onto nascent DNA. Because these two processes occur at all replication forks, their potential to influence a multitude of DNA repair and DNA damage tolerance mechanisms is considerable. The purpose of this review is to provide a description of parental histone segregation and de novo chromatin assembly, and to illustrate how these processes influence cellular responses to DNA replication roadblocks.
    Keywords:  DNA repair; DNA replication; De novo chromatin assembly; Histone chaperones; Histone modifications
  8. FEBS J. 2021 Jun 05.
      Although the increase in the number of identified posttranslational modifications (PTMs) has substantially improved our knowledge about substrate site specificity of single PTMs, the fact that different types of PTMs can crosstalk and act in concert to exert important regulatory mechanisms for protein function has not gained much attention. Here, we show that PKCδ is SUMOylated at lysine 473 in its C-terminal catalytic domain, and the SUMOylation increases PKCδ stability by repressing its ubiquitination. In addition, we uncover a functional interplay between the phosphorylation and SUMOylation of PKCδ, which can strengthen each other through recruiting SUMO E2/E3 ligases and the PKCδ kinase, respectively, to the PKCδ complexes. We identified PIAS2β as the SUMO E3 ligase of PKCδ. More importantly, by enhancing PKCδ protein stability and its phosphorylation through an interdependent interplay of the PTMs, the SUMOylation of PKCδ promotes apoptotic cell death induced by H2 O2 . We conclude that SUMOylation represents an important regulatory mechanism of PKCδ PTMs for the kinase's function in oxidative cell damage.
    Keywords:  Apoptosis; Oxidative damage; PKCδ SUMOylation; PKCδ degradation; PKCδ phosphorylation
  9. DNA Repair (Amst). 2021 May 18. pii: S1568-7864(21)00093-8. [Epub ahead of print] 103137
      Recent studies have suggested that human RNA helicase, DDX3X, is important for DNA repair, but little is known about the nuclear activity of this protein. In vitro analysis of nuclear DDX3X interactions and localization with DNA damage pointed to a direct role for DDX3X in the DNA damage response. We aimed to investigate whether DDX3X plays a direct role in the DNA damage response in live cells. In order to track nuclear DDX3X, we generated a nuclear-export deficient DDX3X mutant construct and performed microirradiation in live cells. We found that DDX3X accumulates at sites of microirradiation shortly after DNA damage induction. We further found DDX3X recruitment to be mediated by its intrinsically disordered domains, similar to other RNA binding proteins that are recruited to sites of DNA damage. Inhibition of liquid-liquid phase separation also reduced DDX3X recruitment. CRISPR/Cas9-mediated knockout of PARP1 ablated DDX3X recruitment, which was restored upon transgenic expression of wild-type PARP1 but not catalytically inactive PARP1, suggesting that DDX3X recruitment is PARP1-dependent.
    Keywords:  DDX3X; DNA damage; RNA helicase
  10. Methods Mol Biol. 2021 ;2323 171-197
      The method development to analyze direct RNA-protein interaction is of high importance as not many homogeneous assay formats are available.The discovery of fluorescent light-up aptamers (FLAPs), short RNA aptamers that switch the fluorescence of small, cell-permeable, and nontoxic organic chromophores on, paved the road for their utilization in direct RNA -protein interactions. The combination with fluorescent proteins as biological fluorophores enabled the development of homogeneous assays that are in principle even encodable on genomic level.Here the rules and methods to design a homogeneous in vitro assay for the detection and quantification of a direct RNA -protein interaction will be described. The design and application of a homogeneous assay to observe and quantify the interaction of the Pseudomonas aeruginosa bacteriophage coat protein 7 (PP7) with its binding RNA sequence (pp7-RNA) will be shown. For this, the Spinach-DFHBI aptamer as RNA fusion and the red fluorescent mCherry as protein fusion was used.The methods presented here do not require any chemical modification of proteins or RNAs which make them relatively easy to use and to adopt on other systems. As all fluorophores are fusion tags to the according biomolecules, standard cloning strategies and molecular biology technologies are sufficient and make this method available for a broad community of researchers.
    Keywords:  DFHBI; Fluorescence Light-up Aptamers (FLAPs); Fluorescence quenching; Förster Resonance Energy Transfer (FRET ); PP7 protein; REMSA; RNA –protein interaction; Spinach aptamer; mCherry
  11. Neurobiol Aging. 2021 Apr 28. pii: S0197-4580(21)00133-0. [Epub ahead of print]105 137-147
      Alzheimer's disease (AD) is the prevailing form of dementia. Protein degradation and antioxidant pathways have a critical role in preventing the accumulation of protein aggregation; thus, failure of proteostasis in neurons along with redox imbalance mark AD. Herein, we exploited an AD Drosophila model expressing human amyloid precursor (hAPP) and beta-secretase 1 (hBACE1) proteins, to better understand the role of proteostatic or antioxidant pathways in AD. Ubiquitous expression of hAPP, hBACE1 in flies caused more severe degenerative phenotypes versus neuronal targeted expression; it also, suppressed proteasome activity, increased oxidative stress and significantly enhanced stress-sensitivity. Overexpression of Prosβ5 proteasomal subunit or Nrf2 transcription factor in AD Drosophila flies partially restored proteasomal activity but did not rescue hAPP, hBACE1 induced neurodegeneration. On the other hand, expression of autophagy-related Atg8a in AD flies decelerated neurodegeneration, increased stress-resistance, and improved flies' health-/lifespan. Overall, our data suggest that the noxious effects of amyloid-beta aggregates can be alleviated by enhanced autophagy, thus dietary or pharmacological interventions that target autophagy should be considered in AD therapeutic approaches.
    Keywords:  APP; Alzheimer's disease; Drosophila; Nrf2; autophagy; proteasome