Coping with the calcium overload caused by cell injury: ER to the rescue

Cells maintain their cytosolic calcium (Ca2+) in nanomolar range and use controlled increase in Ca2+ for intracellular signaling. With the extracellular Ca2+ in the millimolar range, there is a steep Ca2+ gradient across the plasma membrane (PM). Thus, injury that damages PM, leads to a cytosolic Ca2+ overload, which helps activate PM repair (PMR) response. However, in order to survive, the cells must cope with the Ca2+ overload. In a recent study (Chandra et al. J Cell Biol,doi: 10.1083/jcb.202006035) we have examined how cells cope with injury-induced cytosolic Ca2+ overload. By monitoring Ca2+ dynamics in the cytosol and endoplasmic reticulum (ER), we found that PM injury-triggered increase in cytosolic Ca2+ is taken up by the ER. Pharmacological inhibition of ER Ca2+ uptake interferes with this process and compromises the repair ability of the injured cells. Muscle cells from patients and mouse model for the muscular dystrophy showed that lack of Anoctamin 5 (ANO5)/Transmembrane protein 16E (TMEM16E), an ER-resident putative Ca2+-activated chloride channel (CaCC), are poor at coping with cytosolic Ca2+ overload. Pharmacological inhibition of CaCC and lack of ANO5, both prevent Ca2+ uptake into ER. These studies identify a requirement of Cl– uptake by the ER in sequestering injury-triggered cytosolic Ca2+ increase in the ER. Further, these studies show that ER helps injured cells cope with Ca2+ overload during PMR, lack of which contributes to muscular dystrophy due to mutations in the ANO5 protein.

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Hepatitis C Virus Subgenomic Replicons Induce Endoplasmic Reticulum Stress Activating an Intracellular Signaling Pathway

Journal of Virology ◽

10.1128/jvi.76.15.7453-7459.2002 ◽

2002 ◽

Vol 76 (15) ◽

pp. 7453-7459 ◽

Cited By ~ 186

Author(s):

Keith D. Tardif ◽

Kazutoshi Mori ◽

Aleem Siddiqui

Keyword(s):

Endoplasmic Reticulum ◽

Hepatitis C Virus ◽

Hepatitis C ◽

Er Stress ◽

Signaling Pathway ◽

Intracellular Signaling ◽

Transmembrane Protein ◽

Initiation Factor ◽

Α Subunit ◽

Intracellular Signaling Pathway

ABSTRACT Hepatitis C virus (HCV) replicates from a ribonucleoprotein (RNP) complex that is associated with the endoplasmic reticulum (ER) membrane. The replication activities of the HCV subgenomic replicon are shown here to induce ER stress. In response to this stress, cells expressing HCV replicons induce the unfolded protein response (UPR), an ER-to-nucleus intracellular signaling pathway. The UPR is initiated by the proteolytic cleavage of a transmembrane protein, ATF6. The resulting cytoplasmic protein fragment of ATF6 functions as a transcription factor in the nucleus and activates selective genes required for an ER stress response. ATF6 activation leads to increased transcriptional levels of GRP78, an ER luminal chaperone protein. However, the overall level of GRP78 protein is decreased. While ER stress is also known to affect translational attenuation, cells expressing HCV replicons have lower levels of phosphorylation of the α subunit of eukaryotic initiation factor 2. Interestingly, cap-independent internal ribosome entry site-mediated translation directed by the 5′ noncoding region of HCV and GRP78 is activated in cells expressing HCV replicons. These studies provide insight into the effects of HCV replication on intracellular events and the mechanisms underlying liver pathogenesis.

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Maturation of the Na,K-ATPase in the Endoplasmic Reticulum in Health and Disease

The Journal of Membrane Biology ◽

10.1007/s00232-021-00184-z ◽

2021 ◽

Author(s):

Vitalii Kryvenko ◽

Olga Vagin ◽

Laura A. Dada ◽

Jacob I. Sznajder ◽

István Vadász

Keyword(s):

Endoplasmic Reticulum ◽

Intracellular Signaling ◽

Loss Of Function ◽

Α Subunit ◽

Rate Limiting Step ◽

Atpase Subunits ◽

A Cell ◽

Health And Disease ◽

And Function ◽

Rate Limiting

Abstract The Na,K-ATPase establishes the electrochemical gradient of cells by driving an active exchange of Na+ and K+ ions while consuming ATP. The minimal functional transporter consists of a catalytic α-subunit and a β-subunit with chaperon activity. The Na,K-ATPase also functions as a cell adhesion molecule and participates in various intracellular signaling pathways. The maturation and trafficking of the Na,K-ATPase include co- and post-translational processing of the enzyme in the endoplasmic reticulum (ER) and the Golgi apparatus and subsequent delivery to the plasma membrane (PM). The ER folding of the enzyme is considered as the rate-limiting step in the membrane delivery of the protein. It has been demonstrated that only assembled Na,K-ATPase α:β-complexes may exit the organelle, whereas unassembled, misfolded or unfolded subunits are retained in the ER and are subsequently degraded. Loss of function of the Na,K-ATPase has been associated with lung, heart, kidney and neurological disorders. Recently, it has been shown that ER dysfunction, in particular, alterations in the homeostasis of the organelle, as well as impaired ER-resident chaperone activity may impede folding of Na,K-ATPase subunits, thus decreasing the abundance and function of the enzyme at the PM. Here, we summarize our current understanding on maturation and subsequent processing of the Na,K-ATPase in the ER under physiological and pathophysiological conditions. Graphic Abstract

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ER Reorganization is Remarkably Induced in COS-7 Cells Accumulating Transmembrane Protein Receptors Not Competent for Export from the Endoplasmic Reticulum

The Journal of Membrane Biology ◽

10.1007/s00232-014-9710-8 ◽

2014 ◽

Vol 247 (11) ◽

pp. 1149-1159 ◽

Cited By ~ 10

Author(s):

Massimo D’Agostino ◽

Arianna Crespi ◽

Elena Polishchuk ◽

Serena Generoso ◽

Gianluca Martire ◽

...

Keyword(s):

Endoplasmic Reticulum ◽

Transmembrane Protein ◽

Protein Receptors

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The role of endoplasmic reticulum calcium pumps during cytosolic calcium spiking in pancreatic acinar cells.

Journal of Biological Chemistry ◽

10.1016/s0021-9258(18)41521-9 ◽

1993 ◽

Vol 268 (30) ◽

pp. 22262-22264

Author(s):

C.C. Petersen ◽

O.H. Petersen ◽

M.J. Berridge

Keyword(s):

Endoplasmic Reticulum ◽

Acinar Cells ◽

Cytosolic Calcium ◽

Pancreatic Acinar Cells ◽

Endoplasmic Reticulum Calcium ◽

Calcium Pumps ◽

Calcium Spiking

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Protection against hydrogen peroxide cytotoxicity in Rat-1 fibroblasts provided by the oncoprotein Bcl-2: maintenance of calcium homoeostasis is secondary to the effect of Bcl-2 on cellular glutathione

Biochemical Journal ◽

10.1042/bj3400291 ◽

1999 ◽

Vol 340 (1) ◽

pp. 291-297 ◽

Cited By ~ 15

Author(s):

Matthäus M. RIMPLER ◽

Ursula RAUEN ◽

Thorsten SCHMIDT ◽

Tarik MÖRÖY ◽

Herbert DE GROOT

Keyword(s):

Cell Death ◽

Calcium Concentration ◽

Cell Injury ◽

Cytosolic Calcium ◽

Glutathione Metabolism ◽

Transfected Cells ◽

Cytosolic Calcium Concentration ◽

Glutathione Status ◽

Calcium Elevation ◽

Reduced State

The oncoprotein Bcl-2 protects cells against apoptosis, but the exact molecular mechanism that underlies this function has not yet been identified. Studying H2O2-induced cell injury in Rat-1 fibroblast cells, we observed that Bcl-2 had a protective effect against the increase in cytosolic calcium concentration and subsequent cell death. Furthermore, overexpression of Bcl-2 resulted in an alteration of cellular glutathione status: the total amount of cellular glutathione was increased by about 60% and the redox potential of the cellular glutathione pool was maintained in a more reduced state during H2O2 exposure compared with non-Bcl-2-expressing controls. In our cytotoxicity model, disruption of cellular glutathione homoeostasis closely correlated with the pathological elevation of cytosolic calcium concentration. Stabilization of the glutathione pool by Bcl-2, N-acetylcysteine or glucose delayed the cytosolic calcium increase and subsequent cell death, whereas depletion of glutathione by DL-buthionine-(S,R)-sulphoximine, sensitized Bcl-2-transfected cells towards cytosolic calcium increase and cell death. We therefore suggest that the protection exerted by Bcl-2 against H2O2-induced cytosolic calcium elevation and subsequent cell death is secondary to its effect on the cellular glutathione metabolism.

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Structure-Function Analysis of the Endoplasmic Reticulum Oxidoreductase TMX3 Reveals Interdomain Stabilization of the N-terminal Redox-active Domain

Journal of Biological Chemistry ◽

10.1074/jbc.m706442200 ◽

2007 ◽

Vol 282 (46) ◽

pp. 33859-33867 ◽

Cited By ~ 16

Author(s):

Johannes Haugstetter ◽

Michael Andreas Maurer ◽

Thomas Blicher ◽

Martin Pagac ◽

Gerhard Wider ◽

...

Keyword(s):

Endoplasmic Reticulum ◽

Protein Disulfide Isomerase ◽

Function Analysis ◽

Transmembrane Protein ◽

Reduced Form ◽

Disulfide Isomerase ◽

Binding Domains ◽

Redox Active ◽

Protein Disulfide ◽

Protein Disulfide Isomerase Family

Disulfide bond formation in the endoplasmic reticulum is catalyzed by enzymes of the protein disulfide-isomerase family that harbor one or more thioredoxin-like domains. We recently discovered the transmembrane protein TMX3, a thiol-disulfide oxidoreductase of the protein disulfide-isomerase family. Here, we show that the endoplasmic reticulum-luminal region of TMX3 contains three thioredoxin-like domains, an N-terminal redox-active domain (named a) followed by two enzymatically inactive domains (b and b′). Using the recombinantly expressed TMX3 domain constructs a, ab, and abb′, we compared structural stability and enzymatic properties. By structural and biophysical methods, we demonstrate that the reduced a domain has features typical of a globular folded domain that is, however, greatly destabilized upon oxidization. Importantly, interdomain stabilization by the b domain renders the a domain more resistant toward chemical denaturation and proteolysis in both the oxidized and reduced form. In combination with molecular modeling studies of TMX3 abb′, the experimental results provide a new understanding of the relationship between the multidomain structure of TMX3 and its function as a redox enzyme. Overall, the data indicate that in addition to their role as substrate and co-factor binding domains, redox-inactive thioredoxin-like domains also function in stabilizing neighboring redox-active domains.

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An essential dimer-forming subregion of the endoplasmic reticulum stress sensor Ire1

Biochemical Journal ◽

10.1042/bj20050640 ◽

2005 ◽

Vol 391 (1) ◽

pp. 135-142 ◽

Cited By ~ 27

Author(s):

Daisuke Oikawa ◽

Yukio Kimata ◽

Masato Takeuchi ◽

Kenji Kohno

Keyword(s):

Endoplasmic Reticulum ◽

Recombinant Protein ◽

Endoplasmic Reticulum Stress ◽

Transmembrane Protein ◽

Type I ◽

Unfolded Protein ◽

The Unfolded Protein Response ◽

Protein Response ◽

Recombinant Fragments ◽

Made In

The luminal domain of the type I transmembrane protein Ire1 senses endoplasmic reticulum stress by an undefined mechanism to up-regulate the signalling pathway for the unfolded protein response. Previously, we proposed that the luminal domain of yeast Ire1 is divided into five subregions, termed subregions I–V sequentially from the N-terminus. Ire1 lost activity when internal deletions of subregion II or IV were made. In the present paper, we show that partial proteolysis of a recombinant protein consisting of the Ire1 luminal domain suggests that subregions II–IV are tightly folded. We also show that a recombinant protein of subregions II–IV formed homodimers, and that this homodimer formation was impaired by an internal deletion of subregion IV. Furthermore, recombinant fragments of subregion IV exhibited a self-binding ability. Therefore, although its sequence is little conserved evolutionarily, subregion IV plays an essential role to promote Ire1 dimer formation.

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EPA and DHA confer protection against DON-induced endoplasmic reticulum stress and iron imbalance in IPEC-1 cells

British Journal Of Nutrition ◽

10.1017/s0007114521003688 ◽

2021 ◽

pp. 1-29

Author(s):

Jia Lin ◽

Feifei Huang ◽

Tianzeng Liang ◽

Qin Qin ◽

Qiao Xu ◽

...

Keyword(s):

Endoplasmic Reticulum ◽

Er Stress ◽

Rna Sequencing ◽

Cell Injury ◽

Cell Damage ◽

Binding Protein ◽

Sequencing Analysis ◽

Epa And Dha ◽

Expression Of Genes ◽

Transferrin Receptor 1

Abstract This study assessed the molecular mechanism of eicosapentaenoic acid (EPA) or docosahexaenoic acid (DHA) protection against IPEC-1 cell damage induced by deoxynivalenol (DON). The cells were divided into six groups, including the CON group, the EPA group, the DHA group, the DON group, the EPA+DON group, and the DHA+DON group. RNA sequencing was used to investigate the potential mechanism, and qRT-PCR was employed to verify the expression of selected genes. Changes in ultrastructure were used to estimate pathological changes and endoplasmic reticulum (ER) injury in IPEC-1 cells. Transferrin receptor 1 (TFR1) was tested by ELISA. Fe2+ and malondialdehyde (MDA) contents were estimated by spectrophotometry, and reactive oxygen species (ROS) was assayed by fluorospectrophotometry. RNA sequencing analysis showed that EPA and DHA had a significant effect on the expression of genes involved in ER stress and iron balance during DON-induced cell injury. The results showed that DON increased ER damage, the content of MDA and ROS, the ratio of X-box binding protein 1s (XBP-1s)/X-box binding protein 1u (XBP-1u), the concentration of Fe2+, and the activity of TFR1. However, the results also showed that EPA and DHA decreased the ratio of XBP-1s/XBP-1u to relieve DON-induced ER damage of IPEC-1 cells. Moreover, EPA and DHA (especially DHA) reversed the factors related to iron balance. It can be concluded that EPA and DHA reversed IPEC-1 cell damage induced by DON. DHA has the potential to protect IPEC-1 cells from DON-induced iron imbalance by inhibiting ER stress.

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Morphology of Cell Injury: An Approach to the EDIT Programme by the Use of Tobacco Pollen Tubes

Alternatives to Laboratory Animals ◽

10.1177/026119290203000310 ◽

2002 ◽

Vol 30 (3) ◽

pp. 323-329 ◽

Cited By ~ 1

Author(s):

Udo Kristen ◽

Natalie Bischoff ◽

Saskia Lisboa ◽

Enno Schirmer ◽

Sören Witt ◽

...

Keyword(s):

Tip Growth ◽

Cell Injury ◽

Cytosolic Calcium ◽

Pollen Tubes ◽

In Vitro Cytotoxicity ◽

Toxic Compounds ◽

In Vitro System ◽

Tobacco Pollen ◽

Different Types

Tobacco pollen tubes were used as a standard in vitro system to investigate cell growth aberrations caused by some of the Multicentre Evaluation of In Vitro Cytotoxicity (MEIC) programme chemicals and other toxic compounds. Changes in cytoskeletal pattern were observed in the tube cells by using tubu-lin immunofluorescence and rhodamin–phalloidin fluorescence for the localisation of microtubules and actin filaments, respectively. Four different types of cell malformation were found: screw-like growth, isodiametric tip swelling, hook formation, and pollen grain enlargement. We suggest that these malformations resulted from an interference by the chemicals with the cytosolic calcium gradient which controls tip growth and the orientation of the pollen tube. The results may contribute to a general understanding of toxicity-based cell malformations.

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