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

2002 ◽  
Vol 76 (15) ◽  
pp. 7453-7459 ◽  
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.

scholarly journals Protein Synthesis and Endoplasmic Reticulum Stress Can Be Modulated by the Hepatitis C Virus Envelope Protein E2 through the Eukaryotic Initiation Factor 2α Kinase PERK

2003 ◽  
Vol 77 (6) ◽  
pp. 3578-3585 ◽  
Author(s):  
Nicole Pavio ◽  
Patrick R. Romano ◽  
Thomas M. Graczyk ◽  
Stephen M. Feinstone ◽  
Deborah R. Taylor
Keyword(s):  
Endoplasmic Reticulum ◽  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Er Stress ◽  
Envelope Protein ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Virus Envelope ◽  
Expression Levels ◽  
Virus Envelope Protein

ABSTRACT The hepatitis C virus envelope protein, E2, is an endoplasmic reticulum (ER)-bound protein that contains a region of sequence homology with the double-stranded RNA-activated protein kinase PKR and its substrate, the eukaryotic translation initiation factor 2 (eIF2). We previously reported that E2 modulates global translation through inhibition of the interferon-induced antiviral protein PKR through its PKR-eIF2α phosphorylation site homology domain (PePHD). Here we show that the PKR-like ER-resident kinase (PERK) binds to and is also inhibited by E2. At low expression levels, E2 induced ER stress, but at high expression levels, and in vitro, E2 inhibited PERK kinase activity. Mammalian cells that stably express E2 were refractory to the translation-inhibitory effects of ER stress inducers, and E2 relieved general translation inhibition induced by PERK. The PePHD of E2 was required for the rescue of translation that was inhibited by activated PERK, similar to our previous findings with PKR. Here we report the inhibition of a second eIF2α kinase by E2, and these results are consistent with a pseudosubstrate mechanism of inhibition of eIF2α kinases. These findings may also explain how the virus promotes persistent infection by overcoming the cellular ER stress response.

scholarly journals Pancreatic eukaryotic initiation factor-2α kinase (PEK) homologues in humans, Drosophila melanogaster and Caenorhabditis elegans that mediate translational control in response to endoplasmic reticulum stress

2000 ◽  
Vol 346 (2) ◽  
pp. 281-293 ◽  
Author(s):  
Ruchira SOOD ◽  
Amy C. PORTER ◽  
Kun MA ◽  
Lawrence A. QUILLIAM ◽  
Ronald C. WEK
Keyword(s):  
Endoplasmic Reticulum ◽  
Drosophila Melanogaster ◽  
Caenorhabditis Elegans ◽  
Er Stress ◽  
Translational Control ◽  
Mammalian Cells ◽  
Transmembrane Protein ◽  
Initiation Factor ◽  
Eukaryotic Initiation Factor ◽  
Α Subunit

In response to different cellular stresses, a family of protein kinases regulates translation by phosphorylation of the α subunit of eukaryotic initiation factor-2 (eIF-2α). Recently, we identified a new family member, pancreatic eIF-2α kinase (PEK) from rat pancreas. PEK, also referred to as RNA-dependent protein kinase (PKR)-like endoplasmic reticulum (ER) kinase (PERK) is a transmembrane protein implicated in translational control in response to stresses that impair protein folding in the ER. In this study, we identified and characterized PEK homologues from humans, Drosophila melanogaster and Caenorhabditis elegans. Expression of human PEK mRNA was found in over 50 different tissues examined, with highest levels in secretory tissues. In mammalian cells subjected to ER stress, we found that elevated eIF-2α phosphorylation was coincident with increased PEK autophosphorylation and eIF-2α kinase activity. Activation of PEK was abolished by deletion of PEK N-terminal sequences located in the ER lumen. To address the role of C. elegans PEK in translational control, we expressed this kinase in yeast and found that it inhibits growth by hyperphosphorylation of eIF-2α and inhibition of eIF-2B. Furthermore, we found that vaccinia virus K3L protein, an inhibitor of the eIF-2α kinase PKR involved in an anti-viral defence pathway, also reduced PEK activity. These results suggest that decreased translation initiation by PEK during ER stress may provide the cell with an opportunity to remedy the folding problem prior to introducing newly synthesized proteins into the secretory pathway.

scholarly journals Human Cytomegalovirus Protein pUL38 Induces ATF4 Expression, Inhibits Persistent JNK Phosphorylation, and Suppresses Endoplasmic Reticulum Stress-Induced Cell Death

2009 ◽  
Vol 83 (8) ◽  
pp. 3463-3474 ◽  
Author(s):  
Baoqin Xuan ◽  
Zhikang Qian ◽  
Emi Torigoi ◽  
Dong Yu
Keyword(s):  
Endoplasmic Reticulum ◽  
Cell Death ◽  
Er Stress ◽  
Human Cytomegalovirus ◽  
Initiation Factor ◽  
Α Subunit ◽  
Unfolded Protein ◽  
Initiation Factor 2 ◽  
The Unfolded Protein Response ◽  
Protein Response

ABSTRACT The endoplasmic reticulum (ER) is a key organelle involved in sensing and responding to stressful conditions, including those resulting from infection of viruses, such as human cytomegalovirus (HCMV). Three signaling pathways collectively termed the unfolded protein response (UPR) are activated to resolve ER stress, but they will also lead to cell death if the stress cannot be alleviated. HCMV is able to modulate the UPR to promote its infection. The specific viral factors involved in such HCMV-mediated modulation, however, were unknown. We previously showed that HCMV protein pUL38 was required to maintain the viability of infected cells, and it blocked cell death induced by thapsigargin. Here, we report that pUL38 is an HCMV-encoded regulator to modulate the UPR. In infection, pUL38 allowed HCMV to upregulate phosphorylation of PKR-like ER kinase (PERK) and the α subunit of eukaryotic initiation factor 2 (eIF-2α), as well as induce robust accumulation of activating transcriptional factor 4 (ATF4), key components of the PERK pathway. pUL38 also allowed the virus to suppress persistent phosphorylation of c-Jun N-terminal kinase (JNK), which was induced by the inositol-requiring enzyme 1 pathway. In isolation, pUL38 overexpression elevated eIF-2α phosphorylation, induced ATF4 accumulation, limited JNK phosphorylation, and suppressed cell death induced by both thapsigargin and tunicamycin, two drugs that induce ER stress by different mechanisms. Importantly, ATF4 overexpression and JNK inhibition significantly reduced cell death in pUL38-deficient virus infection. Thus, pUL38 targets ATF4 expression and JNK activation, and this activity appears to be critical for protecting cells from ER stress induced by HCMV infection.

scholarly journals Effects of PERK eIF2α Kinase Inhibitor against Toxoplasma gondii

2018 ◽  
Vol 62 (11) ◽  
Author(s):  
Leonardo Augusto ◽  
Jennifer Martynowicz ◽  
Kirk A. Staschke ◽  
Ronald C. Wek ◽  
William J. Sullivan
Keyword(s):  
Endoplasmic Reticulum ◽  
Toxoplasma Gondii ◽  
Er Stress ◽  
Lethal Dose ◽  
Lytic Cycle ◽  
Host Cells ◽  
Initiation Factor ◽  
Α Subunit ◽  
Content Type ◽  
Eif2α Kinase

ABSTRACT Toxoplasma gondii is an obligate intracellular parasite that has infected one-third of the population. Upon infection of warm-blooded vertebrates, the replicating form of the parasite (tachyzoite) converts into a latent form (bradyzoite) present in tissue cysts. During immune deficiency, bradyzoites can reconvert into tachyzoites and cause life-threatening toxoplasmosis. We previously reported that translational control through phosphorylation of the α subunit of T. gondii eukaryotic initiation factor 2 (eIF2α) (TgIF2α) is a critical component of the parasite stress response. Diverse stresses can induce the conversion of tachyzoites to bradyzoites, including those disrupting the parasite's endoplasmic reticulum (ER) (ER stress). Toxoplasma possesses four eIF2α kinases, one of which (TgIF2K-A) localizes to the parasite ER analogously to protein kinase R-like endoplasmic reticulum kinase (PERK), the eIF2α kinase that responds to ER stress in mammalian cells. Here, we investigated the effects of a PERK inhibitor (PERKi) on Toxoplasma. Our results show that the PERKi GSK2606414 blocks the enzymatic activity of TgIF2K-A and reduces TgIF2α phosphorylation specifically in response to ER stress. PERKi also significantly impeded multiple steps of the tachyzoite lytic cycle and sharply lowered the frequency of bradyzoite differentiation in vitro. Pretreatment of host cells with PERKi prior to infection did not affect parasite infectivity, and PERKi still impaired parasite replication in host cells lacking PERK. In mice, PERKi conferred modest protection from a lethal dose of Toxoplasma. Our findings represent the first pharmacological evidence supporting TgIF2K-A as an attractive new target for the treatment of toxoplasmosis.

scholarly journals Regulation of mRNA Translation and Cellular Signaling by Hepatitis C Virus Nonstructural Protein NS5A

2001 ◽  
Vol 75 (11) ◽  
pp. 5090-5098 ◽  
Author(s):  
Yupeng He ◽  
Seng-Lai Tan ◽  
Semih U. Tareen ◽  
Sangeetha Vijaysri ◽  
Jeffrey O. Langland ◽  
...  
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Protein Kinase ◽  
Virus Infection ◽  
Nonstructural Protein ◽  
Mrna Translation ◽  
Mitogen Activated Protein Kinase ◽  
Initiation Factor ◽  
Translation Rate ◽  
Α Subunit

ABSTRACT The NS5A nonstructural protein of hepatitis C virus (HCV) has been shown to inhibit the cellular interferon (IFN)-induced protein kinase R (PKR). PKR mediates the host IFN-induced antiviral response at least in part by inhibiting mRNA translation initiation through phosphorylation of the α subunit of eukaryotic initiation factor 2 (eIF2α). We thus examined the effect of NS5A inhibition of PKR on mRNA translation within the context of virus infection by using a recombinant vaccinia virus (VV)-based assay. The VV E3L protein is a potent inhibitor of PKR. Accordingly, infection of IFN-pretreated HeLa S3 cells with an E3L-deficient VV (VVΔE3L) resulted in increased phosphorylation levels of both PKR and eIF2α. IFN-pretreated cells infected with VV in which the E3L locus was replaced with theNS5A gene (VVNS5A) displayed diminished phosphorylation of PKR and eIF2α in a transient manner. We also observed an increase in activation of p38 mitogen-activated protein kinase in IFN-pretreated cells infected with VVΔE3L, consistent with reports that p38 lies downstream of the PKR pathway. Furthermore, these cells exhibited increased phosphorylation of the cap-binding initiation factor 4E (eIF4E), which is downstream of the p38 pathway. Importantly, these effects were reduced in cells infected with VVNS5A. NS5A was also found to inhibit activation of the p38-eIF4E pathway in epidermal growth factor-treated cells stably expressing NS5A. NS5A-induced inhibition of eIF2α and eIF4E phosphorylation may exert counteracting effects on mRNA translation. Indeed, IFN-pretreated cells infected with VVNS5A exhibited a partial and transient restoration of cellular and viral mRNA translation compared with IFN-pretreated cells infected with VVΔE3L. Taken together, these results support the role of NS5A as a PKR inhibitor and suggest a potential mechanism by which HCV might maintain global mRNA translation rate during early virus infection while favoring cap-independent translation of HCV mRNA during late infection.

Norepinephrine induces endoplasmic reticulum stress and downregulation of norepinephrine transporter density in PC12 cells via oxidative stress

2005 ◽  
Vol 288 (5) ◽  
pp. H2381-H2389 ◽  
Author(s):  
Weike Mao ◽  
Chikao Iwai ◽  
Fuzhong Qin ◽  
Chang-seng Liang
Keyword(s):  
Endoplasmic Reticulum ◽  
Er Stress ◽  
Pc12 Cells ◽  
Initiation Factor ◽  
Α Subunit ◽  
Homologous Protein ◽  
Initiation Factor 2 ◽  
Stress Signals ◽  
Novel Therapeutic Strategies ◽  
Factor C

Cardiac norepinephrine (NE) uptake is reduced in cardiomyopathy. This change is associated with a decrease of NE transporter (NET) receptor and can be reproduced in PC12 cells by extracellular NE. To study whether this effect of NE is mediated via impaired glycosylation and trafficking of NET in the endoplasmic reticulum (ER), we measured the distribution of glycosylated 80-kDa NET and unglycosylated 46-kDa NET in the membrane and cytosolic fractions of PC12 cells. We found that NE decreased glycosylated NET in both membrane and cytosolic fractions and increased cytosolic unglycosylated NET protein. Similar results were produced by tunicamycin and thapsigargin, two agents that induce ER stress by inhibiting N-glycosylation of membrane proteins and disrupting calcium homeostasis, respectively. Also, like the ER stressors, NE not only increased phosphorylation of both the α-subunit of eukaryotic initiation factor-2 and its upstream RNA-dependent protein kinase-like ER kinase over 12 h of treatment but also increased ER chaperone molecule glucose-regulated protein 78 and the nuclear transcription factor C/EBP homologous protein. Antioxidants superoxide dismutase and catalase prevented the downregulation of NET proteins and induction of ER stress signals produced by NE but not by tunicamycin or thapsigargin. The results indicate that the downregulation of membrane NET by NE is mediated by decreased N-glycosylation of NET proteins secondary to induction of ER stress pathways by NE-derived oxidative metabolites. Interventions involving the ER stress pathways may provide novel therapeutic strategies for the treatment of sympathetic dysfunction in heart failure.

scholarly journals NEFA Promotes Autophagosome Formation through Modulating PERK Signaling Pathway in Bovine Hepatocytes

Animals ◽  
2021 ◽  
Vol 11 (12) ◽  
pp. 3400
Author(s):  
Yan Huang ◽  
Chenxu Zhao ◽  
Yaoquan Liu ◽  
Yezi Kong ◽  
Panpan Tan ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Er Stress ◽  
Signaling Pathway ◽  
Lipid Accumulation ◽  
Metabolic Stress ◽  
Perinatal Period ◽  
High Plasma ◽  
Negative Energy ◽  
Initiation Factor ◽  
Sequestosome 1

During the perinatal period, the abnormally high plasma non-esterified fatty acids (NEFA) concentration caused by the negative energy balance (NEB) can impose a significant metabolic stress on the liver of dairy cows. Endoplasmic reticulum (ER) stress is an important adaptive response that can serve to maintain cell homeostasis in the event of stress. The protein kinase R-like endoplasmic reticulum kinase (PERK) pathway is the most rapidly activated cascade when ER stress occurs in cells and has an important impact on the regulation of hepatic lipid metabolism and autophagy modulation. However, it is unknown whether NEFA can affect autophagy through modulating the PERK pathway, under NEB conditions. In this study, we provide evidence that NEFA treatment markedly increased lipid accumulation, the phosphorylation level of PERK and eukaryotic initiation factor 2α (eIF2α), and the expression of glucose-regulated protein 78 (Grp78), activating transcription factor 4 (ATF4), and C/EBP homologous protein (CHOP). More importantly, NEFA treatment can cause a substantial increase in the protein levels of autophagy-related gene 7 (ATG7), Beclin-1 (BECN1), sequestosome-1 (p62), and microtubule-associated protein 1 light chain 3 (LC3)-II, and in the number of autophagosomes in primary bovine hepatocytes. The addition of GSK2656157 (PERK phosphorylation inhibitor) can significantly inhibit the effect of NEFA on autophagy and can further increase lipid accumulation. Overall, our results indicate that NEFA could promote autophagy via the PERK pathway in bovine hepatocytes. These findings provide novel evidence about the potential role of the PERK signaling pathway in maintaining bovine hepatocyte homeostasis.

2,4,6-Trichlorophenol Cytotoxicity Involves Oxidative Stress, Endoplasmic Reticulum Stress, and Apoptosis

2014 ◽  
Vol 33 (6) ◽  
pp. 532-541 ◽  
Author(s):  
Xiaoning Zhang ◽  
Xiaona Zhang ◽  
Zhidan Niu ◽  
Yongmei Qi ◽  
Dejun Huang ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Endoplasmic Reticulum ◽  
Er Stress ◽  
Messenger Rna ◽  
Nuclear Translocation ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Heme Oxygenase 1 ◽  
Related Factor ◽  
Α Subunit

This study aims to evaluate the cytotoxicity and potential mechanisms of 2,4,6-trichlorophenol (2,4,6-TCP) in mouse embryonic fibroblasts. Our results show that 2,4,6-TCP causes morphological changes and reduces cell viability. The overproduction of reactive oxygen species, the upregulation of nuclear factor-E2-related factor 2 (Nrf2) and heme oxygenase 1 (HMOX1) messenger RNA (mRNA) expressions, and the nuclear translocation of Nrf2 protein demonstrate that 2,4,6-TCP induces oxidative stress, and the Nrf2/HMOX1 pathway might be involved in 2,4,6-TCP-induced antioxidative response. Simultaneously, our data also demonstrate that 2,4,6-TCP upregulates the expressions of binding immunoglobulin protein, inositol-requiring enzyme/endonuclease 1α, and C/EBP homologous protein; stimulates α subunit of eukaryotic translation initiation factor 2 phosphorylation; and induces the splicing of Xbp1 mRNA, suggesting that endoplasmic reticulum (ER) stress is triggered. Moreover, 2,4,6-TCP alters the mitochondrial membrane potential and increases the apoptosis rate, the caspase 3 activity, and the Bax/Bcl-2 ratio, demonstrating that the mitochondrial pathway is involved in the 2,4,6-TCP-induced apoptosis. Thus, these results show that 2,4,6-TCP induces oxidative stress, ER stress, and apoptosis, which together contribute to its cytotoxicity in vitro.

Sign in / Sign up

Export Citation Format

Share Document